Interdiab

© Filodiritto Editore – Proceedings
PROCEEDINGS
OF
6th International Conference on

Interdisciplinary Management of
Diabetes Mellitus and its Complications
INTERDIAB
(Bucharest, Romania, 5-7 March 2020)
Editors
Cristian SERAFINCEANU and Anca PANTEA STOIAN
FILODIRITTO
INTERNATIONAL PROCEEDINGS
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First Edition February 2020
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INDEX
Foreword 10
Early Detection of Left Atrial Dysfunction by Speckle Tracking Echocardiography

in Asymptomatic Subjects with Type 1 Diabetes Mellitus 14
NEAGOE Oana, MIREA Oana, DONOIU Ionuț, BERCEANU Mihaela,
ISTRATOAIE Octavian, MOTA Maria, MILITARU Constantin
Evaluation of the Nutritional Status – Decision Factor in the Therapeutic Approach

in Senior Patient with Hip Fracture 20
PÎSLARU Anca Iuliana, ILIE Adina Carmen, SANDU Ioana Alexandra,
ȘTEFĂNIU Ramona, ALEXA Ioana Dana, ALEXA Ovidiu, PÎNZARU Roxana
Efficacy and Safety of Adding Sitagliptin in Patients with Type 2 Diabetes Mellitus
with Inadequate Glycemic Control in Metformin Monotherapy 26
ALBAI Oana, TIMAR Bogdan, TIMAR Romulus
Is in Vitro Fertilisation Pregnancy Different to Natural Pregnancy Concerning

Gestational Diabetes Risk? A Short Review 34
MATEI Alexandra, IONESCU Cringu Antoniu, GHEORGHIU Diana,
CATRINA Eduard Lucian, AL AZAWI Alla, ROSU George-Alexandru,
ZYGOUROPOULOS Nikolaos, CALIN Florin Daniel, PANTEA STOIAN Anca,
NAVOLAN Dan, PACU Irina
Association between Helicobacter Pylori Infection, Obesity and Gestational Diabetes 38

CODREANU Ana-Maria, MATEI Alexandra, BORONTEA Minodora,
OLARU Luciana, OLARU Flavius
The Relationship Between Oral Health and Quality of Life in Romanian

Diabetic Patients Hospitalised in a Tertiary Antidiabetic Center 44
PANTEA STOIAN Anca, STOICA Roxana Adriana, ȘTEFAN Simona Diana,
PITURU Silviu, ISTRATI Ionela, SERAFINCEANU Cristian
Metformin Treatment During Pregnancy and Its Effect on Fetal Growth:

A Literature Review 51
IONESCU Crîngu Antoniu, ROȘU George-Alexandru,
ZYGOUROPOULOS Nikolaos, MĂDAN Victor, SOCEA Bogdan,
JACOTĂ-ALEXE Florentina, GHEORGHIU Diana Claudia,
GHEORGHIU Nicolae, HARADJA Horațiu, CAIMACAN Adriana,
POPESCU (BĂNACU) Ina, BĂNACU Mihail, CĂLIN Florin Daniel,
DIMITRIU Mihai
Plant-Based Diets and Type 2 Diabetes Prevention and Management 57

HANCU Anca Mihaela, SERAFINCEANU Cristian, RIZZO Manfredi,
PAPANAS Nikolaos, PANTEA STOIAN Anca
3
Male Infertility, Metabolic Syndrome and Other Related Disorders 66

PACU Irina, IONESCU Crîngu Antoniu, PACU Ovidiu, DIMITRIU Mihai,
BĂNACU Mihail, SOCEA Bogdan, MĂDAN Victor, HANGANU Irina,
ZYGOUROPOULOS Nikolaos
Uncontrolled Diabetes Mellitus – Negative Predictive Factor for Colorectal

Cancer Recurrence after Curative Surgical Treatment 73
SUCEVEANU Andra-Iulia, SUCEVEANU Adrian Paul, PAREPA Irinel,
ARDELEANU Valeriu, MICU Ioan Sergiu, DUMITRU Andrada,
CATRINOIU Doina, STANCIU Adina Elena, NITIPIR Cornelia,
MAZILU Laura
Assessment of Physical Activity in the Elderly an Element in Maintaining

Vascular Resilience? 80
AURELIAN Sorina Maria, MIHALACHE Ruxandra, ZAMFIRESCU Andreea,
AURELIAN Justin, CAPISIZU Ana
Chronic Kidney Disease and Related Conditions in a Roma Population with

Diabetes Mellitus 86
RUSU Emilia, COSOREANU Andrada, BALEANU Maria, MARINESCU Mihai,
ENACHE Georgiana, RUSU Florin, BEJINARIU Catalina, JINGA Mariana,
RADULIAN Gabriela
The Metabolic Syndrome – Clinical and Biochemical Correlations 92

PASARICA Ilinca, PERTEA Leonard-Iosif, VLASE Alexandru,
BARBU Roxana-Mihaela, MUNTEANU Dragos, CERNOMAZ Andrei
Computational Approach for Small Data in Animal Models with Induced

Metabolic Disorders 98
GHICA Manuela, BĂNCESCU Irina, UDEANU Denisa Ioana,
TĂEREL Adriana, ARSENE Andreea Letiția, ANUȚA Valentina,
VELESCU Bruno Ștefan, GERGHICEANU Florentina, MITITELU Magdalena,
IONIȚĂ Corina Ana
The Relationship between Novel Haematological Markers and Microvascular

Complications among Patients with Diabetes in Sibiu County 107
SITTERLI-NATEA Carmen-Narcisa, CALUTIU Nicoleta-Georgiana
The Oral Antidiabetic Treatment in Patients with Type 2 Diabetes Mellitus

and Peripheral Artery Disease 113
DIACONU Camelia Cristina, HORODINSCHI Ruxandra-Nicoleta,
BRATU Ovidiu Gabriel, BACALBASA Nicolae, ILIESCU Laura,
STANESCU Ana Maria Alexandra
Vascular Calcification in Diabetes Mellitus Type2 – Molecular Mechanisms

and Clinical Implications 118
CASOINIC Florin, BUZOIANU Anca, HANCU Nicolae
4
Diabetes Mellitus: Unforgettable Became Inevitable – a Case Report with

a 2-Years Follow-Up 126
CHELARU Sorina-Alina, CATRINOIU Doina, NEAMŢU Mirela, ION Ileana
Spectrum of Bacteria Associated with Diabetic Foot Syndrome

and Their Susceptibility to Antibiotics 130
ZAHA Dana Carmen, POPA Amorin Remus, JURCA Alexandru,
POPA Loredana, POPOVICIU Mihaela, FERICIAN Anca, DAINA Cristian Marius,
MAGHIAR Octavian, VESA Cosmin Mihai
Cardiovascular Risk Assessment in a Group of Non-alcoholic Fatty Liver

Patients from Transylvania 135
DASCĂLU Daciana Nicoleta
Evolution Particularities and Prognostic Significance of New-Onset Atrial Fibrillation

in a Group of Diabetic Female Patients with Acute ST-Elevation Myocardial Infarction
Treated by Primary Percutaneous Transluminal Coronary Angioplasty 139
MACOVEI Liviu, POPA Delia Melania, TRINCA Cristina,
ADOAMNEI Dumitru Marius, NASTASA Dan George, LOVIN Nicusor,
BAZYANI Amin, ACHITEI Ionut Silviu, BURLACU Alexandru, NEDELCIUC Igor
The Importance of Physical Exercise for the Correction of Spinal Deviations

in Children with Type 1 Diabetes 145
AMARICAI Elena
Type 2 Diabetes Mellitus and the Risk of Hepatocellular Carcinoma in Chronic

Hepatitis C Patients Treated with Direct Acting Antivirals 148
ILIESCU Elena Laura, TOMA Letitia, DIACONU Camelia, ZGURA Anca,
BACALBASA Nicolae, MERCAN-STANCIU Adriana
Novel Therapeutics Aspects in Advanced and Metastatic Non-small Cell Lung

Cancer – Experience of Oncology Department of Chronic Disease
St. Luke’s Hospital Bucharest 154
RAHNEA-NITA Gabriela, CIUHU Anda-Natalia,
RAHNEA-NITA Roxana-Andreea, STOIAN Alexandru-Rares
Palliative Treatment of Severe Dysphagia in Advanced Cancer Patients 161

RAHNEA-NITA Gabriela, CIUHU Anda-Natalia,
RAHNEA-NITA Roxana-Andreea, STOIAN Alexandru-Rares
The Impact of Chemotherapy on the Quality of Life in Patients

with Gynaecological Cancer 167
RAHNEA-NITA Roxana-Andreea, RAHNEA-NITA Gabriela,
ANGHEL Rodica Maricel
Hypertension and Diabetes Mellitus throughout History 177

GAIȚĂ Laura-Adriana, TIMAR Romulus Zorin, TIMAR Bogdan
5
Pharmacoepidemiologic Implications at Patients with Metabolic Syndrome 183

GĂNCEANU-RUSU Ana-Roxana, LUPUŞORU Elena-Cătălina,
DIMA Nicoleta, MĂRĂNDUCĂ Minela-Aida, CLIM Andreea, POP Ana-Maria,
BĂDESCU Minerva-Codruţa, TĂNASE Daniela-Maria, OUATU Anca,
REZUŞ Ciprian
Natural Killer Cells in Type 1 Diabetes – Phenotypic Characteristics 189

ISVORANU Gheorghiţa, MANOLE Emilia, SURCEL Mihaela,
MUNTEANU Adriana Narcisa, NEAGU Monica Teodora
Risk of Hypoglycaemia in Patients with Stage Three of Chronic Kidney Disease

and Type 2 Diabetes Mellitus Treated with Different Classes
of Oral Antidiabetic Drugs 195
GHERBON Adriana, FRANDES Mirela
The Impact of Metabolic Syndrome on Pregnancy 203

POPESCU (BANACU) Ina, PACU Irina, IONESCU Cringu Antoniu,
MĂDAN Victor, SOCEA Bogdan, DIMITRIU Mihai, BACALBASA Nicolae,
POPESCU Mihail, ZYGOUROPOULOS Nikolaos, DAN Adelina,
BANACU Mihail
Serum Adipokines in Obese Bariatric Patients – Correlations

with Metabolic Parameters 207
HRISTOV Ioana, TIMOFTE Daniel Vasile, CREȚU-SILIVESTRU Iustina Silvia,
MIHAI Bogdan, ANISIE Ecaterina, MOCANU Veronica
Insulin Resistance and Male Infertility 214

PACU Irina, GHEORGHIU Diana, MĂDAN Victor, CIOBANU Ana Maria,
DAN Adelina-Loredana, ZYGOUROPOULOS Nikolaos, MATEI Alexandra,
ROȘU George-Alexandru, PACU Ovidiu, NAVOLAN Dan, CĂLIN Florin Daniel
Oral Mucosa Vascularization in Early Diabetic Diagnosis 218

VYSOCHANSKAYA Julia, YEROSHENKO Galina, RYBALOV Oleg
Clinical and Pathological Characteristics of Patients with Type 2 Diabetes

and Early Stage Breast Cancer – a Single Centre Experience 224
MAZILU Laura, SUCEVEANU Adrian-Paul, PAREPA Irinel-Raluca,
GHEORGHE Andreea Daniela, PANTEA STOIAN Anca, HAINAROSIE Razvan,
STEFANESCU Dragos Cristian, NITIPIR Cornelia, ARDELEANU Valeriu,
SUCEVEANU Andra Iulia
Clinical and Pathological Features of Patients with Colorectal Cancer and Diabetes
– a Single Centre Experience 230
MAZILU Laura, SUCEVEANU Adrian-Paul, PAREPA Irinel Raluca,
GHEORGHE Andreea Daniela, PANTEA STOIAN Anca, ARDELEANU Valeriu,
SUCEVEANU Andra Iulia
6
Statins and Risk Biomarkers for Coronary Heart Disease (CHD) in Diabetes
Mellitus Patients with Normal LDL Cholesterol Levels. 1 Year Prospective Study
in Constanta County, Romania 235
PAREPA Irinel Raluca, SUCEVEANU Andra Iulia, PANTEA-STOIAN Anca,
MAZILU Laura, CATRINOIU Doina, IONESCU Paris, IONESCU Mihaela
The Relationship between Arterial Stiffness and 25-OH Vitamin D3 in Type 2

Diabetes Mellitus Patients 240
IURCIUC Stela, TUDOR Anca, RADA Maria, BADALICA-PETRESCU Marius,
PAH Ana, STOICHESCU-HOGEA Gheorghe, CRACIUN Laura,
IURCIUC Mircea, ORAVITAN Mihaela
Maternal Obesity: A New Challenge of Modern-Day Medicine – Case Report 245

DIMITRIU Mihai, NICULAE Mihai-Bogdan, GHEORGHIU Diana-Claudia,
HARADJA Horatiu, CIRSTOVEANU Catalin Gabriel, DAN Adelina-Loredana,
IONESCU Andra-Maria
Coping Strategies, Illness Perceptions and Optimism: Contribution

to Self-Management and Adherence in Diabetic Patients 250
POPA-VELEA Ovidiu, APOSTOL Alina Florina, DIACONESCU Liliana Veronica
Cause or Complication? Psychosocial Aspects of Diabetic Patients

with Cardiovascular Diseases 256
PÁL Tünde, PREG Zoltán, SZABÓ Monica Iudita Maria, NYULAS Kinga,
BÁLINT SZENTENDREY Dalma, GERMÁN-SALLÓ Márta
Mild Cognitive Impairment Is More Frequent in Middle Aged Diabetics

and Females 263
PÁL Tünde, GERMÁN-SALLÓ Márta, SZABÓ Monica Iudita Maria,
NEMES-NAGY Enikő, NYULAS Kinga, BÁLINT SZENTENDREY Dalma,
PREG Zoltán
The Prevalence and Characteristics of Depression in Diabetes Mellitus Patients.

Is Routine Screening of Depression Necessary in The DM Patients? 268
LEȚI Maria-Mădălina, BODNĂRESCU-COBANOGLU Mihaela,
DOBRESCU Iuliana, POP Anca Lucia, POPA Denisa, ROSU Isabella Andreea,
IORGUT Cristina, ZETU Cornelia
Diabetes Mellitus and Pregnancy 278

RADU Mihaela Corina, BOERU Adrian Calin, MARIN Mihaela Liliana,
MANOLESCU Loredana Sabina Cornelia
Incidence and Severity of Sudomotor Dysfunction in Early Stages of Sensorimotor

Neuropathy in Type 2 Diabetes 286
DOROS Rodica, STEGARU Daniela, IACOBINI Andra Evelyn, MOTOC Razvan,
BRAILEANU Bogdana, ANDREI Catalina, CACEAUNE Elena, ELIAN Viviana,
ZETU Cornelia
7
Hypercortisolism Is Associated with Increased Cardiovascular Mortality

and Morbidity: A Retrospective Study 293
DUMITRIU Roxana, DUȘCEAC Roxana, SALMEN Teodor,
POIANĂ Cătălina
Heart Failure in Patients with Diabetes Mellitus: Paraclinical Findings 298

SALMEN Teodor, STEGARU Daniela, PIETROSEL Valeria-Anca,
ZARZU Alexandra, ALEXE Vlad, MILICESCU Alexandra, DOROS Rodica,
MIHAI Andrada
Glucose Control in Patients with Acromegaly Resistant to First-Line

Somatostatin Receptor Analogues Monotherapy 304
GĂLOIU Simona, MĂRGĂRIT Emma, BACIU Ionela, NICULESCU Dan,
TRIFĂNESCU Raluca, RADIAN Șerban, CĂPĂȚÎNĂ Cristina,
CARAGHEORGHEOPOL Andra, POIANĂ Cătălina
Physical Activity Level and Patterns of Patients with Type 2 Diabetes 309
SZABO Monica, MÁTÉ Beáta, BALOGH Rozália, GERMÁN SALLÓ Márta
Fast-food and Diabetes in Paediatrics – A Road to Disaster 316

MIHAI Cristina Maria, PINZARU Anca Daniela, CHISNOIU Tatiana
Chemical-Toxicological Research on the Evaluation of Serum Malondialdehyde

Levels in Patients with Diabetic Arteriopathy 320
IONIȚĂ Ana Corina, NICOLESCU Teodor Octavian, IONIȚĂ Elena Iuliana,
NICOLESCU Florica, MITITELU Magdalena
Research on the Hypolipidemic Action of Almond Oil and Almond Seeds 326
IONIȚĂ Ana Corina, MITITELU Magdalena, NICOLESCU Teodor Octavian,
NICOLESCU Florica, MOROȘAN Elena, OZON Emma Adriana,
IONIȚĂ Elena Iuliana
Research on the Influence of Some Sweeteners on Body Weight and Blood

Glucose in Laboratory Mice 332
MITITELU Magdalena, NICOLESCU Teodor Octavian,
UDEANU Denisa Ioana, NICOLESCU Florica
Study of Hypolipemiant Activity of Some Mussel Extracts 338

MITITELU Magdalena, NICOLESCU Teodor Octavian, MOROȘAN Elena,
POP Anca Lucia, NICOLESCU Florica
Comparative Assessment of Continuous Hospitalization and Day Hospitalization

in Patients with Diabetes Mellitus 344
DAINA Lucia Georgeta, VENTER Alina Cristiana, POPA Amorin Remu,
VESA Cosmin Mihai, BUHAȘ Camelia Liana, SABAU Monica,
BONTA Marinela, DAINA Cristian Marius
Impact of Diabetes Mellitus on Hospital Indicators 350

DAINA Cristian Marius, POPA Amorin Remus, VESA Cosmin Mihai,
TIMAR Calin, COTRĂU Petru, DAINA Lucia Georgeta
8
Major Differences Regarding Lipid Profile, Apolipoprotein-B and Insulin

Resistance between Type II Diabetes Patients with Altered and Normal
Glomerular Filtration Rate 355
GHERDAN Violeta Valentina, POPA Amorin Remus, JURCA Alexandru,
POPA Loredana, POPOVICIU Mihaela, VESA Cosmin Mihai, FERICIAN Anca,
DAINA Cristian Marius, ZAHA Dana Carmen
The Prevalence of Cardiovascular Risk Factors Control among Type 2 Diabetes

Mellitus Patients – Epidemiological Study in a Tertiary Care Hospital
from North Western Romania 361
POPOVICIU Mihaela Simona, POPA Loredana, VESA Cosmin Mihai,
MAGHIAR Octavian, ZAHA Andreea Atena, BILUȚĂ Adina, BUNGAU Alexa,
POPESCU Ioachim Mircea, POPA Amorin Remus
Challenges in Treating Erectile Dysfunction in Diabetic Patients – Review 367

MĂDAN Victor, PAVALEAN Mihai, PACU Ovidiu, PAVALEAN Mihaela,
BUCURICA Sandica, DIMITRIU Mihai, PACU Irina
New Data about Urinary Bladder Complications of Diabetes mellitus – A Review 372
MĂDAN Victor, PAVALEAN Mihai, PACU Ovidiu, PAVALEAN Mihaela,
BUCURICA Sandica, DIMITRIU Mihai, PACU Irina
Does Obesity Cause Type 2 Diabetes? 377

MICIC Dragan D., POLOVINA Snezana, MICIC Dusan D.
The Amount of Weight Loss after Metabolic Surgery Matters 380

POLOVINA Snezana, MICIC Dragan D., MICIC Dusan D.
9
FOREWORD
Association for Renal, Metabolic and Nutritional Studies (ASRMN) is a scientific entity
based in Romania that organizes every year (since 2015) the International Conference on
Interdisciplinary Management of Diabetes and its Complications – INTERDIAB.
During its existing, INTERDIAB has become more than just a Conference, as it has been
addressing specialists, researchers, experts in the field of diabetes mellitus and connected topics
(Romanian and international), bringing to the fore original research as a means of developing
and knowledge.
And so was born the Proceedings INTERDIAB Volume, promoting the original works of
young researchers and also creating a forum for discussion and debate between specialists
involved in the complex care of the patients with diabetes mellitus: diabetologists,
endocrinologists, cardiologists, nephrologists, gastroenterologists, surgeons, oncologists,
neurologists and many others.
Since its first edition, the original research articles published in Proceedings INTERDIAB
Volume have received positive opinion from Thomson Reuters, so they were indexed in Web
of Science (2015, 2016 and 2017 editions). The 2018 and 2019 editions are in full process of
being indexed by Clarivate Analytics.
Each year, a new dimension of interdisciplinarity in the diagnosis and management of
diabetes mellitus is being brought into discussion at INTERDIAB. The 2015 and 2016 editions
were devoted entirely to interdisciplinary approaches in diabetic chronic kidney disease while
in 2017 it focused on diabetes mellitus as cardiovascular disease and in 2018 on surgery and
diabetes: at crossroads.
This year, INTERDIAB has reached its sixth edition and, just as in 2019, is embracing the
same topic: diabetes mellitus in internal medicine, as the general acceptance acknowledges
this topic as best defining INTERDIAB essence/mark. Given the topic generous
interdisciplinary framework, we’ve managed to gather in this volume 60 articles of original
research.
But selecting the best articles is never an easy duty. Each article we receive is being sent to
the peer review process to make sure every single one is being seen by at least two reviewers
whose area of expertise cover the article topic. Each reviewer releases a review report for each
article they see and at the end of the reviewing process, the Scientific Committee gather to
analyze all review reports. We use a standard type of review report used by other prestigious
international publications as well. What is to be mentioned is that in almost 100% cases,
reviewers’ opinion concurs, even if they belong to different professional environments and/or
from different countries. This is a very good sign for us that the articles are being reviewed
properly, that every article is being fully read. Each article is being reviewed in blind, so that
the authors names and/or their affiliation won’t influence the review’s result.
During the review process, it happens sometimes that the reviewers to ask the authors to
come back with clarifications and/or completions to their articles. Thus, we make sure the
articles’ scientific quality is being improved and sometimes it happens that even if after the first
review an article not to meet the publishing criteria, but, supported by the reviewers’ comments,
the author to improve the article’s scientific quality and in the end to get published.
The reviewing process is being handled by an impartial agency that is also taking care of the
INTERDIAB Conference management, to make sure no conflict of interest arises between
members of the Scientific Committee and articles authors.
At the end of the reviewing process, after analyzing all review reports, the Scientific
Committee decide which articles meet the criteria for Web of Science and are being sent to be
10
indexed. Some of these articles that best meet the INTERDIAB Conference scientific needs are
even proposed to be held as oral presentations within the Conference.
Hopefully, year by year we manage to invite more and more well-known Romanian and
international opinion leaders to be part of the Scientific Committee and thus participate as
reviewers and, year by year, as the scientific rigor is increasing, the articles quality also
increases.
We are confident that the original research performed by multiple groups in different
specialties is the best way to understand the extremely complex nature of the metabolic
pathology associated to diabetes mellitus. Our wish is to keep alive INTERDIAB tradition of
being a leader in promoting interdisciplinary research work, as there are only few scientific
manifestations of its kind in this part of Europe.
Professor Cristian Serafinceanu – Carol Davila University of Medicine and Pharmacy,

Bucharest – Romania
President of INTERDIAB Conference
President of Association for Renal, Metabolic and Nutritional Studies (ASRMN)
11
INTERDIAB 2020 VOLUME EDITORS

- Prof. Cristian SERAFINCEANU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Anca PANTEA STOIAN – Carol Davila University of Medicine and
Pharmacy, Bucharest, Romania
SCIENTIFIC COMMITTEE – ARTICLES’ REVIEWERS

- Prof. Antonio CERIELLO – University of Udine, Italy
- Prof. Francesco COSENTINO – Karolinska University Hospital, Stockholm, Sweden
- Prof. Andrej JANEZ – Ljubljana University Medical Centre, Slovenia
- Prof. Gyorgy JERMENDY – Bajcsy-Zsilinszky Hospital, Budapest, Hungary
- Prof. Katarina LALIC – Faculty of Medicine, University of Belgrade, Serbia
- Prof. Nebojsa LALIC – Faculty of Medicine, University of Belgrade, Serbia
- Prof. Dragan MICIC – Faculty of Medicine, University of Belgrade, Serbia
- Prof. Dimitri MIKHAILIDIS – University College London (UCL), UK
- Prof. Snezana POLOVINA – Research Associate at School of Medicine, University of
Belgrade, Serbia
- Prof. Manfredi RIZZO – University of Palermo, Italy
- Prof. Peter TOTH – University of Illinois, College of Medicine, Peoria, USA
- Prof. Istvan WITTMANN – University of Pécs, Hungary
- Assoc. Prof. Yajnavalka BANERJEE – College of Medicine, Dubai, UAE
- Assoc. Prof. Nikolaos PAPANAS – Democritus University of Thrace, Greece
- Dr. Niki KATSIKI – Aristotle University of Thessaloniki, Greece
- Dr. Iuliana POPESCU – University of Kentucky, Lexington, USA
- Dr. Julia VYSOCHANSKAYA – Odontologia Somatica LTD, Russia
- Acad. Maya SIMIONESCU – Romanian Academy
- Prof. Doina CATRINOIU – Ovidius University, Faculty of Medicine and Pharmacy,
Constanţa, Romania
- Prof. Adrian COVIC – Grigore T. Popa University of Medicine and Pharmacy, Iaşi,
Romania
- Prof. Antoniu Crîngu IONESCU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Prof. Dan DUMITRAŞCU – Iuliu Haţieganu University of Medicine and Pharmacy,
Cluj-Napoca, Romania
- Prof. Carmen FIERBINŢEANU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Prof. Dan GAIŢĂ – Victor Babeş University of Medicine and Pharmacy, Timişoara,
Romania
- Prof. Nicolae HÂNCU – Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-
Napoca, Romania
- Prof. Daniel LIGHEZAN – Victor Babeş University of Medicine and Pharmacy,
Timişoara, Romania
- Prof. Eugen MOŢA – University of Medicine and Pharmacy, Craiova, Romania
- Prof. Maria MOŢA – University of Medicine and Pharmacy, Craiova, Romania
12
- Prof. Antoniu PETRIŞ – Grigore T. Popa University of Medicine and Pharmacy, Iaşi,
Romania
- Prof. Cătălina POIANĂ – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Prof. Gabriela ROMAN – Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-
Napoca, Romania
- Prof. Cristian SERAFINCEANU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Prof. Romulus TIMAR – Victor Babeş University of Medicine and Pharmacy,
Timişoara, Romania
- Prof. Liliana TUŢĂ – Ovidius University, Faculty of Medicine and Pharmacy,
Constanţa, Romania
- Prof. Dragoş VINEREANU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Cornelia BALA – Iuliu Haţieganu University of Medicine and Pharmacy,
Cluj-Napoca, Romania
- Assoc. Prof. Anca CERGHIZAN – Iuliu Haţieganu University of Medicine and
Pharmacy, Cluj-Napoca, Romania
- Assoc. Prof. Camelia DIACONU – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Liliana GÂRNEAŢĂ – Carol Davila University of Medicine and
- Assoc. Prof. Cristian GUJA – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Cristina LĂCĂTUŞU – Grigore T. Popa University of Medicine and
Pharmacy, Iaşi, Romania
- Assoc. Prof. Mircea MANUC – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Bogdan MIHAI – Grigore T. Popa University of Medicine and Pharmacy,
Iaşi, Romania
- Assoc. Prof. Anca PANTEA-STOIAN – Carol Davila University of Medicine and
- Assoc. Prof. Diana PĂUN – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Corina POP – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Assoc. Prof. Simona POPA, – University of Medicine and Pharmacy, Craiova, Romania
- Assoc. Prof. Bogdan TIMAR – Victor Babeş University of Medicine and Pharmacy,
Timişoara, Romania
- Assoc. Prof. Ioan Andrei VEREŞIU – Iuliu Haţieganu University of Medicine and
Pharmacy, Cluj-Napoca, Romania
- Assoc. Prof. Adrian VLAD – Victor Babeş University of Medicine and Pharmacy,
Timişoara, Romania
- Lect. Viviana ELIAN – Carol Davila University of Medicine and Pharmacy, Bucharest,
Romania
- Lect. Răzvan HAINĂROŞIE – Carol Davila University of Medicine and Pharmacy,
Bucharest, Romania
- Lect. Andrada MIHAI – Carol Davila University of Medicine and Pharmacy, Bucharest,
Romania
13
Early Detection of Left Atrial Dysfunction by Speckle Tracking

Echocardiography in Asymptomatic Subjects with Type 1
Diabetes Mellitus
*NEAGOE Oana1, MIREA Oana1, DONOIU Ionuț1, BERCEANU Mihaela1,

ISTRATOAIE Octavian1, MOTA Maria2, MILITARU Constantin1
1 Department of Cardiology, University of Medicine and Pharmacy, Craiova (ROMANIA)
2 Department of Diabetes and Nutrition Diseases, University of Medicine and Pharmacy, Craiova (ROMANIA)
Emails: oananeagoe91@yahoo.com, oana.mirea83@gmail.com, i.donoiu@gmail.com, mihaela.florescu0222@yahoo.com,
droctavist@yahoo.com, mmota53@yahoo.com, cccmilitaru@yahoo.com
* All authors contributed equally to the manuscript conception
Abstract
Background
Incidence of type 1 diabetes mellitus (DM1) is expected to double in the next 20 years and
the risk for developing heart failure and cardiovascular ischemic events is higher in this group
of patients compared to healthy subjects. Diagnosis of cardiac pathology prior to clinical
manifestations is crucial in patients with diabetes.
Objectives
The purpose of this study was to investigate by using measurements derived from
conventional echocardiography and 2D-speckle tracking echocardiography (2D-STE) whether
left atrial (LA) function is impaired in patients with DM1.
Methods
We included 43 young asymptomatic adults diagnosed with DM1 (mean interval from
diagnosis 9±6 years) and 50 healthy young adults. 2D-STE was obtained using the GE Vivid
S6 ultrasound machine and processed using the EchoPAC version 13.0 (GE Healthcare).
Results
There were no differences regarding LA volumes between the groups while LA strain during
reservoir phase was significantly reduced in patients with DM1 compared to healthy subjects
(p<0.01).
Conclusions
Young adults with DM1 without any known cardiovascular pathology display impaired LA
function which may precede the development of diastolic dysfunction.
Keywords: diastolic function, left atrium deformation, speckle tracking echocardiography, type 1 diabetes mellitus
Introduction
Diabetes mellitus (DM) represents a major public health problem, statistics showing a steep
increase of DM prevalence in the past decades, from 4.7% in 1980 to 8.5% in 2014 [1].
Although the majority of subjects presents type 2 diabetes mellitus (DM2), newer studies
show that type 1 diabetes mellitus (DM1) accounts for 5-10% of the total cases of diabetes
14
worldwide [2] and according to EURODIAB registry data, the incidence is expected to double
in the next 20 years [3].
The association between diabetes and cardiovascular events has been extensively studied
and demonstrated. Multiple large-scale studies demonstrated that DM independently increases
by at least two-fold the risk for myocardial infarction and stroke [4] and is strongly associated
with the development and progression of heart failure [5]. Although the mechanism is not fully
understood, DM leads to myocardial contractile dysfunction, myocardial stiffness and, in time,
to heart failure (HF) with preserved left ventricular ejection fraction (LVEF) [6], which
accounts for more than 50% of HF cases [7]. The coexistence of DM and HF leads to a higher
risk for mortality and repeated hospitalization for HF decompensation [8].
Although DM1 is mostly diagnosed during early age and the triggering mechanisms are
different compared to DM2, the latter is also associated with a 10-fold higher risk for
cardiovascular disease in comparison to healthy subjects [9]. The continuously increasing
number of young subjects with DM1 and the lack of large epidemiological studies to clearly
define to which extent the relation between long term DM1 and conventional cardiovascular
risk factors will increase the risk for cardiac events, indicate that subjects with DM1 represent
a vulnerable group in which diagnosis and medical management could be challenging. Thus,
the early detection of diastolic and systolic myocardial impairment in diabetic pathology is of
major importance, as lifestyle changes and prompt medical interventions may prevent or delay
the development of heart failure.
Echocardiography represents a valuable tool for cardiac assessment, providing information
about both systolic and diastolic function. Nonetheless, conventional cardiac ultrasound is less
accurate in detecting sub-clinical cardiac pathology. Two-dimensional speckle tracking
echocardiography (2D-STE) has a superior capacity to determine subtle cardiac dysfunction
compared to conventional echocardiography by evaluating myocardial longitudinal,
circumferential and radial deformation [10]. Furthermore, latest software algorithms of
myocardial deformation analysis allow the measurement of left atrium (LA) deformation [11].
LA dysfunction assessed by 2D-STE was associated with diastolic dysfunction, high risk for
atrial fibrillation (AF), congestive heart failure, stroke and cardiovascular death [12].
The purpose of this study was to investigate whether left atrium function is impaired in
subjects with DM1 as compared with healthy controls.
Methods
Study population
We enrolled 43 patients, aged between 15-40 years old, diagnosed with DM1 (referred as
study group), with no history of cardiovascular disease. Additionally, we enrolled 50 healthy
subjects with no history of cardiovascular disease or DM, considered control group.
Anthropometric information such as age, gender, height, weight and body surface area
(BSA) were collected. Clinical assessment, as well as blood pressure measurement, were
performed prior to echocardiography examination. Biological test results for glycated
haemoglobin, creatinine, urea, cholesterol, were obtained from the patients’ medical charts.
Echocardiographic measurements
The echocardiographic examination was performed at Cardiology Center of Craiova County
Emergency Hospital with a Vivid S6 ultrasound machine, equipped with a 3.5 MHz transducer
(GE Vingmed Ultrasound, Horten, Norway) and all the measurements were made offline using
EchoPAC version 13.0 (GE Healthcare), from apical 4- and 2-chamber view according to the
current recommendations for chamber quantification [13, 14].
15
2D-STE measurements of LA deformation were made from apical 4-chamber view,

according to current expert recommendations [14]. End-diastole, was visually defined as the
frame of mitral valve closure while end-systole was measured as aortic valve closure. The
region of interest of LA longitudinal strain was determined as the endocardial border. (Fig. 1A).
As recommended [14], we have assessed LA longitudinal strain during reservoir phase
(LASr) and during atrial contraction (LASct) (Fig. 1B).
A B
Fig. 1. Measurement of left atrium (LA) longitudinal strain. A. LA range of interest. B. LA strain during
reservoir phase (white arrow); strain during contraction phase (yellow arrow)
Statistical analysis
All characteristics are provided either as mean and/or standard deviation for continuous
variables, or as absolute number or percentage for categorical variables, taken into
consideration to be normally distributed and to have equal variances.
Results
Clinical and biologic characteristics of the subjects

We included 42 young subjects with DM1, with a mean duration of diabetes of 9±6 years
and an average value of HbA1c of 8.9±1.6%. Biologic data is summarized in Table 1.
Diabetic patients were slightly but significantly older than the subjects in the control group
(mean age 31.2±8 vs. 28.3±6 years, p<0.05). There were no important differences of BSA, body
mass index or heart rate between the two groups. However, subjects with DM1 had significantly
higher values for both systolic and diastolic blood pressure in comparison to control group
(126±14 vs. 118±8mmHg; 81±8 vs. 69±6 mmHg, p<0.01).
Table 1. Clinical and paraclinical characteristics of the subjects

Parameter Unit Controls DM1
(n=50) (n=43)
Age (years) 28.3±6 31.2±8*
BSA (m2) 1.88±0.2 1.88±0.2
Heart Rate (bpm) 78±11 80 ±11
SBP (mmHg) 118±8 126±14*
DBP (mmHg) 69±6 81±8*
LDL-cholesterol (mg/dl) - 115±32
Triglycerides (mg/dl) - 101±53
Urea (mg/dl) - 32±10
Creatinine (mg/dl) - 0.97±0.1
HbA1c (%) - 8.9±1.6
BSA, body surface area; DBP, diastolic blood pressure; SBP, Systolic blood pressure.
*: p<0.05 vs. controls
16
Echocardiographic parameters
A summary of echocardiographic measurements is presented in Table 2. LV wall thickness
and mass were higher in patients with DM1 (p<0.01). Additionally, LV volumes were lower in
the study group (p<0.005) while LV ejection fraction showed no significant difference.
Pulsed Doppler measurements showed a decreased early mitral filling velocity and
prolonged E wave deceleration time (p<0.01) in patients with DM1. As for the ratio e/e’, it was
higher in diabetic patients, being an indicator of slightly more elevated LV filling pressures.
LA measurements derived from conventional echocardiography showed that there were no
differences regarding volumes between the groups. However, 2D-STE measurements of LA
demonstrated that LA strain during reservoir phase was significantly reduced in patients with
DM1 compared to healthy subjects (p<0.01). Nevertheless, LA strain during contraction phase
showed no significant difference between the groups.
Table 2. Echocardiographic parameters of the subjects

Parameters Controls DM1
(n=50) (n=43)
IVSd (mm) 8.4±1.6 9.7±2.0*
PWd (mm) 8.3±1.4 9.3±2.2*
LVMi (g/m2) 69±14 76±24 *
LV EDVi (ml/m )
2
53±9 46±11*
LV ESVi (ml/m )
2
22±6 18±6*
LV EF (%) 60±7 60±7
LV Ep (cm/s) 80±19 73±17*
LV Edt (ms) 160±26 184±52*
LV Ep/Ap 1.7±0.5 1.3±0.4*
LV Ea (cm/s) 16±2 12±4*
LV Ep/Ea 5±2 6±2*
LA ESVi (ml/m )
2
22±5 21±6
LA EDVi (ml/m2) 9±3 9±3
LA Res (%) 145±41 154±43
LASr (%) 34.2±7.4 28.7±9.1*
LASct (%) 10.1±4.3 9.8±5.1
Ap: pulsed Doppler transmitral peak late diastolic wave; EDVi: end-diastolic volume index; EF: ejection fraction;
Ep: pulsed Doppler transmitral peak early diastolic wave; Ea: tissue Doppler peak early diastolic wave; ESVi:
end-systolic volume index; IVSd: interventricular septum thickness, diastole; LA: left atrial; LV: left ventricular;
LVMi left ventricular mass index; PWd: postero-lateral wall thickness, diastole; Res: Reservoir expansion; Sr:
strain during reservoir phase; Sct: strain during contraction phase.*: p<0.05 vs. controls
Discussion
DM1 is usually diagnosed at early ages, implying long-term insulin administration.

Although it was demonstrated that DM1 increases significantly the risk for cardiovascular
disease in comparison to healthy subjects [9], the mechanisms and interactions between
cardiovascular disease onset and progression and DM1 are still not clearly defined.
Moreover, there are contradictory data concerning the effects of insulin administration on
the heart [15]. Therefore, it is of major importance to identify new methods for finding subjects
at risk for developing cardiac pathology in the subclinical phase in order to implement
preventive strategies.
Conventional cardiac ultrasound can provide important information regarding the systolic
and diastolic myocardial function, but its value is greatest in the presence of cardiac manifest
disease.
17
Regardless of the fact that conventional echocardiographic measurements are widely

available and essential in assessing cardiac function and structure, in particular situations novel
software algorithms are required in order to detect subtle myocardial dysfunction.
One of the observations that we made was that LV wall thickness and mass are increased in
diabetic patients compared to healthy subjects, possibly as a result of higher blood pressure in
addition to long-term diabetes [16], being well known that LV hypertrophy is frequently
encountered in diabetic cardiomyopathy [6]. Moreover, our study showed that LV relaxation
was mildly altered in diabetic patients, in according to previous studies [17], while ejection
fraction was normal and similar between groups.
LA function and left atrial-ventricular coupling are major determinants of LV diastolic
function [18], and are predictors of long-term morbidity and mortality in general population
[19]. Newer studies show also promising results about the superiority of LA strain to detect LA
remodelling and to predict cardiovascular events [20] in comparison to conventional
measurement.
In our study, both groups had similar LA volumes according to measurements derived from
conventional echocardiography, while 2D-STE measurements proved a significantly reduced
LA strain during reservoir phase in patients with DM1 compared to healthy subjects.
However, concerning the LA strain during contraction phase, there were observed no
differences between the groups. These findings were in accordance with previous studies [21].
LA reservoir function is proved to be in strong correlation with LV function and ejection
fraction [20]. Moreover, there were studies performed that demonstrated the strong relation
between LA reservoir function and AF occurrence in diabetic patients [22].
Conclusions
Our study shows evidence that left atrium longitudinal strain is impaired in young
asymptomatic subjects with DM1 and the measurement could be useful in the detection of
subjects at risk for developing heart failure.
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2. Maahs, D.M., West, N.A., Lawrence, J.M., Mayer-Davis, E.J. (2010). Epidemiology of Type 1
Diabetes. Endocrinol Metab Clin North Am. 39(3), pp. 481-497.
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incidence of childhood type 1 diabetes in 26 European centres in the 25-year period 1989-2013: a
multicenter prospective registration study. Diabetology 62, pp. 408-417.
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(2019). Diabetic cardiomyopathy. Heart 105(4), pp. 337-345.
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260-269.
8. MacDonald, M.R., Petrie, M.C., Varyani, F., et al., (2008). Impact of diabetes on outcomes in patients
with low and preserved ejection fraction heart failure: an analysis of the Candesartan in Heart failure:
Assessment of Reduction in Mortality and morbidity (CHARM) programme. Eur Heart J. 29, pp. 1377-
1385.
9. Laing, S.P., Swerdlow, A.J., Slater, S.D., Burden, A.C., Morris, A., Waugh, N.R., et al., (2003).
Mortality from heart disease in a cohort of 23,000 patients with insulin-treated diabetes. Diabetologia
46(6), pp. 760-765.
18
10. Gunasekaran, P., Panaich, S., Briasoulis, A., Cardozo, S., Afonso, L. (2017). Two-dimensional speckle
tracking echocardiography (2D-STE) has a superior capacity to determine subtle cardiac dysfunction
compared to conventional echocardiography by evaluating myocardial longitudinal, circumferential and
radial deformation. Curr Cardiol Rev. 13(1), pp. 32-40.
11. Gan, G.C.H., Ferkh, A., Boyd, A., Thomas, L. (2018). Left atrial function: evaluation by strain analysis.
Cardiovasc Diagn Ther. 8(1), pp. 29-46.
12. Abhayaratna, W.P., Seward, J.B., Appleton, C.P., Douglas, P.S., Oh, J.K., Tajik, A.J., Tsang, T.S.
(2006). Left atrial size: physiologic determinants and clinical applications. J Am Coll Cardiol. 47(12),
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13. Lang, R.M., Badano, L.P., Mor-Avi, V., Afilalo, J., Armstrong, A., Ernande, L. et al., (2015).
Recommendations for cardiac chamber quantification by echocardiography in adults: an update from
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Thomas, J.D. and Voigt, J.-U. (2018). Standardization of left atrial, right ventricular, and right atrial
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19
Evaluation of the Nutritional Status – Decision Factor in the

Therapeutic Approach in Senior Patient with Hip Fracture
PÎSLARU Anca Iuliana1,3, ILIE Adina Carmen1,3*, SANDU Ioana Alexandra1,3,

ȘTEFĂNIU Ramona1,3, ALEXA Ioana Dana1,3, ALEXA Ovidiu2,3,
PÎNZARU Roxana2,3
1 Department of Geriatrics, “Dr. C.I. Parhon” Clinical Hospital, Iasi (ROMANIA)
2 Department of Orthopaedics and Traumatology, “Sf. Spiridon” Clinical Hospital, Iasi (ROMANIA)
3 “Gr. T. Popa” University of Medicine and Pharmacy, Iasi (ROMANIA)
Emails: morosanu_anca@yahoo.com,* adinacarmenilie@yahoo.com, ioana0sandu@gmail.com,

ramona.stefaniu@yahoo.ro, ioana.b.alexa@gmail.com, ovidiu_alexa@yahoo.com, roxana.bulgarescu@yahoo.com
Abstract
Malnutrition is part of the great geriatric syndromes and is widespread among the seniors
with hip fracture. The scope of the study is the evaluation of the nutritional status and its
correlation with the degree of pre-operative frailty in this population segment. 44 senior patients
were included in the Orthopaedics Section, Iasi, in which the comprehensive evaluation was
evaluated, focusing on evaluating the nutritional status (MNA – Mini Nutrition Assessment)
and frailty (Groningen-IGF frailty index) preoperative.
Most patients evaluated were female, over 80 years old. The risk of malnutrition was about
50% in the studied group and one third of these patients had malnutrition. At the same time,
most of the patients were fragile and their precarious nutritional status was statistically
significantly correlated with frailty.
In conclusion, it could be emphasized that protein-caloric malnutrition and frailty are
negative prognostic factors for postoperative evolution and during the recovery period, thus,
this study emphasizing the importance of finding and monitoring the nutritional status of the
frail senior population.
Keywords: nutritional status, frailty, hip fracture, senior patients
Introduction
Malnutrition is considered a geriatric syndrome in the senior population in general and in

particular in those with a hip fracture, which results in the influence of the therapeutic decision,
the immediate postoperative evolution and the ability to recover.
The high prevalence of malnutrition in senior patients with hip fracture has been well
documented [1-3], as well as the consequences of this related to duration of hospitalization [4],
functional recovery, disability and mortality [5].
Protein-caloric malnutrition in the seniors must be detected with the help of specific tests
(Mini Nutrition Assessment-MNA, bioimpedance) because the anthropometric determinations
do not faithfully reflect the balance between low and fat mass of medical assistance [4-6].
Practically, the association of the poor nutritional status, the cognitive impairment and the
functional dependence, which is frequently noticed in the senior patients, marks an extreme
frailty situation, with a risk of complications related to hospitalization, permanent disability and
death after fracture. [2, 5, 7, 8]
Malnutrition with sarcopenia, osteopenia and balance and gait disorders are part of the
clinical manifestations of frailty. The frailty syndrome is reported to affect 7% of elderly
20
patients aged over 65 years, and 25-40% of those aged 80 years or over. The borders between
age and frailty appear to be so indistinct that it is widely assumed that at a specific age, all
people become frail [9].
Early detection and recognition of frailty is necessary, in order to prevent its serious
consequences, such as: repeated falls, fractures, multiple hospitalizations, institutionalization
and death. Implicitly, frailty in older adults may be necessary in preventing disability after an
acute event.
The proposed objective is to evaluate the preoperative nutritional status in the elderly patient
who suffered a hip fracture and the correlation with the degree of frailty.
Methodology
The prospective study, from which we present the preliminary data collected in June-October
2019, on a lot of 44 patients, aged between 62-93 years, hospitalized in the Orthopaedics
Section of the “Sf. Spiridonˮ Clinical Hospital Iasi, after suffering a hip fracture.
A comprehensive evaluation was performed, with special attention being given to assessing
the nutritional status (MNA - Mini Nutrition Assessment) and frailty (Groningen-IGF frailty
index), in the preoperative period. Depending on the score obtained in the MNA test, patients
were considered to have normal nutritional status (˃24 points), risk of malnutrition (17-24
points) or malnourished (<17 points) and all frail at an IGF≥4. The completion of the
questionnaires was done after obtaining the informed consent of the patient, being made by the
same trained medical team.
The data was collected by the medical staff involved in the study and was analysed using
SPSS 18.0 software. Anova Test, Chi-square Test and Paired T-test, Kruskall-Wallis and
Pearson correlation for quantitative analyse where required. The statistical significance was
defined in 95% interval of confidence (p<0.05).
Results
The age group ranged from 62 to 93 years, with an average of 80.36 years ± 7. 59. Gender
distribution was predominantly female (70.5%), the ratio F/M=2.4/1.
In men, equal weights were recorded in the 6, 7 and 8 age groups (30.8% each), while in
women there was a significantly higher weight in the 8-age decade (64.5%) (p=0.05) (Fig. 1).
Fig. 1. Distribution of the lot by age and gender groups
21
The percentage distribution of cases according to the nutritional status highlighted the
following aspects (Fig. 2):
- malnutrition risk accounted for 45.5% of the total study group;
- malnutrition was identified in 22.7% of cases.
normal risk of malnutrition malnutrition
22,7%
31,8%
45,5%
Fig. 2. The structure of the lot according to the nutritional status
The evaluation of the nutritional status, showed statistically significant percentage

differences, 60% of the married or divorced patients suffered from malnutrition, while 46.7%
of the married patients and 54.2% of widow patients had risk of malnutrition (p=0,022) (Fig.
3).
100%
6,7
90% 25
80%
70% 46,7 60
60%
malnutrition
50% 54,2 risk of malnutrition
40% 0
normal
30%
20% 46,7
40
10% 20,8
0%
Married Widower Divorced / unmarried
Fig. 3. Correlation of nutritional status with marital status
Preoperative frailty according to the IGF score varied in the range 1-12, with an average of
6.3232±3.10 (Fig. 4).
22
Fig. 4. Histogram of IGF score in preoperative patients
79.5% of cases had frailty (IGF≥4 score) (Fig. 5).
robust frail
20,5%
79,5%
Fig. 5. Structure of the lot according to the presence of frailty
The MNA and IGF scores were in indirect correlation, of high intensity (r=+0.601; p=0.001),
statistically significant, noting that about 60% of the patients associated frailty with
malnutrition (Fig. 6).
Fig. 6. Correlation of frailty with MNA score
23
Discussions
In their study, Zanetti M et al. demonstrate the strong negative impact of malnutrition on
mortality in the senior population with hip fracture and strengthens the idea of extended
preoperative nutrition screening [10]. In line with these conclusions, Gumiero et al., [6] showed
that the use of the MNA nutrition test is superior to other instruments for mortality 6 months
after hip fracture.
In their observational study Goisser S. et al., pointed out that more than half of the patients
with hip fracture were malnourished or at risk of malnutrition [11]. Thus, the scores obtained
in our study are in agreement with the previous ones, with a higher weight for patients with
malnutrition risk.
One of the most important contributions to malnutrition is solitary life. Solitude, as well as
depression, are statistically significantly correlated with the precarious nutritional status, an
aspect supported by the other studies in the field.
Our study demonstrates once again that there is a close relationship between protein-caloric
malnutrition and the degree of frailty, an issue mentioned by Bollwein et al., [12] in their study,
where it appears that the percentage of people at risk of malnutrition increased progressively
with the level of frailty.
Conclusions
The pre-operative recognition of the elements that suggest the vulnerability of the geriatric
patient (e.g. nutritional status, frailty, disability) can provide additional information in the pre-
operative prediction of complications and subsequent evolution, favouring individualized
decisions. Consequently, nutritional screening and detection of frailty should be included in any
stratification of admission risk in the case of hip fractures.
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24
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25
Efficacy and Safety of Adding Sitagliptin in Patients with Type 2

Diabetes Mellitus with Inadequate Glycaemic Control in
Metformin Monotherapy
ALBAI Oana1,2, TIMAR Bogdan1,2, TIMAR Romulus1,2,3

1 “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
2 Center of Diabetes, Nutrition and Metabolic Diseases, Timisoara (ROMANIA)
3 Department of Diabetes, Nutrition and Metabolic Diseases, County Hospital, Timisoara (ROMANIA)
Email: oana_olari@yahoo.com
Abstract
Background and Aims

Type 2 diabetes mellitus is a metabolic disorder becoming more prevalent worldwide.
Although, it is known that a tight glycaemic control decreases the incidence and progression
of diabetes complications, in some patients this cannot be reached despite intensifying of the
pharmacological therapy. The treatment options were enriched with new agents, acting on the
incretion pathway. Sitagliptin, a dipeptidyl peptidase-4 inhibitor, increases level of GLP-1 and
GIP hormones, being an option for improving the glycaemic control. The main purpose of this
study was to assess the efficacy and safety of adding sitagliptin in patients with type 2 diabetes
mellitus treated only with metformin.
Material and Method

The study included 148 patients with type 2 diabetes mellitus, with an average age of
56.7±5.9 years and a history of DM of 7.2±3.1 years. Patient’s characteristics were measured
at baseline and after 26 weeks after the adding of sitagliptin to the previous metformin
monotherapy.
Results
We observed an average decrease in HbA1c with 0.7%. We also noted a significant decrease
in the value of fasting and postprandial glycemia, as well as an average weight loss of 2.2 kg.
Conclusion
The combined therapy, sitagliptin and metformin is demonstrated to be efficient, sustainable,
safe for the patients and its results are to be observed in short time after its initiation.
Keywords: diabetes mellitus; dipeptidyl peptidase-4 inhibitor; sitagliptin; metformin
1. Background and Aims
Type 2 Diabetes Mellitus (T2DM) is a prevalent metabolic disorder which requires lifelong
treatment for preventing the development of its acute and chronic complications. Currently, 425
million individuals are diagnosed with T2DM worldwide and their number it’s estimated to
reach 629 million in 2045 [4].
All opinions are pointing that the intensive management of hyperglycaemia, with the
consecutive achievement of a good glycaemic control, prevents the outset or the progression of
chronic complications, especially the microvascular ones. The United Kingdom Prospective
26
Diabetes Study (UKPDS) demonstrated that a decrease of HbA1c with one percentage point is
associated with a 37% relative risk reduction in microvascular complication of patients with
T2DM. Also, its ten years follow-up indicated that, despite an early loss of glycaemic
differences, a continued reduction in microvascular risk and emergent risk reductions for
myocardial infarction and death from any cause were observed in initially intensive treated
cohort [6,7].
The American Diabetes Association (ADA) and the European Association for the Study of
Diabetes (EASD) developed a series of guidelines for the management of hyperglycaemia in
patients with T2DM [4], which are recommending as a therapeutic target an HbA1c value lower
than 7%, value which corresponds to average blood glucose of 8.3-8.9 mmol/L (150-160
mg/dL). Also, these guidelines are suggesting a fasting blood-glucose target of <7.2 mmol/L
(<130 mg/dL) and a post-prandial value <10 mmol/L (<180 mg/dL) [4].
The therapy with a single hypoglycaemic agent is often insufficient to obtain and/or maintain
a satisfactory glycaemic control in patients with T2DM. Metformin is a first-line medication,
the mostly used to treat patients with T2DM. It acts mainly by decreasing hepatic glucose output
and by reducing insulin-resistance (IR), without causing weight gain and with a low risk of
hypoglycaemia events.
If the monotherapy cannot be adjusted to reach and/or maintain HbA1c target values <7%
after 3 months, the next step should be to combine it with another oral antihyperglycemic agent.
Adding sitagliptin in patients with T2DM treated only with Metformin improves their
glycaemic control, these two substances having complementary mechanisms of action. The
American Association of Clinical Endocrinologists (AACE) recommends adding sitagliptin in
the second line antihyperglycemic therapy [3].
Sitagliptin is an oral antihyperglycemic agent that reversibly inhibits the dipeptidyl
peptidase-4 enzyme (DPP-4), slowing down the degradation of the incretin hormones GLP-1
and GIP, increasing in this way their concentration and action. The therapy with sitagliptin
increases up to two-fold the postprandial plasma concentrations of endogenous GLP-1
compared to placebo therapy in patients T2DM. Through all these mechanisms, sitagliptin
significantly improves the pancreatic beta cell function and the glycaemic control [12, 13].
Our study’s aim was to evaluate the optimization of the glycaemic control after the addition
of sitagliptin in patients with T2DM treated with metformin monotherapy. As secondary
endpoints we assessed the effect of combined metformin and sitagliptin therapy on body weight,
safety and tolerability.
2. Material and method
2.1 Study design

We enrolled 148 patients with T2DM: 79 men (53.4%) and 69 women (46.6%), with an
average age 56.7±5.9 years and a mean duration of DM 7.2±3.1 years. All patients signed and
informed consent form, previously approved by the local ethics committee.
The enrolment process was a population based consecutive one, having as inclusion criteria
the previous diagnosis of T2DM, treated only with metformin (at a dose of at least 1500
mg/day), aged 35 to 70 years, having a body mass index (BMI) between 20 and 40 kg/m 2 and
having an unsatisfactory glycaemic control (HbA1c between 7 and 9%). Patients with
documented severe cardiovascular disease (CVD), heart failure, a glomerular filtration rate
(GFR) less than 45 ml/min and with a 3-fold increase of the level of their serum transaminases
(AST, ALT) were not enrolled in the study.
All patients received counselling regarding the change of their lifestyle (diet, physical
exercise) needed.
27
2.2 Studied parameters

At baseline and after 26 weeks of combined therapy we measured using standardized
methods the following parameters: weight, waist circumference (WC), BMI, systolic blood
pressure (SBP), diastolic blood pressure (DBP), HbA1c, fasting and postprandial glycemia,
serum creatinine, GFR, AST, ALT, total cholesterol (TC), triglycerides (TG) and HDLc.
LDLc was calculated using Friedewald’s formula (if the TG values were lower than 400
mg/dL): TC-(TG/5)-HDLc (mg/dL) or it was measured directly if TG values were higher than
400 mg/dL.
2.3 Statistical analysis

Data were collected and analysed using SPSS v.17 software suite (SPSS Inc. Chicago, IL,
USA) and are presented as mean ± standard deviations (for continuous variables with Gaussian
distribution), or percentages (categorical variables).
In order to assess the significance of the differences between groups, t-student (means,
Gaussian populations), Mann-Whitney U (not-Gaussian populations), and chi-square with
Yates correction (proportions) tests were used. Continuous variable distributions were tested
for normality using Shapiro-Wilk test and for equality of variances, with Levene’s test.
A p-value <0.05 was considered the threshold for statistical significance.
3. Results
Patient’s baseline characteristics are presented in Table 1.
Table 1. Patient’s baseline characteristics

Number 148
Men (%) 79 (53.4%)
Age (years) 56.7±5.9
Diabetes duration (years) 7.2±3.1
BMI (kg/m2) 30.8±3.1
Abdominal circumference (cm) 101.2±10.6
Men 108±11.4
Women 94.5±8.6
Weight (kg) 93.4±15.8
HbA1c (%) 8.4±1.0
Fasting glycemia (mg/dL) 164±44.7
Postprandial glycemia (mg/dL) 236.4±54.8
TC (mg/dL) 208±34.2
TG (mg/dL) 214±96.7
HDLc (mg/dL) 44.8±6.9
LDLc (mg/dL) 120.4±42.6
Variables are Gaussian distributed. Data is presented as mean ± SD
In the study group, 28 patients (18.9%) were normal weight (BMI <25 kg/m 2), 69 patients
(46.6%), overweight (BMI between 25-29.9 kg/m2) and 51 patients (34.5%) were obese (BMI
≥30 kg/m2).
The mean HbA1c decreased with 0.7 percentage points, from 8.4%±1.0 to 7.7%±0.8 after
26 weeks of treatment (p<0.001) (Fig. 1).
28
HbA1c
(%) HbA1c0
9 HbA1c1
8,4
7,7
5
Fig. 1. Average HbA1c, before and after the 26 weeks
The association of sitagliptin in patients with T2DM previously treated only with metformin
was associated with a significant decrease of fasting glycemia, observed both after 12 and 26
weeks of intensified treatment. The observed average decrease was with 28 mg/dL in the
studied interval, having statistical significance (p<0.001) (Fig. 2).
Fig. 2. Evolution of fasting glycemia in the patients included in the study group
We also observed significant improvements regarding the 2-hours postprandial glycemia:

from an average value of 247.4 mg/dL at baseline, at the end of the study a mean postprandial
glycemia of 187.1 mg/dL was observed (p<0.001) (Fig. 3).
Fig. 3. Evolution of postprandial glycemia in the study group
29
The average weight was reduced with 2.2 kg: from 93.4±8.8 kg to 91.2±6.2 kg, however this
reduction was not significant (p=0.16) (Fig. 4).
G (kg)
99 p<0,16
G0
97 G1
95
93,4
93
91,2
91
89
87
85
Fig. 4. Average weight (kg), before and after the 26 weeks
Clinically significant reductions were observed in HbA1c values in all age groups. By adding
sitagliptin to previous metformin monotherapy, HbA1c decreased with 0.9% in patients <50
years (p<0.001), 0.6% in those aged between 50 and 60 years (p<0.001) and 0.6% in those aged
between 60 and 70 years (p=0.008) (Fig. 5).
% HbA1c0 (%)
9 8,4 8,6
8,2 HbA1c1 (%)
8,5
8
8 7,5 7,6
7,5
7
6,5
6
5,5
5
<50 ani 50-60 ani 60-70 ani
Fig. 5. HbA1c improvements in different age groups
The treatment with sitagliptin was associated with a significant decrease in HbA1c,
irrespective of the time elapsed since the patients were diagnosed with T2DM (Fig. 6). In all
these groups the decreases were statistically significant (p<0.001).
30
%
9 8,5 HbA1c0 (%)
8,3
8,4 HbA1c1 (%)
8,5
7,8
8 7,6 7,7
7,5
7
6,5
6
5,5
5
<5 ani 5-10 ani >10 ani
Fig. 6. HbA1c decreases regarding disease duration
After the initiation of sitagliptin treatment we observed also a change in the distributions of
the HbA1c in the studied group. The proportion of patients with inadequate glycaemic control
decreased along with the increase of the proportion of patients within guidelines
recommendation (Table 2).
Table 2. The distribution of HbA1c values, before and after sitagliptin treatment
HbA1c value/No. patients (%) <7% 7-7.9% 8-9%
Metformin 0 78 (52.7%) 70 (47.3%)
Metformin + Sitagliptin 64 (43.2%) 66 (44.6%) 18 (12.2%)
p-value <0.001 <0.001 <0.001
The percentage of patients who achieved HbA1c target, without a body weight increase of
over 2% and without having hypoglycaemic events, was 35.1%. The percentage of patients with
a decrease of HbA1c more than 0.3%, without an increase of over 2% in body weight and no
hypoglycaemia was 79.7%.
The comparison between studied parameters at baseline and after 26 weeks of dual therapy
is presented in Table 3.
Table 3. Comparison between the main characteristics studied in patients in Metformin monotherapy,
versus the combined therapy Metformin + Sitagliptin
Baseline After sitagliptin + p
metformin
HbA1c (%) 8.4±1 7.7±0,6 <0.001
Weight (kg) 93.4±8.8 91.2±6.2 0.16
Fasting glycemia (mg/dL) 164±44.7 136.2±32.8 <0.001
Postprandial glycemia (mg/dL) 236.4±54.8 187±48.9 <0.001
TC (mg/dL) 208±34.2 198±30.7 0.008
TG (mg/dL) 214±96.7 184.1±91.6 0.007
HDLc (mg/dL) 44.8±6.9 46.7±7.2 0.021
LDLc (mg/dL) 120.4±42.6 112.3±34.7 0.074
Variables are Gaussian distributed. Results are presented as mean ± SD. p was calculated using paired t-
student test.
31
The incidence of hypoglycaemic episodes was very low (2%): no severe hypoglycaemia or
major cardiovascular events were observed. The treatment with sitagliptin was not interrupted
in any of these patients.
4. Discussions
Many studies presented the effects of the therapy with sitagliptin in patients with T2DM.
Two similar studies which enrolled 741, respectively 521 patients aimed to evaluate the
efficacy of the sitagliptin monotherapy during 24 and 18 weeks. Mean baseline HbA1c was 8%
and 8.1%. In both trials, patients received sitagliptin 100 mg/day, sitagliptin 200 mg/day or
placebo. HbA1c decreased by 0.6% after 18 weeks, and by 0.8% after 24 weeks of study [5,
19].
Another study was conducted on 353 patients with a mean baseline HbA1c of 7.5%, who
received in combination with metformin (1500 mg) either sitagliptin 100 mg/day or glipizide 5
mg/day. After 52 weeks of observation, HbA1c decreased by 0.7%, thus demonstrating the non-
inferiority of sitagliptin compared to glipizide. Furthermore, the patients treated with metformin
and sitagliptin had fewer hypoglycaemic events. The weight loss observed in this subgroup was
1.5 kg, compared to weight gain of 1.1 kg in the subgroup treated with glipizide added to
background metformin [17].
At least three mechanisms are involved in the etiopathogenesis of T2DM: reduction of
insulin secretion, IR and hypersecretion of glucagon [18]. The reasoning for the combined
therapy with sitagliptin and metformin is based on two strategies, such as: metformin acts
primarily by decreasing hepatic glucose production, thus improving liver and muscle IR [9, 11],
while DPP-4 inhibitors act by increasing GLP-1 levels, stimulating insulin secretion and
reducing glucagon release [1, 2]. Furthermore, it seems that metformin increases GLP-1 serum
level by stimulating gut secretion [14, 15].
Charbonnel et al., studied the effects of the combined therapy sitagliptin and metformin in a
group of 701 subjects with an average HbA1c of 8%. The patients received metformin
monotherapy 1500 mg/day, to which sitagliptin 100 mg/day or placebo was added. After 24
weeks of assessment, the therapy with sitagliptin reduced HbA1c by 0.7%, compared to
placebo. Also, 47% of the subjects receiving sitagliptin reached the therapeutic target HbA1c
<7%, compared to only 18% in patients treated with placebo [8]. The treatment with sitagliptin
was not accompanied by side effects as hypoglycaemia, gastrointestinal reactions or significant
changes in body weight. An improvement in serum levels of C-peptide, basal insulin and
proinsulin/insulin ratio was also observed.
A large study conducted on 1091 patients with T2DM and an average HbA1c of 8.8%,
assessed for a period of 24 weeks the effects on glycaemic control of the metformin therapy in
various doses and sitagliptin as monotherapy or in combination. HbA1c was lowered by 2.9%
in the subgroup of patients who received a combined therapy (sitagliptin plus metformin).
Also, the combined therapy significantly improved the beta cell function, as measured by
HOMA-B index [10].
5. Conclusions
The association between metformin and sitagliptin proved to be highly effective in patients
with T2DM with unsatisfactory glycaemic control treated only with metformin. Sitagliptin
therapy was well tolerated, without significant side effects.
Both preparations are easily accepted due to their oral administration and the patient’s
compliance to treatment increases further by taking a single tablet containing sitagliptin and
metformin in combination.
32
We can affirm that the initiation of the combined therapy with sitagliptin and metformin as
first line treatment is more efficient compared to the metformin monotherapy in many patients
with T2DM.
Further studies are needed to investigate the effects of this combined therapy on the
preservation of pancreatic beta-cells function, as well as on the occurrence of cardiovascular
complications.
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2009; 70: pp. 343-353.
2. Bergman AJ, Stevens C, Zhou Y, et al., Pharmacokinetic and pharmacodynamic properties of multiple
oral doses of sitagliptin, a dipeptidyl peptidase-IV inhibitor: a double-blind, randomized, placebo-
controlled study in healthy male volunteers. Clin Ther, 2006; 28: pp. 55-72.
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monotherapy on glycaemic control in patients with type 2 diabetes. Diabetes Care, 2006; 29: pp. 2632-
2637.
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added to ongoing metformin therapy in patients with type 2 diabetes. Curr Med Res Opin, 2008; 24: pp.
537-550.
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peptidase-4 inhibitor, sitagliptin, compared with the sulfonylurea, glipizide, in patients with type 2
diabetes inadequately controlled on metformin alone: a randomized, double-blind, noninferiority trial.
Diabetes Obes Metab, 2007;9: pp. 194-205.
11. Nauck MA. Unravelling the science of incretin biology. Am J Med. 2009; 122: pp. S3-S10.
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metformin therapy in patients with inadequately controlled type 2 diabetes on metformin alone. Diabetes
Care, 2009; 32: pp. 1649-1655.
13. Hinke SA, McIntosh CH, Hoffmann T, et al., On combination therapy of diabetes with metformin and
dipeptidyl peptidase IV inhibitors. Diabetes Care, 2002; 25: pp. 1490-1491.
14. Ahren B et al. Inhibition of dipeptidyl peptidase IV improves metabolic control over a 4-week study
period in type 2 diabetes. Diabetes Care, 2002; 25: pp. 869-875.
15. Ahren B. Dipeptidyl peptidase-4 inhibitors. Diabetes Care, 2007; 30: pp. 1344-1350.
16. Mannucci E, Ognibene A, Cremasco F, et al., Effect of metformin on glucagon-like peptide 1 (GLP-1)
and leptin levels in obese nondiabetic subjects. Diabetes Care, 2001; 24: pp. 489-494.
17. Mudaliar S, Henry RR. Incretin therapies. Effects beyond glycaemic control. Am J Med, 2009; 122: pp.
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18. Charbonnel B, Karasik A, Liu J, Wu M, Meininger G. Efficacy and safety of the dipeptidyl peptidase-4
inhibitor sitagliptin added to ongoing metformin therapy in patients with type 2 diabetes inadequately
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33
Is in Vitro Fertilisation Pregnancy Different to Natural Pregnancy

Concerning Gestational Diabetes Risk? A Short Review
MATEI Alexandra1,2, IONESCU Cringu Antoniu1,2, GHEORGHIU Diana2,

CATRINA Eduard Lucian1, AL AZAWI Alla2, ROSU George-Alexandru1,2,
ZYGOUROPOULOS Nikolaos2, CALIN Florin Daniel1,2,
PANTEA STOIAN Anca1, NAVOLAN Dan3, PACU Irina1,2
1 “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
2 Department of Obstetrics & Gynaecology, “Sf. Pantelimon” Clinical Emergency Hospital, Bucharest (ROMANIA)
Emails: ale.matei@gmail.com, antoniuginec@yahoo.com, cdgheorghiu@hotmail.com, alazawi.alla@yahoo.com,

george.rosu@ymail.com, nz@zitaquality.ro, dkcalin@gmail.com, ancastoian@yahoo.com, navolan@yahoo.com,
irinapacu@hotmail.com
Abstract
The prevalence of gestational diabetes mellitus (GDM) varies worldwide depending on

racial and ethnic groups, screening practice, population characteristics and diagnostic criteria.
The prevalence has been rising due to an increase in mean maternal age, weight and obesity.
The importance of screening and diagnosis of GDM resides in the possibility of severe
maternal and adverse fetal outcome, studies have shown that the relative risk of developing
GDM is approximately twice higher in in vitro fertilisation (IVF) singleton pregnancies than in
unassisted singleton pregnancies. Different procedure-related variables like the source of the
zygote (autologous vs donor), quality of the egg (fresh vs frozen) and the number of transferred
embryos has shown to have over time a significant impact on GDM risk, questioning medical
practitioners the safety of assisted reproductive techniques.
Keywords: diabetes mellitus, pregnancy, in vitro fertilisation, assisted reproductive techniques
Introduction
Infertility has become a pathology of the modern era, recent studies reporting that only in
the United States of America it affects approximately 6.1 million people, equivalent to 10% of
the reproductive-age population [1]. As a consequence, the use of assisted reproductive
techniques (ART) including IVF has increased, leading worldwide medical practitioners
towards confronting new challenging medical scenarios regarding adverse pregnancy
outcomes.
In Europe, one in 50 children born is the result of ART treatments [2]. Although data on the
continent is not homogeneous and Romania is still experiencing a pioneering transition in
infertility management compared to other European states, in 2014 our country reported a
number of 19 IVF clinics where 1201 IVF procedures were performed in that year alone. The
European Society of Human Reproduction and Embryology (ESHRE) concluded in the last
2014 annual report that compared to 2013 not only the number of reporting clinics has increased
(1169 in 2013 to 1279 in 2014, +9.5%), but also the overall number of reported treatments (686
271 in 2013 to 776 556 in 2014, +13.1%) [2].
Twenty-four years after the first IVF baby was born in our country, “cross-border egg
donation is still the missing link” for our national IVF procedures, possibly responsible for the
reduced delivery rates as well (39% pregnancies per aspiration and 27.8% delivery rate per
34
aspiration in 2014) [2, 3]. The complexity of the IVF technique, which involves ovarian
hyperstimulation with corresponding supraphysiological levels of oestradiol as well as a zygote
and early embryo micromanipulation, comprises elements susceptible to be primary drivers for
the adverse maternal and fetal outcome. This explains the raised awareness and focus on
studying the safety and possible collateral adverse effects of ART procedures.
Referring to the risk of GDM in ART pregnancies, in the early 2000s, specialists in the field
have faced contradictory situations. While a multivariate analysis on singletons born from ART
showed an increased risk of GDM compared to those conceived naturally, studies assigned on
multiple pregnancies were still under a broad spectrum of uncertainties, many of them
questioning whether or not a possible increased risk of GDM could be iatrogenic or intrinsic
[4]?
It was later, in 2018, that the first and largest study on transcriptomic profiles of the first-
trimester placenta could prove that there are few differences in global gene expression among
spontaneous versus IVF pregnancies; there was only one gene found to be differently expressed
between the two groups: CACNA1I, encoding a calcium channel responsible for release of
hormones or neurotransmitters among others [5].
Whether or not molecular alterations related to ART procedure itself or maternal
characteristics of women who conceive by ART are responsible for the increased incidence of
GDM in assisted pregnancies, studies have shown that the relative risk of developing GDM is
approximately twice higher (RR:1.48, CI:1.33-1.66; RR:2.23, CI:1.85-2.69) in IVF singleton
pregnancies than in unassisted singleton pregnancies [6, 7].
Solutions have been sought to reduce the risk of adverse obstetrical outcomes. This way, a
controlled technique of multifetal pregnancy reducing had been approved earlier in 1980. This
would decrease the inherent risk of preterm birth and/or low birth weight that twin pregnancies
are at higher risk of. Vieira et al., compared pregnancy evolution between elective twin
pregnancy reduction and ongoing twin gestations. Both groups associated with IVF and
spontaneous pregnancies. The study concluded that the risk of GDM for twin pregnancies was
not significantly different than the one for 2-to-1 reduced singleton pregnancies (p=0.08, OR
1.94 CI:0.92-4.09), after adjusting by age, body mass index, race, use of IVF, use of chorionic
villus sampling, prior term birth, and previous preterm birth [8].
Furthermore, considering the different embryo transfer (ET) methods: fresh versus frozen,
it has been demonstrated that both protocols associated an increased risk of GDM compared to
spontaneous pregnancies: 7.6% vs 4.6% for frozen ET and 8.6% vs 4.6% for fresh ET (p<0.01)
[9]; all three patient subgroups comprised of both singleton and twin pregnancies.
Overall, female patients attempting pregnancies via ART procedures are more often
advanced in age with reduced ovarian reserve. Speciality literature confirms that age,
overweight and multiple pregnancies are GDM risk factors in IVF subjects [10]. It was a matter
of time until researchers turned their attention towards donor-oocyte IVF. Few available studies
report conflicting results and confounding factors like maternal age were considered to be
excluded. Yadav et al., published in 2018 that GDM was found more frequently in donor-oocyte
IVF group as compared to self-oocyte IVF group 34% vs 15%, p=0.001; however, when
regression analysis model was used for age-matched results, it was not significant (p=0.234)
[11].
GDM is considered the most common complication in pregnancy, and it leads to undesirable
consequences on embryo integrity by interfering with motor neuron function, glucose
metabolism, embryo growth, and so on [10]. Approaching this obstetrical matter from a
different perspective leads to the evaluation of risk factors and corresponding ways to fight
them. In 2019, Charkamyani proposed the empowerment of clinicians to take practical
measures on promoting a healthy lifestyle during pregnancy. His studies pointed out that
physical activity-related practices in women undergoing IVF led to a significant reduction in
35
GDM; concerning the implementation of a nutritional program in the same context, the same
results could not be obtained [10, 12].
Advising overweight and obese patients who undergo IVF to lose weight was the target of
another study which found that IVF appeared to be an independent risk factor for developing
GDM in overweight women [13].
In our country, unfortunately, we experience a concerning deficit in medical education:
whether this implies 3rd-trimester pregnant women who lack proper obstetrical surveillance and
confront themselves with tragic situations [14] or patients who disregard gynaecologic
symptoms and finish by adding up to national increased cervical cancer incidence clusters [15],
the alarm signs need to be drawn. Nutrition education and weight control are the first
approachable steps to be followed. A healthy body mass index can maintain physiologic
endocrinologic functions reflecting on the genital system as well. Furthermore, literature case
reports prove that even polycystic ovarian disease can be correlated with sporadic malignant
tumours which can raise significant challenges for clinicians due to the nature of their
complexity and elusive diagnosis [16]. On the other hand, there is no correlation with pre-
invasive cervical lesions or myelofibrosis [15, 17].
The last aspect to consider is related to further obstetrical complications and comorbidities
associated in GDM following singleton pregnancies achieved by ART. Evocative for this matter
is a prospective cohort study which compared naturally conceived pregnancies with IVF
pregnancies, both groups being previously diagnosed with GDM. The results showed that the
IVF group had a higher risk for pregnancy induced-hypertension, preeclampsia, antepartum
haemorrhage, emergency caesarean section and unfavourable perinatal outcomes like preterm
birth, low birth weight, neonatal hypoglycaemia and Neonate Intensive Care unit admission.
Also, the insulin requirement was higher in women with IVF pregnancies compared to
unassisted pregnancies [18, 19].
Conclusion
To conclude, the role of ART techniques in achieving pregnancies in female patients with
poor ovarian reserve or following other unsuccessful treatments is nowadays the Holy Grail in
reproductive medicine. Alongside succeeding to improve pregnancy rates which is the primary
purpose of the procedure, researched are making breakthrough progress in investigating the
safety and impact of ART on the obstetrical framework as well as over foetal outcome
throughout life. Good knowledge of Guidelines and permanent literature update are a must for
any physician practising in the field. Over time, obstetrics has kept showing that sometimes
instead of taking a step forward by accessing advanced treatment options, it is more efficient to
look back and prevent.
REFERENCES
1. Faddy MJ, Gosden MD, Gosden RG. (2018). A demographic projection of the contribution of assisted
reproductive technologies to world population growth. Reprod Biomed Online 36, pp. 455-8.
2. De Geyter Ch, Calhaz-Jorge C, Kupka M.S., et al., (2018). ART in Europe, 2014: results generated from
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37
Association between Helicobacter Pylori Infection, Obesity

and Gestational Diabetes
CODREANU Ana-Maria1*, MATEI Alexandra2,4*, BORONTEA Minodora1,

OLARU Luciana1,3, OLARU Flavius1
1 Department of Obstetrics, Gynaecology and Neonatology, “Victor Babes” University of Medicine and Pharmacy,
Timisoara (ROMANIA)
3 Department of Gastroenterology, Military Emergency Hospital, Timisoara (ROMANIA)
4 Department of Obstetrics and Gynaecology, “Sf. Pantelimon” Clinical Emergency Hospital, Bucharest (ROMANIA)
* CAM and equally contributed to this article and should both be regarded as the first author.
Emails: ale.matei@gmail.com, luciana_olaru@yahoo.com
Abstract
Gestational diabetes (GDM) is a carbohydrate intolerance of variable severity with onset or

first recognition during pregnancy. The gram-negative bacillus Helicobacter pylori (H. pylori)
is one of the leading causes of chronic gastric infection. It is estimated that 50% of the world
population is infected with H pylori, with a higher prevalence in the developing countries. H.
pylori infection has been linked to carcinogenesis and also to toll-like receptors activation via
lipopolysaccharides, leading to metabolic changes and insulin resistance. The objective of this
study was to assess the scientific literature which evaluates the association between H. pylori
infection and gestational diabetes, respectively obesity. We questioned the PubMed database
(December 10th, 2019) using the keywords “gestational diabetes and Helicobacter Pylori”
obtaining four articles and by “Helicobacter Pylori and gestational weight gain” receiving three
reports. While studies revealed an association between H pylori infection and gestational
diabetes occurrence, other studies showed controversial results referring to the association with
obesity.
Keywords: Helicobacter Pylori, pregnancy, obesity, gestational diabetes
Introduction
Gestational diabetes (GM) is recognised as carbohydrate intolerance of variable severity

with onset or first recognition during pregnancy. American College of Obstetricians and
Gynaecologists recommend a two-step approach to screen and diagnose gestational diabetes.
Two-step approach begins using 50-gram oral glucose. After 1-hour post glucose ingestion,
the plasma glucose level is evaluated, and followed by 100 grams oral glucose and after 3 hours
plasmatic glucose level is measured. Screening should be performed between 24- and 28-
weeks’ gestation in those women not known to have glucose intolerance earlier in pregnancy
[1, 2]. Other guidelines recommend an earlier pregnancy screening at the time of the first
prenatal visit [3].
Treatment of gestational diabetes is targeting maintaining the fasting plasma glucose levels
95 mg/dL or the 2-hour postprandial plasma glucose 120 mg/dL.
Because of the relationship between pregnancy complications and maternal glycaemic
control, efforts to achieve glucose targets are typically more aggressive during pregnancy [4].
Insulin has been considered standard therapy in cases where the fasting levels persistently
exceed 95 mg/dL. The starting dose is typically 0.5-1.0 units/kg/day a combination of
38
intermediate-acting and short-acting insulin. The American College of Obstetricians and

Gynaecologists recommends that insulin be considered in women with 1-hour postprandial
levels that persistently exceed 140 mg/dL or those with 2-hour levels above 120 mg/dL [5].
The mother and the fetus, suffer serious complications directly linked to diabetes: gestational
hypertension, preterm birth, macrosomia, malformation, fetal-growth restriction, stillbirth,
perinatal death [4]. Gestational diabetes screening should be performed in all pregnancies as
well as first [6-8] or second-trimester aneuploidies screening [9, 10]. Also, foetuses from
pregnancies with gestational diabetes need intense surveillance in the antenatal [11, 12] and
postnatal period [13, 14] as described elsewhere.
The gram-negative bacillus H. pylori is the central cause of chronic gastric infection. It is
estimated that 50% of the world population is infected with H. pylori, with a higher prevalence
in the developing countries. H pylori infection has been linked to carcinogenesis and also to
toll-like receptors activation via lipopolysaccharides, leading to metabolic changes and insulin
resistance [2].
Objective
This study aimed to realise a review of studies examining the association between the
presence of H pylori infection and gestational diabetes and obesity.
Materials and Methods
To realise the study, we questioned the PubMed database (December 10th, 2019) using the
keywords “gestational diabetes and Helicobacter Pylori” obtaining four articles and by
“Helicobacter Pylori and gestational weight gain” receiving three reports.
Results and Discussion
It is known and proved the relationship between H pylori infection and extra-gastric diseases
frequently encountered during the last decade in pregnancy, like iron deficiency anaemia,
thrombocytopenia, fetal malformations, miscarriage, preeclampsia, gestational diabetes and
fetal growth restriction [15].
Helicobacter pylori infection and gestational diabetes

In the first study regarding gestational diabetes and H pylori infection realised on 2820
pregnant women recruited before 20 weeks’ gestation, from October 2008 to August 2010, was
investigated the association between the development of the significant pregnancy-related
pathological conditions during the late second or third trimester and maternal infection with H
pylori. The conclusion was that GDM was the most common maternal complication (5.7%) and
was significantly higher in H. pylori-positive women [16].
In the second study realised on a systematic literature search for relevant publications on
PubMed, EMBASE, Cochrane Library and also WebScience databases through 17th November
2018, which identified 31 studies with a total of 22845 participants, showed a significant
association of H Pylori infection of pregnant women with preeclampsia, fetal growth restriction,
gestational diabetes, spontaneous abortion and birth defect [17].
Considering the risk of adverse pregnancy complications in pregnant women infected with
H. pylori, we should take into account the screening and treating for H. pylori infection before
pregnancy.
Another study realised in Sweden from 1st July 2017 to 31st January 2018, on 166 pregnant
women infected who were tested for H. pylori IgG antibodies and was analysed the quantitative
39
insulin sensitivity check index and glucose levels. The results suggest that the seroprevalence
of H. pylori IgG was 66% among the study population, and 12% of women were found to have
GDM [2]. There was no difference concerning the demographics, body mass index,
haemoglobin aspects between the GDM group and the other group. This study concludes that
an increased prevalence of H. pylori is a risk factor for the development of GDM [2].
Concerning that H pylori infection is frequently acquired before pregnancy, it is considered
that hormonal and immunological modifications during pregnancy could activate latent H.
pylori with a negative impact not only on maternal health but also on the fetus and also
consequences can be noticed later in life [15].
Table 1. Studies which assess the association between H. pylori infection and GDM
Nr. Description of study Conclusion References
1 Meta-analysis, 22.845 participants. H pylori infection during [17]
Statistically significant association between H pylori pregnancy can raise the risk on
infection with: adverse pregnancy outcomes.
-preeclampsia (OR: 2.51; 95% CI: 1.88‐3.34;
P<0.001),
-fetal growth restriction (OR: 2.28; 95% CI: 1.21‐
4.32; P=0.01),
-gestational diabetes mellitus (OR: 2.03; 95% CI:
1.56‐2.64; P<0.001),
-spontaneous abortion (OR: 1.50; 95% CI: 1.05‐
2.14; P=0.024), and
-birth defect (OR: 1.63; 95% CI: 1.05‐2.54; P=0.03).
Sensitivity analysis showed the significant
association between H pylori infection and low
birthweight (OR: 1.59; 95% CI:1.05‐2.40; P=0.03)
2 Firstly, investigators on pregnancy focused their H. pylori infection may have a [15]
awareness, mainly on the link between hyperemesis role in the pathogenesis of these
gravidarum and H. pylori infection. Following, disorders through different
researchers turned their attention to other mechanisms: depletion of
pregnancy-related disorders, such as iron deficiency micronutrients (iron and
anaemia, thrombocytopenia, fetal growth defects vitamin B12) in the case of
and malformations, miscarriage and, more recently, maternal anaemia and fetal
to preeclampsia. neural tube defects; local and
systemic induction of pro-
inflammatory cytokines release
and oxidative stress in
gastrointestinal disorders and
preeclampsia.
3 GDM was the most common maternal complication GDM was the most common [16]
(5.7%) and the only pregnancy-related disorder with maternal complication (5.7%)
a significantly higher rate of H. pylori-positive and was significantly higher of
women (41.3%) compared with subjects who did not H. pylori-positive women.
develop the disease (27.7%; P<0.001; OR=1.829,
95% CI=1.320-2.533)
4 A cross-sectional study, a total number of 166 This study concludes that an [2]
women were recruited to the study. The median age increased seroprevalence of H.
was 27 years and 26 weeks. 12% of these women pylori is a risk factor for the
had gestational diabetes mellitus, 65.7% of women development of gestational
were H. pylori IgG seropositive. diabetes mellitus.
Helicobacter pylori infection and obesity or weight gain

The 5th study was a prospective cohort study conducted between May 2012 and May 2013
at the antenatal clinic of Kawempe Health Center IV. The study was conducted in Uganda, on
pregnant women that were in the early second and late third trimester of gestation. The study
demonstrated that the rate of gestational gain weight (GWG) of H. pylori-positive at pregnant
40
women was significantly lower than that of H. pylori-negative at pregnant women. One of the
mechanisms through H pylori infection may lead to low GWG is by reducing the production of
ghrelin and increasing the production of gastric leptin. Ghrelin increases appetite and facilitates
fat storage, while leptin reduces hunger and leads to weight loss [18, 19, 20].
Another study conducted in the Netherlands between 2002 and 2006 describes an association
between H pylori-positive and the reduction of total weight gain in women with daily vomiting.
From the total number of pregnant women with H pylori-positive, the women with CagA
positive were more predisposed to experience daily vomiting compared with women who were
H pylori-positive, but CagA negative. It was not observed any link between H. pylori and fetal
sex, mother’s level of education, parity, smoking and body mass index [21].
Li-Wei Chen, the author of the following study, aims to evaluate the association between H
pylori infection and overweight/obesity. The study was performed from March 2014 to
November 2016 in the north western region of Taiwan. The association between H. pylori and
overweight/obesity is multifactorial, most of the subjects have more than 50 years old, were
male or they had a higher prevalence of insulin resistance or diabetic mellitus, dyslipidaemia
or metabolic syndrome [22, 23].
The results of another study shown that H pylori infection is not associated with obesity or
weight gain. Prevalence of obesity was 19.9% at the patients with H. pylori-positive and 19.1%
at the patients with .H pylori negative. Compared with the H. pylori-negative group, the subjects
in the H. pylori-positive group lost weight more efficiently [24].
The last study contains data collected from 10 European countries, Japan, the USA and
Australia. This study demonstrated that the prevalence of gastric H. pylori colonisation in
various countries is inversely related to the incidence of obesity. The rate of obesity or
overweight were inversely and significantly correlated with the prevalence of H pylori infection
[25].
Table 2. Studies which assess the association between H. pylori infection, obesity and weight gain
Nr. Description of study Conclusion References
1 A prospective cohort study, demonstrate that the H. pylori infection was found to [18]
mean ± sd rate of GWG of H. pylori-positive be independently associated
pregnant women (288.2±81.8 grams/week) was with low rates of gestation
significantly lower than that of H. pylori-negative weight gain (P=0.006).
pregnant women (317.0±74.1 grams/week),
P=0.013.
2 An observational retrospective study, from a total H. pylori infection was not [21]
of 3039 subjects, 12.8% were obese. The associated with
prevalence of H. pylori infection was 53.9% in the overweight/obesity.
obese subjects. The change of BMI in the H. pylori
(+) group was not significantly greater than that in
the H. pylori (-) group after adjustment for
potential confounding factors [RR = 0.988,
95%CI: 0.924-1.057, P=0.729]. The prevalence of
obesity decreased by 1.1% in the H. pylori (+)
group and 0.5% in the H. pylori (-) group.
3 Forty-nine studies with data from 10 European The rate of obesity and [22]
countries, Japan, the US and Australia were overweight were inversely and
identified. The mean H. pylori rate was 44.1% significantly (r=0.29, P<0.001)
(range 17-75%), the mean rates for obesity and correlated with the prevalence
overweight were 46.6 (±16) % and 14.2 (±8.9) %. of H. pylori infection.
41
Conclusion
Recently, the spectrum of H. pylori-related diseases has expanded to include

gastroesophageal reflux disease and obesity. The parallel epidemic of gastroesophageal reflux
disease and obesity combined with decreasing H pylori infection in western countries suggests
HP might play a protective role in the development of obesity and gastroesophageal reflux
disease [26].
Blaser and Atherton proposed that H. pylori infection might influence food intake and calorie
homeostasis through defective signalling of appetite-related hormone in the stomach.
A possible inverse relationship between exposure to H pylori and the occurrence of obesity
has been further addressed in studies of morbid obesity. Preliminary data by histological
examinations have shown that the infection rate of H. pylori in these patients ranged from 24%
to 38%. However, controversial results existed, and whether H. pylori infection plays a role in
childhood and adult adiposity remains to be determined [27].
Some biological mechanisms can explain the association between H pylori infection and
insulin resistance. First, changes in glucose metabolism might lead to chemical alterations in
the gastric mucosa that facilitates the detection of H. pylori infection. A second explanation is
an increase in the levels of the proinflammatory cytokines in response to H. pylori gastric
infection that results in structural alterations to the insulin receptors inhibiting their interaction
with insulin [2]. Placenta tissue from pregnant women with gestational diabetes needs a more
detailed examination [28].
A few studies demonstrated that the elimination of H pylori infection improved glucose
metabolism in patients with type 2 diabetes. Still, the association between H pylori infection
and insulin resistance remains controversial.
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43
The Relationship Between Oral Health and Quality of Life in

Romanian Diabetic Patients Hospitalized in a Tertiary
Antidiabetic Center
PANTEA STOIAN Anca1, STOICA Roxana Adriana1,
ȘTEFAN Simona Diana1,2, PITURU Silviu1, ISTRATI Ionela1,
SERAFINCEANU Cristian1,2
2 “Prof. N.C. Paulescu” National Institute of Diabetes, Nutrition and Metabolic Diseases, Bucharest (ROMANIA)
Emails: ancastoian@yahoo.com, roxana88stoica@gmail.com, simona_ds2002@yahoo.com, piturus@yahoo.com,
ionelaistrati@gmail.com, criserafinceanu@gmail.com
Abstract
Introduction
Quality of life (QuoL) in diabetic patients is generally decreased. There are numerous causes
for that, including chronic illness, anxiety regarding adaptation, social consequences, the time
needed for a diagnosis and the risk of complications. Diabetic patients also have several oral
conditions that can alter their QuoL.
Material and Methods

This were a single-centre, cross-sectional study. We selected 100 patients between January
2019 and June 2019 that were hospitalized at the National Institute of Diabetes, Nutrition and
Metabolic Diseases “Prof. N.C. Paulescu”, in Bucharest. The purpose of our study was to assess
the oral health status and the QuoL in patients with diabetes, for a better understanding of the
factors involved. We used a questionnaire adapted for the Romanian population, and we
collected data from the medical files.
Results
The mean age was 66.4±9.0 years; 71% of patients had type 2 diabetes, and 29% had type 1
diabetes; mean HbA1c was 9.5±2.6%. A high percentage of patients had frequent xerostomia
(40%), burning sensation of the oral cavity (20%) or altered taste perception (20%).
Spontaneous dental bleeding was present in 14% of patients while as 25% had frequent
halitosis. Moreover, 52% of patients were forced to change the consistency of the food, due to
the tooth or gum pain or partial/complete edentation. All of these factors contribute to a poor
QuoL. The diabetic patient is often affected by periodontal disease, 41% of patients enrolled in
our study lost their teeth due to mobility, due to the periodontal support: ligaments, alveolar
bone and dental cement. In 55% of cases, tooth loss had a mixed ethiology, being affected by
the caries processes. There were no statistically significant differences in QuoL among patients
with partial and those with complete edentation.
Conclusion
Patients with diabetes often have symptoms suggestive of poor oral health (xerostomia,
dysgeusia, halitosis and/or spontaneous bleeding). Visits to the dentist are infrequent, despite
the presence of partial edentations, which affect the mastication process in over 60% of patients.
We suggest that better education of these patients is required.
Keywords: oral health, diabetes, quality of life
44
Introduction
Quality of life (QuoL) is an individual's perception regarding their standard of life, in the
context of the culture, communities and value systems in which they live and concerning their
goals, expectations, standards and concerns. [1]. QuoL has more dimensions: psycho-
emotional, physical and material well-being, personal affirmation, social integration and
independence while respecting fundamental human rights. It is well established that QuoL in
diabetic patients is generally decreased. There are numerous causes: like chronic illness, the
anxiety regarding the adaptation and the social consequences, also the time needed to make a
diagnosis and the risk of complications [2].
According to the World Health Organization, oral health is an indicator of overall health,
well-being and QuoL [3]. Oral health is influenced by general health. It shares numerous risk
factors with non-communicable diseases NCDs (cardiovascular diseases, cancer, chronic
respiratory diseases and diabetes), tobacco use, alcohol consumption and unhealthy diets high
in free sugars [3]. NCDs are increasing worldwide.
In 2019, the International Diabetes Federation estimated that around 463 million adults
worldwide have diabetes [4], Type 2 diabetes (T2DM) remaining one of the most frequent
NCDs. For Romania, the results of the PREDATORR epidemiological study estimated that the
prevalence of diabetes in adults is 11.6%, while that of overweight and obesity is 66% [5].
In addition to vascular complications, diabetic patients are suffering from oral conditions
that can affect QuoL and oral health. Gingivitis, oral candidiasis, dysgeusia, local burning
sensation, xerostomia, mucormycosis, flat lichen, angular cheilitis are most commonly seen in
patients with diabetes mellitus [6, 7].
Periodontitis or inflammation of periodontal tissue is 3 times more common in diabetics [7].
The mechanisms by which diabetes mellitus determines periodontitis are the decreased
capacity for periodontal tissue renewal and inadequate local immune response due to the
increased production of proinflammatory cytokines [8].
This was a single-centre, cross-sectional study. We selected 100 patients previously known
with type 1 diabetes mellitus (T1DM) or T2DM between January 2019 to June 2019 that were
hospitalized in the Nephrology or Diabetology Department at the National Institute of Diabetes,
Nutrition and Metabolic Diseases “Prof. N.C. Paulescu”, in Bucharest. For diagnosis and study
inclusion, we used the criteria of the American Diabetes Association 2019 guidelines [9].
The purpose of our study was to assess the oral health status and the quality of life in patients
with diabetes, for a better understanding of the involving factors. We used a questionnaire
adapted for the Romanian population, and we collected data from the medical files. All patients
signed an informed consent form. Clinical and laboratory data (age, gender, weight, height,
BMI, HbA1c, glycemia and haemoglobin) were collected from medical files.
Statistical analyses were carried out using SPSS® version 20.0 software. A p<0.05 was
considered significant. We used mean ± standard deviation to describe continuous variables
with a normal distribution and median for skewed variables. The variables were tested for
normal distribution. For group comparisons, we used paired Student’s t-test, chi-square or
Mann-Whitney U test depending on the Gaussian distribution.
45
We included 100 patients in our study, 71% of patients had T2DM, and 29% had T1DM,
with a mean age of 66.4±9.0 years. Patients had a relatively long duration of diabetes (14.5±8.5
years). The clinical characteristics of the study population are presented in Table 1.
The mean HbA1c showed poor glycaemic control (9.5±2.6%) and the patients were
hospitalized in a tertiary diabetes center to improve metabolic status.
Table 1. Clinical characteristics of the study population

Variable Value
BMI (kg/m2) 30.5±6.6

Age (years) 66.4±9.0
Diabetes duration (years) 14.5±8.5
SBP (mmHg) 147.8±25.92
DBP (mmHg) 86.16±13.89
Glycemia (mg/dL) 242±100
HbA1c (%) 9.5±2.6
PCR (mg/L) 13.3±16.58
VSH (mm/h) 19.4±11.81
Fibrinogen (mg/dL) 465±91
WBC (*1000/μL) 7.78±2.28
Neutrophil count (*1000/μL) 4.91±2.09
Legend: BMI=body mass index, SBP=systolic blood pressure, DBP=diastolic blood pressure, PCR=protein C-
reactive, VSH=erythrocyte sedimentation rate, WBC=white cells count.
A high percentage of patients had frequent xerostomia (40%), burning sensation of the oral
cavity (20%) or altered taste perception (20%). Spontaneous dental bleeding was present in
14% of patients while as 25% had frequent halitosis. All of these symptoms are associated with
periodontal disease and support the results of epidemiological studies that have a higher
frequency of this pathology in patients with diabetes [10, 11].
Most patients (64%) were treated with insulin (Fig. 1). Also, the majority of patients had
distal sensory diabetic polyneuropathy (77%), diabetic nephropathy (57%) or proliferative
diabetic retinopathy (42%), as shown in Fig. 2. The presence of these complications is an
additional argument for the reduced adherence to the medical, nutritional intervention, in
contrast with the self-reported grade for adherence to diet (Fig. 3). Besides, some studies have
shown an association between periodontal disease and the prevalence of stroke, nephropathy,
or heart failure [12].
46
Fig. 1. Distribution of patients according to the treatment
Fig. 2. The presence of diabetes complications
Polyneuropathy Nephropathy Retinopathy
Fig. 3. Distribution of patients according to diet adherence (Grade 4 to 10)
Moreover, 52% of patients were forced to change the consistency of the food, due to the
tooth or gums pain or partial/complete edentation. All of these indicate the poor quality of life
of diabetic patients (Fig. 4 and 5).
47
Fig. 4. The distribution of patients according to the perception of the tooth and gingival pain (dark orange)
and the need to interrupt the meals (light orange)
Fig. 5. The level of mastication, speech (yellow) and the need to change the consistency of food (orange)
The use of dental services was low, as shown in Figure 6; most patients (72%) had their last
visit to the dentist more than a year ago. Some of them have not been to a dentist for more than
10 years, the leading cause being – difficulty to attend (31%) or fear (28%). A total of 38% of
patients fulfilled the brushing of the oral cavity or the dental prosthesis 1-2x/week, while 8%
of them – had abandoned this habit. The level of education (25% of the patients had a low level)
probably influences this behaviour.
The results of our research are comparable to those of other populations where patients with
diabetes were also reported to have poor oral health [13]. People in developed countries had
better status than those with lower incomes [14].
Fig. 6. Distribution of patients according to the last visit to the dentist
<1 month 1-6 months 6-12 months >12 months
The diabetic patient is often affected by periodontal disease, 41% of patients enrolled in our
study lost their teeth due to mobility, due to the periodontal support: ligaments, alveolar bone
48
and dental cement. In 55% of cases, tooth loss had a mixed ethiology, being affected by the
caries processes. The Mann Whitney U test revealed no statistically significant differences in
QuoL between patients with partial and those with complete edentation.
Considering that diabetic patients have a lower QuoL when compared with the general
population [15], other factors may be more important than the presence of edentation. Also,
some of the patients had a dental prosthesis which could influence their perception of the QuoL.
In correlation with the number of daily brushing performed and the weak control of diabetes,
we observed that the prevalence of the bacterial plaque and the dental tartar was 78% in our
patients.
An equal number of patients had cavities and non-cavities lesions (atria, abrasion, dental
erosions); 65% of patients had the mobility of the remaining teeth, while 75% of them had
gingival retractions, conditions caused by the evolution of the periodontal disease. This
confirms the results of the study published by Costa et al., where poor control of diabetes was
associated with the faster progression of periodontitis and edentation [16].
Fig. 7 shows that 26% of the patients have total edentations of the jaw, 12% of these patients
having a total acrylic prosthesis in the upper jaw.
A percentage of 12% of the people with diabetes enrolled, had mandibular edentulism, and
almost all of them (11%), wore total acrylic prostheses. 22% of the patients, were completely
bimaxillary edentate, and only 19% of them were protected, the other 3% refused to wear them,
accusing an unpleasant sensation.
Fig. 7. Patient distribution according to the presence of the total edentation (dark orange)
and its prosthesis (light orange)
Regarding the relationship between QuoL, oral health and the complications of diabetes
mellitus, the symptoms of diabetic distal sensory neuropathy are considered more significant
(the mean of self-reported grades by patients was 7.17 as opposed to the actual grades 6.58,
p<0.05) than the presence of proliferative diabetic retinopathy or the diabetic nephropathy.
We also analysed the group of patients with complete edentation (n=48) against the others
(n=52). No significant differences were obtained in the above parameters or connection with
QuoL. Regarding other life and work conditions that might interfere, the mean value of the
systolic and diastolic blood pressure was statistically significantly different between the
smoking patients and the non-smoking ones but not QuoL. Alcohol consumption influenced
BMI but not the other factors related to oral health. We have not obtained significant
correlations between the extent which the dietary restrictions were followed and the other
laboratory data, or between the frequency of daily dental brushing and the other data. We would
like to mention that the mean number of daily brushings was different, based on the education
level, but not based on professional status. No differences existed between male and female
genders.
49
Conclusions
Patients with diabetes experience many symptoms suggestive of poor oral health
(xerostomia, dysgeusia, halitosis and/or spontaneous bleeding). Visits to a dentist are low
despite the presence of partial edentations, which affect the mastication process in over 60% of
patients. We suggest that better education of the patients is required. Among diabetes
complications, distal sensory diabetic polyneuropathy has the highest impact on QuoL, most
likely through secondary difficulties with walking.
Contributions
All the authors had equal contributions to this paper. The authors declare no conflict of
interest.
REFERENCES
1. https://www.who.int/healthinfo/survey/whoqol-qualityoflife/en/, Accessed: 2020-01-11

2. Poudel P, Griffiths R, Wong VW, et al., (2018). Oral health knowledge, attitudes and care practices of
people with diabetes: a systematic review. BMC Public Health.;18(1), p. 577.
3. https://www.who.int/health-topics/oral-health/#tab=tab_1; Accessed: 2020-01-11
4. International Diabetes Federation, IDF Diabetes Atlas, 9th Edition, 2019, Available from
http://www.diabetesatlas.org/, Accessed: 2020-01-17
5. Mota M, Popa SG, Mota E, et al., (2016). Prevalence of diabetes mellitus and prediabetes in the adult
Romanian population: PREDATORR study. J Diabetes. 8(3), pp. 336-44.
6. Sischo L, Broder HL (2011). Oral Health-related Quality of Life – What, Why, How, and Future
Implications. J Dent Res, 90(11), pp. 1264-1970.
7. Kudiyirickal MG, Pappachan JM (2014). Diabetes mellitus and oral health. Endocrine, 49(1), pp. 27-34.
8. Weinspach K, Staufenbiel I, Memenga-Nicksch S, et al., (2013). Level of information about the
relationship between diabetes mellitus and periodontitis – results from a nationwide diabetes information
program. Eur J Med Res.18(6) p. 33.
9. American Diabetes Association (2019). Standards of care-Diagnosis and Classification of Diabetes
Mellitus. Diabetes Care 42(Supplement 1), pp. S13-S28.
10. Saremi A, Nelson RG, Tulloch-Reid R et al., (2005). Periodontal disease and mortality in type 2 diabetes.
Diabetes Care 28, pp. 27-32.
11. Moore PA, Weyant RJ, Mongelluzzo MB et al., (1999). Type 1 diabetes mellitus and oral health:
assessment of periodontal disease. J. Periodontol.70, pp. 409-417.
12. King GL (2008). The role of inflammatory cytokines in diabetes and its complications. J. Periodontol,
79(8 Suppl.), pp. 1527-1534.
13. Allen EM, Ziada HM, O’Halloran D, Clerehugh V, Allen PF (2008). Attitudes, awareness and oral health-
related quality of life in patients with diabetes. J Oral Rehabil. 35(3), pp. 218-223.
14. Moore PA, Orchard T, Guggenheimer J, Weyant RJ (2000). Diabetes and oral health promotion: a survey
of disease prevention behaviours. J Am Dent Assoc. 131, pp. 1333-1341.
15. Rubin RR, Peyrot M (1999). Quality of life and diabetes. Diabetes Metab Res Rev, 15(3)
16. Costa FO, Miranda Cota LO, Pereira Lages EJ et al., (2013). Progression of periodontitis and tooth loss
associated with glycaemic control in individuals undergoing periodontal maintenance therapy: a 5-year
follow-up study. J Periodontol 84, pp. 595-605.
50
Metformin Treatment During Pregnancy and Its Effect on Fetal

Growth: A Literature Review
IONESCU Crîngu Antoniu1,2, ROȘU George-Alexandru1,

ZYGOUROPOULOS Nikolaos1, MĂDAN Victor2,3, SOCEA Bogdan2,4,
JACOTĂ-ALEXE Florentina1, GHEORGHIU Diana Claudia1,
GHEORGHIU Nicolae2,6, HARADJA Horațiu1, CAIMACAN Adriana1,
POPESCU (BĂNACU) Ina1,2, BĂNACU Mihail1,2, CĂLIN Florin Daniel1,2,
DIMITRIU Mihai1,2
1 Department of Obstetrics & Gynaecology, “St. Pantelimon” Clinical Emergency Hospital, Bucharest (ROMANIA)
3 Department of Urology, “Dr Carol Davila” Central Military Emergency University Hospital, Bucharest (ROMANIA)
4 Department of General Surgery, “Sf. Pantelimon” Clinical Emergency Hospital, Bucharest (ROMANIA)
5 “Maria Sklodowska Curie” Children Clinical Emergency Hospital, Bucharest (ROMANIA)
6 Department of Orthopaedics and Traumatology, Elias Emergency University Hospital, Bucharest (ROMANIA)
Emails: antoniuginec@yahoo.com, george.rosu@ymail.com, nz@zitaquality.ro, victmad@gmail.com,

bogdansocea@gmail.com, falexe.jakota@gmail.com, cdgheorghiu@hotmail.com, ngheorghiu@hotmail.com,
hharadja@gmail.com, adriana.caimacan@gmail.com, ina.popescu@yahoo.com, mbanacu@gmail.com, dkcalin@gmail.com,
drmihaidimitriu@yahoo.com
Abstract
The prevalence of pregnant women with gestational diabetes mellitus or type 2 diabetes has
increased in the last two decades. This has led to a higher frequency of maternal and fetal
complications associated with this comorbidity that could put at vital risk the mother and the
fetus. The treatment of choice for this pathology in pregnancy is insulin, due to its lack of
significant transplacental passage and safety in pregnancy confirmed by the long history of use.
A few randomised controlled trials have investigated metformin, a biguanide that is used as
first line of treatment for type 2 diabetes mellitus in case of nonpregnant patients, in the
treatment of gestational diabetes, type 2 diabetes mellitus associated with pregnancy and for
reducing the insulin resistance in obese pregnant patients and pregnant patients with polycystic
ovary syndrome. Due to is the free transplacental passage, metformin could have effects of the
fetal side, although the risk of teratogenicity is low. In this study, we analysed the effect of
metformin on fetal growth and birth weight reported in 9 randomised controlled trials that were
published in the past ten years. The nine randomised controlled trials included several 1574
pregnant women that took metformin treatment during pregnancy for gestational diabetes, type
2 diabetes and to reduce insulin resistance in the case of obese pregnant women and pregnant
women with polycystic ovary syndrome. This study concludes that metformin has reduced or
no effect on fetal growth and birth weight compared with standard insulin treatment in diabetes
mellitus or placebo in the cases of pregnant women with obesity or previous polycystic ovary
syndrome without diabetes.
Keywords: diabetes mellitus, pregnancy, metformin, PCOS, obesity, fetal growth
Introduction
The number of pregnant women with gestational and type 2 pregestational diabetes mellitus
(DM) has increased in the last 20 years [1, 2]. This medical comorbidity of pregnancy is placing
the mother and fetus at risk for several antenatal, intrapartum and neonatal complications,
51
especially in cases with poor glycaemic control during pregnancy [3-6]. The most frequent
complications identified are preeclampsia, fetal macrosomia, need for caesarean section and
neonatal hypoglycaemia [7]. Treatment of DM in pregnancy reduces severe perinatal morbidity,
improves women’s health and decreases maternal and neonatal complications [8, 9, 10, 11].
Insulin is the only option for treating DM associated with pregnancy, because of its lack of
significant transplacental passage, long history of use and safety in pregnancy [12-14].
For type 2 DM (T2DM) not associated with pregnancy, the first line of treatment is
metformin [15]. Metformin is a biguanide that improves peripheral insulin sensitivity and
reduces hepatic gluconeogenesis, fasting insulin and testosterone levels [16, 17]. Metformin
has been studied in several small randomised trials as an alternative medication to insulin in
patients with gestational DM [18-24], and it has also been used in pregnant patients with
polycystic ovary syndrome (PCOS) [25]. However, metformin is not yet accepted as a treatment
for gestational DM and T2DM in pregnancy [26-28], primarily because of the small number of
patients included in the randomised controlled trials conducted until recently [17, 28-30]. Those
randomised controlled trials have also analysed, as primary or secondary outcomes, the effect
of metformin on the foetuses, since metformin crosses the placental barrier [31]. Nowadays,
metformin is categorised by the food and drug administration (FDA) regarding its safety for
use in pregnancy [32].
We conducted a literature review to evaluate the effect of metformin treatment during

pregnancy on fetal growth and birth weight compared with insulin, in the case of DM, and with
placebo in the trial of obese patients or patients with PCOS.
The search was performed in Medline and Embase databases. The search was restricted by
language, searching only for full-text articles in English. We searched by free text search terms
as “diabetes mellitus” and “pregnancy” and “metformin”; “metformin treatment” and
“pregnancy”; “metformin” and “obese” and “pregnancy”; “metformin” and “polycystic ovary
syndrome” and “pregnancy”. We only included randomised controlled trials whose results were
published between January 1, 2010 and January 5, 2020. The outcomes that we searched for
were the birth weight, the number or percentage of small for gestational age and large for
gestational age neonates in the metformin groups and insulin or placebo groups.
Using our search protocol, we initially identified 63 randomised controlled trials (RCT).
After we applied the inclusion criteria, 9 RCT that analysed as a primary or secondary
outcome the effect of metformin treatment during pregnancy on fetal growth and fetal birth
weight were identified. For each study, we also reported the primary outcome, when this was
not the effect of metformin on birth weight. A total of 1574 patients were included in those
studies.
Three randomised controlled trials [33-35] analysed the effect of metformin treatment on
fetal growth and birth weight in pregnant patients with gestational DM. A study conducted by
Ainuddin (2015b) [33] compared treatment with metformin alone or metformin plus insulin or
insulin alone in women with gestational DM. A total of 150 gestational diabetic patients were
included in their prospective randomised control open labelled study [33]. They showed that
mean birth weight was significantly lower in metformin-treated groups (3.4±0.4 kg – metformin
alone vs 3.3±0.5 kg – metformin + insulin – vs 3.7±0.5 kg – insulin alone, p<0.01) [33]. The
conclusion was that metformin in an effective and cheap treatment option for women with
gestational DM with or without supplemented insulin, being generally well tolerated and
52
accepted by all women participating in the study and having advantages of less maternal weight
gain, no risk of maternal hypoglycaemia, with fewer neonatal complications and with less
neonatal hypoglycaemia and neonatal intensive care unit (NICU) admissions [33].
Another randomised controlled trial study on gestational DM treated with metformin
compared with insulin, conducted by Spaulonci [34], evaluated glycaemic control in women
receiving metformin or insulin for gestational DM and the predictors for supplemental insulin
treatment in women initially treated with metformin. They included 94 patients with gestational
DM [34]. The study showed that metformin treatment during pregnancy does not influence new
born weight (3143.7±446.6 vs 3237,6±586 g; p=0.39) [34]. There were no foetuses with
macrosomia in the metformin group compared with 3 cases (6.5%) in the insulin group
(p=0.242) [34]. In the group receiving metformin, 6 neonates (13%) were small for gestational
age (SGA), 38 (82.6%) were adequate for gestational age (AGA) and 2 (4.3%) were large for
gestational age (LGA) [31]. In the group receiving insulin 4 neonates (8.7%) were SGA, 39
(84.8%) were AGA and 3 (6.5%) were LGA (p=0.735) [34]. The conclusion was that metformin
provided adequate glycaemic control with lower mean glucose level throughout the day, less
maternal weight gain and lower frequency of neonatal hypoglycaemia [34].
A third study examined the in vivo placental transfer of metformin in patients with
gestational DM treated with metformin and its effect on maternal glycaemic control and weight
gain. The primary outcome was the birth weight of new-borns from diabetic mothers treated
with metformin [35]. The study conducted by Tertti (2014) included 217 pregnant patients
randomly allocated to one of the study arms: metformin treatment and insulin treatment [35].
They concluded that between the metformin group and insulin-treated group were no
differences regarding the fetal growth and birth weight (3556±430 g – metformin group vs
3588±447 g; p=0.93) [35].
One randomised controlled trial study conducted by Ainuddin et al., (2015a) assessed the
effect of metformin treatment compared with insulin in pregnant patients with T2DM regarding
the perinatal outcome, maternal complications, additional insulin requirement and treatment
acceptability [26]. Patients (n=206) with T2DM in pregnancy were selected [26].
The mean birth weight represented one of the neonatal outcomes the that was analysed; this
parameter did not differ significantly in all the compared groups (metformin alone, metformin
+ insulin, insulin alone – p=0,345) [26]. However, significantly smaller for gestational age
babies were found in metformin-treated groups compared with the insulin alone group (31.2 vs
14.4 vs 2%; p<0.01) [26]. The incidence of macrosomia was similar in the 3 groups (12.5 vs
33.3 vs 27%; p=0.28) [26]. The main conclusion of this study was that metformin alone or with
add-on insulin is an effective, safe and cheap treatment option for patients with T2DM in
pregnancy [26].
Two studies conducted by Hickman et al. (2012) and Ibrahim et al., (2013) on the
effectiveness of metformin treatment on pregnant women with DM, T2DM or gestational DM,
concluded that metformin alone or with insulin compared with insulin alone does not affect the
fetal growth and development and the birth weight [36, 37]. Hickman’s study concluded that
metformin is a safe, well-tolerated treatment for the control of overt T2DM and early gestational
DM in pregnancy and that is preferred over conventional insulin therapy by the patient [36].
Ibrahim’s study found no significant difference between the 2 groups analysed (insulin +
metformin vs insulin alone) in diabetic pregnant women with insulin resistance, regarding the
fetal macrosomia (10 out of 43 vs 12 out of 39; p=0.443) [34]. In this study, only 1 congenital
malformation was identified in the metformin group – a case of cleft lip [37]. This study
conclusion was that adding metformin to insulin therapy in women with insulin-resistant DM
in pregnancy seems to be effective for appropriate glycaemic control and is associated with
better neonatal outcomes (less frequent neonatal hypoglycaemia, NICU admission and neonatal
53
respiratory distress syndrome) [37]. The total number of patients included in these 2 studies
was 118 pregnant women with DM [36-38].
Since metformin treatment in obese patients decreases insulin resistance, 2 studies
investigated the effect that metformin compared with placebo in obese pregnant women without
DM [39-41]. The studies conducted by Syngelaki with Nicolaides et al., (2016) and Chiswick
et al., (2015) included 659 patients with body mass index (BMI) of ≥35 kg/m2, respectively 30
kg/m2 or more [39, 40]. The patients received 2.5 to 3 g of metformin/day from 12-18 weeks of
gestation until delivery [39, 40]. The Syngelaki’s study showed that in pregnant women without
DM who had a BMI of ≥35 kg/m2, metformin did not reduce the median neonatal birth weight
Z score or the incidence of large for gestational age neonates (p=0.66), but was associated with
less maternal gestational weight gain compared with placebo (4.6 vs 6.3 kg; p<0.001) [39]. The
study conducted by Chiswick – the EMPOWaR study – concluded that metformin, compared
with placebo, has no significant effect on birth weight percentile in obese pregnant women (Z
score of birth weight percentile 0.2462 vs 0.2680; p=076) [40]. Also, their results challenge the
Pedersen hypothesis [42] which state that maternal hyperglycaemia drives fetal
hyperglycaemia, and hence fetal hyperinsulinemia and fetal overgrowth, providing the first
experimental evidence that factors other than maternal glucose are important in fetal
overgrowth. Ultrasound estimates of fetal growth and stillbirth are challenging [43, 44]. Early
trimester markers and myelofibrosis are not influenced [45, 46].
Only 1 randomised controlled study analysed the effect of metformin treatment on
pregnancy outcomes in women with PCOS that took metformin from the first trimester to
delivery to reduce the higher incidence of preeclampsia, gestational DM and preterm delivery
[42]. The study by Vanky et al., (2010) conducted included 257 pregnant patients randomly
assigned to metformin or placebo [45]. One of the secondary outcomes of Vanky’s study was
the effect of metformin on fetal growth [47]. The study group found no difference in the birth
weight between the metformin and the placebo group (p=0.32). Nevertheless, significantly
more neonates with macrosomia were found in the placebo group compared with the metformin
group (7 out of 130 vs 2 out 127 patients).
Conclusions
The majority of the randomised controlled studies published in the last 10 years demonstrate
that metformin is a safe and effective treatment during pregnancy. It decreases insulin resistance
and improves glycaemic control in pregnant women with DM, PCOS or obesity. It also has a
reduced or no effect on fetal growth and birth weight compared with standard insulin treatment
in DM or placebo in pregnant women with obesity or previous PCOS without DM.
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56
Plant-Based Diets and Type 2 Diabetes Prevention

and Management
HANCU Anca Mihaela1, SERAFINCEANUCristian2, RIZZO Manfredi3,4,

PAPANAS Nikolaos5, PANTEA STOIAN Anca2
1 Nutriscience Clinic, “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
2 Department of Diabetes, Nutrition and Metabolic Diseases, “Carol Davila” University of Medicine and Pharmacy,
Bucharest (ROMANIA)
3 Biomedical Department of Internal Medicine and Medical Specialties, School of Medicine, University of Palermo, Palermo
(ITALY)
4 Division of Endocrinology, Diabetes and Metabolism, School of Medicine Columbia, University of South Carolina South
Carolina (USA)
5 Second Department of Internal Medicine, Diabetes Centre, Clinic of Diabetic Foot, University Hospital of Alexandroupolis,
Democritus University of Thrace (GREECE)

Emails: ancahancu@gmail.com, criserafinceanu@gmail.com, manfredi.rizzo@unipa.it, papanasnikos@yahoo.gr,
ancastoian@yahoo.com
Abstract
A plant-based diet (PBD) emphasizing intake of food from plant origin, mainly based on
whole grains, fruits, vegetables, nuts, seeds and legumes is excluding animal origin foods.
This eating model excludes dairy, poultry, red meat and fish. How this eating model could
bring benefits in prevention or management of type 2 diabetes (T2D) and what
recommendations are in new guidelines will be detailed in this article. It could be possible that
a plant-based diet could modulate glucose metabolism and insulin levels, offering protection
against T2D. Specific components like high fiber intake, monounsaturated acids (MUFA’s) and
polyunsaturated acids (PUFA) definitely could play a role.
Meanwhile, micronutrients with anti-inflammatory and antioxidant activity will bring
additional benefits, together with reduced intake of nitrites and nitrates. The explanation for
how this diet could improve glycaemic control and promoting insulin sensitivity is through
weight and adiposity control, reduced inflammation and oxidation. How the microbiome is
influenced by diet is already proved, and a plant-based diet could build the potential for healthy
gut microbiota. The American Diabetes Association Standards 2020 recognize benefits for PBD
in the management of T2D, including this model in recommended medical nutrition therapy for
these patients.
Keywords: Mediterranean diet, plant-based diets, nephropathy, retinopathy, neuropathy, lifestyle, medical nutrition therapy
Introduction-Background
The plant-based diet (PBD) emphasizes the consumption of foods from plant origin,
avoiding ultra-processed food and is mainly based on whole grains, fruits, vegetables, nuts,
legumes, seeds consumption excluding food from the animal origin [1]. These types of diets do
not include dairy products, fish, eggs, poultry or red meat. The debate regarding recommending
such a diet in diabetes prevention will be challenging, mainly considering benefits already
proven by dairy products or fish, excluded by such diets, in preventing diabetes [2].
Mozzafarian mentioned as dietary priorities for cardiometabolic health more than 2
servings/week of fish and 2-3 servings/day of dairy.
57
Microbiota balance is influenced by live cultures included in fermented dairy. These cultures
are acting as a protective mechanism against host dysbiosis. Meanwhile, systemic inflammation
present in cardiometabolic diseases will be influenced by a healthy gut.
Bioactive metabolites, peptides, exo-polysaccharides, released by lactic acid bacteria LAB
could exert cardiometabolic and anti-inflammatory effects [3].
Could a PBD be considered as a recommendation in type 2 diabetes (T2D)? Is there enough
evidence-base? Could T2D be prevented or managed better by adopting PBD? Should this type
of diet be prioritized vs other diets, or just included among other recommendations?
We realized our research on PubMed. Articles were selected based on the significance and
the year of publication, considering subjects as PBDs, Mediterranean diet, T2D, comorbidities,
inflammation and insulin resistance. The new American Diabetes Association (ADA) Standards
2020 [4-6] have been taken into consideration as the main guide.
PBD and T2D incidence and management – The Adventist Health Study (AHS) (2002-
2007), an observational study, investigated the relationship between vegetarian diets and T2D
incidence, by multiple logistic regression analysis. The AHS, which included 41387
individuals, showed variable T2D incidence according to diet: Lacto-Ovo vegetarian compared
with non-vegetarian 0.618 (0.503-0.760). Vegan vs non-vegetarian 0.381 (0.236-0.617). Semi-
vegetarian vs non-vegetarian 0.486 (0.312-0.755). Pesco-vegetarian vs non-vegetarian 0.790
(0.575-1.086). In conclusion, a substantial reduction in T2D incidence is associated with
vegetarian diets [7]. Recently, in another observational study (2007-2009) including 4384
Taiwanese Buddhist participants, the complete avoidance of fish and meat, in Taiwanese
vegetarian diet was inversely correlated with T2D and impaired fasting glucose (IFG)
prevalence in men and postmenopausal women. The Taiwanese diet was completed by alcohol
and tobacco avoidance. The prevalence of T2D in different groups was: 4,3 vs 8,1% for men;
0,6% vs 2,3% for premenopausal women and 2,8% vs 10% in menopausal women.
This vegetarian diet was demonstrated to be negatively associated with T2D in men (OR for
T2D: 0.49, 95% CI: 0.28-0.89); in premenopausal women (OR for T2D: 0.26, 95% CI: 0.06-
1.21); and in menopausal women (OR for T2D: 0.25, 95% CI: 0.15-0.42). In T2D, a good
glycaemic control achieved by vegetarian diets pattern is demonstrated by 6 clinical trials that
involved 255 individuals [8]. Five of these were vegan like the other ovo-lacto-vegetarian. The
vegan diet achieved a statistically significant reduction in HbA1c vs control, p<0,01, reduction
of 0,39% vs control diets concluding that a vegan diet could bring benefits for glycaemic
management in T2D patients.
Healthy Lifestyle adoption, mainly by a correct diet, could play an important role in
delaying or improving T2D comorbidities. PBD may support the prevention of microvascular
and macrovascular complications due to intrinsic properties, as PBD are rich in micronutrients
with anti-inflammatory and antioxidant properties. A significant reduction in T2D incidence
from 28 to 58% in different countries has been shown after prevention strategies focused merely
on lifestyle intervention based mainly on diet and physical activity [9-12].
Microvascular Complications
Nephropathy, the microvascular complication appears with a prevalence of 40% at diabetic

patients. Diabetics may be vulnerable at the harmful effect of high protein diet, which could be
associated with glomerular hyperfiltration, leading to higher risk of de novo chronic kidney
disease (CKD) or accelerating pre-existing disease [13]. Traditionally, a low protein diet (LPD)
for the management and prevention of CKD in T2D is defined with a range of 0,6-0,8 g/kg/day
58
proteins [14], consistent with guidelines recommending 10-15% of energy to be taken from
proteins [14] drawing the conclusion that an additional tool in managing CKD diabetic patients
could be plant based diet. Recent evidences from studies are suggesting that a low intake of
animal-based foods and meat could bring benefits for kidney, albuminuria being associated
with animal fat ingestion. Moreover, carnitine and choline, resulted from meat are transformed
by gut microbiome into trimethylamine (TMA) and TMA-N oxide, in a direct relation with
renal fibrosis and atherosclerosis [15]. In the Nurses’ Health Study, saturated fats and red and
processed meats are in line with rapid decline of glomerular filtration rate (GFR), >3
ml/min/1.73m2/year and albuminuria, contrasting with decreased risk of GFR for a dietary
pattern including whole grains, vegetables and fruits. The Atherosclerotic Risk in Communities
Study (ARIC) followed for 23 years 12000 adults, revealing that high intake of nuts, low fat
dairies and legumes has been associated with a lower risk of CKD (HR 0.81, 0.72-0.92) but red
and processed meat ingestion was in line with high CKD risk (HR 1.23, 1.06-1.42) [16]. Not
surprising are results obtained by substitution analysis; one serving of red/processed meat
replaced by a serving of legumes reduced the risk of CKD by 31%. Similar, if replacement was
with whole grains, risk reduction was 21% [17].
The largest observational study, with >63000 individuals [18] followed for 15.5 years
showed a high risk of developing end stage renal disease (ESRD) for those in the highest
quartile of red meat consumption. In this study, named, a 50,4 reduction in ESRD was proven
by the substitution analysis, when 1 serving of red meat was replaced by soy and legumes.
Moreover, issues related to hyperkalaemia or any protein deficiency during PBD are not
supported by the literature. In conclusion, the pleiotropic benefits of PDB, could be considered
useful in CKD, due to reduced serum phosphate level, blood pressure decrease and survival
improvement.
Diabetic retinopathy risk was inversely correlated with plant foods and dietary fibers [19].
It was not identified as a specific study addressing diabetic retinopathy and PBD.
However, a post hoc analysis of the PREvencion con DIeta MEDiterranea (PREDIMED)
showed a 44% reduction of diabetic retinopathy (HR: 0.56; 95% CI: 0.32, 0.97) in the group
with Mediterranean Diet enriched with extra-virgin olive oil group compared with the low-fat
control diet group [20]. Another subgroup analysis showed that Med Diet with nuts added is
beneficial for diabetic patients without baseline hypertension. More studies are needed in order
to explain better how these dietary models could be helpful in the prevention of diabetic
retinopathy.
The primary strategy to prevent diabetic neuropathy, is focused on lifestyle improvement
and optimal glycaemic control [21]. Only one small non-randomized trial showed that
neuropathic pain was attenuated after a nutritional intervention with a vegan diet, no
cholesterol, low fat and high in fiber. Additional studies are needed to deliver future definite
recommendations regarding PBD. Gastroparesis, typical of diabetic neuropathy needs
particular dietary recommendations; a randomized clinical trial has demonstrated that diets with
small size particles could reduce vomiting, bloating, heartburn, regurgitation and postprandial
fullness [22]. A small study (45 patients) identified foods that enhance symptoms in
gastroparesis: peppers, tomato juice, broccoli, oranges and orange juice and generally spicy and
acidic food. Some recommended foods proved to be tolerated without enhancing symptoms are:
graham or saltine crackers, gluten-free foods, pretzels, tea, clear soup, potatoes, apple sauce
and white rice [23] all could be part of a PBD.
Macrovascular Complications – CVD
A 66% lower risk of peripheral arterial disease has been reported for patients with high
adherence to Med Diet (11%) compared with those who did not adhere strictly (score 0-8) in a
59
small study on 432 patients with T2D [24]. The ATTICA study [25] revealed an inverse
association between adherence to Med Diet and 10 years of cardiovascular risk in non-diabetic
participants, but the inverse association seen at diabetic patients was non-significant.
A 29% lower risk for major cardiovascular events in a PREDIMED – subgroup has been
seen compared with diabetic patients following a low-fat diet. However, no effect was seen in
non-diabetic subjects [26].
Diabetes and mortality – Although diabetes is one of the top 10 mortality causes, dietary
models’ impact on diabetes mortality is insufficiently studied. In summary, actual evidence: 4
prospective studies-lower risk of death from ischemic heart disease in vegetarians compared
with non-vegetarians, independently of diabetes diagnosis at baseline [27]; in the Women’s
Health Initiative Study [28] – high adherence to Med Diet was without impact on women with
heart failure, with or without diabetes; but DASH diet adherence was inversely correlated with
low mortality risk for non-diabetic women [28].
A possible mechanism suggested for PBD is through glucose metabolism and insulin
levels, offering protection against T2D. Specific components like high fiber intake,
monounsaturated acids (MUFA’s) and polyunsaturated acids (PUFA’s) could play a role.
Meanwhile, micronutrients with anti-inflammatory and antioxidant activity will bring
additional benefits, together with reduced intake of nitrites and nitrates. The explanation for
how this diet improves glycaemic control and promoting insulin sensitivity is through weight
and adiposity control, reduced inflammation and oxidation. How the microbiome is influenced
by diet is already proved, and a PBD could build the potential for healthy gut microbiota.
Ultra-processed food (UPF) and T2D risk [29] New published results associated UPF
consumption with T2D risk. In a large population-based prospective study, involving 104707
participants from Nutri-Net Sante cohort followed for 10 years, in the period 2009-2019, data
about dietary intake have been obtained through 24h dietary records. Statistically significant
results, with an absolute increment of 10 in UPF percentage in the diet High T2D risk was
associated with the absolute consumption of UPF in g/day. This large prospective observational
study correlated the proportion of UPF with T2D risk. In line with these results, more efforts
should be made in limiting UPF consumption. [29].
Functional Foods
Functional foods could be defined as foods with ingredients biologically active that are
associated with physiological health benefits and are related to the prevention of different
chronic diseases like T2D. The active ingredients that are included in functional foods are
“nutraceuticals” and implies processes of purifying, extracting, assaying and concentrating
these ingredients [30]. Regular intake of functional foods is demonstrated to prevent or delay
T2D and complications. It will improve blood pressure, glycaemic control, antioxidant enzymes
activity, suppressing pro-inflammatory cytokines, maintaining a healthy gut microbiota [31].
Polyphenols present in vegetables, fruits, tree nuts and olive oil seem to confer health
benefits [32, 33]. Functional foods can exert a protective role in T2D prevention as part of
lifestyle intervention. Clinical evidence demonstrated the efficacy of foods rich in lignans-
polyphenols, such as flaxseeds in the reduction of C reactive protein (CRP) levels, glucose and
insulin. The epidemiological evidence is associating total flavonoid intake with T2D risk [34].
Resveratrol, highly present in grape products, reduces insulin secretion and increases glucose
transport [35], leading to essential benefits in the prevention and management of T2D. Coffee,
the highest source of dietary polyphenols, is inversely associated with T2D risk [36]. The group
named polyphenols include flavonoids, phenolic acids, lignans and stilbene [37]. The potential
antioxidant role in the prevention of chronic diseases, often characterized by increased ROS,
including T2D, is evidenced by epidemiological studies [38-41].
60
Planet Sustainability and PBD – The EAT-Lancet Commission, with “HEALTHY

DIETS” from sustainable food systems was recently published [42] Food Planet Health means
a strategy considering dietary shifts, and a transformation until 2050 through a diet for healthy
people and a healthy planet. This diet will emphasize the consumption of vegetables, fruits,
legumes, nuts and will reduce by 50% the consumption of red meat and sugar. This diet rich in
plant-based food will offer health for people and will bring benefits to the environment [42].
In line with the newest released 2020 ADA standards for medical nutrition therapy in
diabetes, it will be essential to maintain a healthy weight and glycaemic, blood pressure and
lipid goals, by promoting healthy eating patterns. The earlier focus on nutrients or single foods
is now replaced now by eating models. Maintaining the pleasure of eating, with attention to the
language used about food choices is also important [5]. These guidelines consider that no ideal
percentage between macronutrients should be specified and the recommendations should start
with the eating model close to actual preferences in order to obtain the best adherence to the
new eating model. All aspects should be taken into consideration when giving advice to a
diabetic patient: religion, health, tradition, beliefs also metabolic goals and total calories [43-
45]. Main aspects that have to be taken into consideration when choosing an eating model are:
to decrease as much as possible refined grains and added sugars, reduce ultra-processed foods,
increase non-starchy vegetables [43].
The ADA 2020 standards for medical nutrition therapy include as examples: The
Mediterranean style [46, 47] PBDs or vegetarian [48, 49] or low carbohydrate diets [50-52] are
all considered healthy evidence-based eating models. Guidelines are emphasizing the
importance of individualization, with focus on individual goals, needs and preferences.
ADA Standards 2020 T2D Prevention [4]. The Diabetes Prevention Program [53] and
other prevention studies, like Da Qing Study [54] confirmed that lifestyle is a cornerstone for
preventing T2D, with individualized hypocaloric plan leading to achieve optimal weight and
normalize cardiometabolic markers, like lipids, blood pressure and inflammatory markers [4].
T2D incidence reduction by 58% was demonstrated through DPP along 3 years of intensive
lifestyle intervention. A significant result, observed in Da Qing Study was a reduction in all-
cause mortality and also cardiovascular mortality during 30 years of follow up.
DPP goals were 7% of weight loss and 150 min of physical activity per week. These
achievements were obtained during 6 months of interventions. Calorie goals have been
calculated by obtaining a daily deficit of 500-1000 kcal. The recommendation was focused on
the first step of total fat reduction. Then, the calorie balance and calorie restriction concept were
introduced [4].
DPP used an individual model of a lifestyle intervention that allowed tailoring interventions
according to population diversity. Persons with overweight and obesity are at high risk of
developing T2D [55, 56] and a hypocaloric meal plan with physical activity is necessary. Long
term lifestyle changes leading to weight reductions are difficult to be maintained and need
specialized support. What should eat models be used in pre-diabetes? Intervention trials are
suggesting the Mediterranean Model [57-63] and low fat and hypocaloric models. according to
ADA 2020 standards, PBD is not excluded but not explicitly recommended for T2D prevention.
The Healthy Index is an excellent tool to approach the healthiness of the food consumed.
Whole grains, nuts, legumes, vegetables and fruits as well as ultra-processed food avoidance
are evaluated by this Index. [63]
Conclusions
Improved glycaemic control during the Med Diet, vegan diets or vegetarian diets is strongly
supported by a meta-analysis of randomized controlled trials RCT's. Management of T2D
should take into consideration the benefits offered by plant-based models. The prevention in
61
T2D should start by adopting a healthy lifestyle; this could include plant-based dietary patterns
that would be helpful but does not exclude other models and studies are not emphasizing any
superiority vs the Med Diet.
In our opinion, after literature revision, as patient preference will maximize adherence to
lifestyle intervention, The PBD should be strongly recommended when it will guarantee the
highest adherence to a healthy lifestyle due to patient preference. Otherwise, the Mediterranean
model is the most complex, bringing all benefits and being a long-term option both in the
prevention and management of T2D.
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65
Male Infertility, Metabolic Syndrome and Other Related

Disorders
PACU Irina1,2, IONESCU Crîngu Antoniu1,2, PACU Ovidiu3,

DIMITRIU Mihai1,2, BĂNACU Mihail1,2, SOCEA Bogdan1,2, MĂDAN Victor3,
HANGANU Irina2, ZYGOUROPOULOS Nikolaos2
3 Department of Urology, “Dr. Carol Davila” Central Military Emergency Hospital, Bucharest (ROMANIA)
Emails: irinapacu@hotmail.com, antoniuginec@yahoo.com, ovidiupacu@live.com, drmihaidimitriu@yahoo.com,

bogdansocea@gmail.com, victmad@gmail.com, nz@zitaquality.ro
Abstract
Metabolic syndrome (MetS) represents a cluster of factors that have a negative impact on
human health which extends beyond the cardiovascular system. Growing evidence has linked
it to the increasing prevalence of male infertility. This brief review article considers the
literature regarding MetS, its components and other related pathologies with male infertility.
Effects of obesity, insulin resistance (InR), dyslipidaemia (DysL), hypertension (HT) and
inflammation – though not strictly part of MetS – on male infertility were examined and support
evidence explained in brief. Currently, there are limited prospective studies examining the
effects of treating MetS on male reproduction or research in understanding the underlying
pathophysiology linking MetS and male infertility. These should be a focus of further
investigation to elucidate the role of therapeutic intervention.
Keywords: metabolic syndrome, male infertility, inflammation
Introduction
Infertility according to the World Health Organisation (WHO) is the inability to achieve a
clinical pregnancy despite 12 months of unprotected intercourse [1]. Both male and/or female
factors can contribute to infertility. Male factor infertility – causative or contributing – is
estimated to occur in 20-50% of infertile couples [2]. In recent years there is some evidence
that the prevalence of male infertility may be increasing due to declining traditional sperm
parameters (volume, concentration, motility percentage and normal morphology) as well as
deteriorating sperm chromatin integrity [3, 4]. The coincidence of decreasing sperm quality and
increasing prevalence of MetS and its related disorders has prompted several scientists to
postulate that MetS is contributing to male infertility, albeit directly or indirectly. This could
occur through an association between the different components of MetS and sperm production
and function or through the pathophysiological pathways instigated by MetS affecting male
fertility [2, 3, 5].
The term MetS, refers to a cluster of abnormalities associated with an increased risk for
cardiovascular disease [3] namely: obesity, insulin resistance, systemic hypertension and
dyslipidaemia [6]. Nevertheless, disparity still exists regarding the exact diagnostic criteria used
in clinical practice. The following 3 guidelines are most applied to recognize this condition each
with its merits and disadvantages: WHO [7], National Cholesterol Education Program Adult
Treatment Panel III (NCEP ATP III) [8] and International Diabetes Federation (IDF) [9]. Table
1 summarizes the different criteria for the classification of metabolic syndrome in men.
66
Furthermore, patients with MetS according to the NCEP ATP III criteria, frequently have a pro-
inflammatory state with elevated cytokines and acute phase reactants (e.g., C-reactive protein)
something that will discussed in more detail below [10].
Table 1. Criteria for the classification of metabolic syndrome in men. Adapted from Martins,
A.D. et al., (2019) [2]
WHO [11] NCEP ATP III [12] IDF [13]
Central Obesity Waist/Hip ratio >0.9 Waist circumference Waist circumference
and/or >102 cm dependent on
BMI >30kg/m2 ethnicity
Raised Blood ≥140/90 mmHg Treatment of Treatment of
Pressure previously diagnosed previously diagnosed
hypertension hypertension
or or
≥130/80 mmHg ≥130/85 mmHg
Raised FPG Impaired glucose Previously diagnosed type 2 diabetes mellitus
tolerance or
Impaired fasting glucose FPG ≥100 mg/dL
Type 2 Diabetes mellitus
Reduced HDL <40 mg/dL Specific treatment for cholesterol
cholesterol Or
<40 mg/dL
TG ≥150 mg/dL Specific treatment for lowering TG
or
≥150 mg/dL
Diagnostic Insulin resistance Any 3 out of 5 of the Central obesity & any
criteria evidence is an absolute above criteria other 2 criteria of the
requirement requirement above
BMI: Body Mass Index, FPG: Fasting Plasma Glucose, HDL: High-density Lipoprotein-cholesterol,
TG: Triglycerides
The prevalence of MetS is influenced significantly by the adopted diagnostic criteria in

addition to age, ethnicity, urbanisation rate as well as education level, lifestyle (diet and
exercise) and socioeconomic status. [11].
Obesity & Infertility
The exact pathophysiology for how obesity affects male infertility remains not clearly
defined. Several mechanisms have been suggested; among the ones most commonly agreed are:
Obesity and increased adipose tissue lead to perturbations on the hypothalamic-pituitary-
gonadal axis (HPG) [12]; Obesity leading to increased oxidative stress and production of
reactive oxygen species (ROS) affecting sperm quantity and quality [13-16]; Increase in
suprapubic and scrotal fat creating an environment which hinders normal spermatogenesis [17-
20].
Excess adipose tissue increases peripheral conversion of testosterone to estrogenic which in
turn inhibits the HPG leading to secondary hypogonadism. Several studies both in animal
models [13] and longitudinal studies in humans [21, 22] linked obesity and disruption of the
central endocrine regulation of male fertility by decreasing total testosterone (tT), free
testosterone (fT) and sex hormone-binding globulin (SHBG) [21-23]. Further, a linear
association between BMI and incidence of oligospermia as well as a correlation between waist
circumference with low sperm concentration and low total sperm count was found [24].
Similar associations were reported in other studies [14, 25-28].
In addition, sperm motility, morphology and DNA fragmentation causing a decrease in
sperm quality in obese males have been associated secondary to oxidative stress [13, 15, 16].
67
The DNA damage is further compounded by mechanical factors which are hypothesized to
impact adversely fertility such as increased scrotal temperature [17] and scrotal lipomatosis
[18].
Insulin Resistance, Diabetes & Infertility
InR and diabetes appear to negatively affect male infertility through similar
pathophysiologic processes as obesity [3]. Both a prospective study [29] and similar cohort
studies [30] showed that the risk of developing diabetes was significantly increased in patients
with decreased fT and low SHBG. Similarly, a retrospective study showed that 33% of type 2
diabetes mellitus patients had hypogonadotropic hypogonadism [31]. It is important to note that
it has not been elucidated whether hypogonadism is a risk factor for diabetes. Further, InR and
diabetes appear to negatively affect sperm quantity and quality (DNA fragmentation) [17] while
treatment with metformin seems to reverse these effects [32]. Larger randomised control studies
are required to confirm the above positive result. Sequelae of diabetes – mainly neurological –
such as erectile dysfunction [33] and ejaculatory dysfunction [34] impact further on fertility.
Hence diabetic patients presenting with azoospermia or oligospermia should be assessed for
possible retrograde ejaculation [3].
Dyslipidaemia & Infertility
DysL-defined as elevated TG and decreased HDL cholesterol is another key component of

MetS though less researched than obesity and diabetes regarding to male infertility [3].
Animal studies [35, 36] have demonstrated that hypercholesteremia and its resultant elevated
levels of ROS may alter normal sperm generation and function. Clinical studies – correlating
well to prior animal studies – demonstrated that increased free cholesterol and phospholipid
levels are associated with decreased sperm head size and percentages of sperm with intact
acrosome [37] as well as increased time to pregnancy. Overall DysL in male infertility seems
to be acting in two levels: a molecular level by affecting normal sperm development and
function and a metabolic level by increasing ROS. The suggestion by some animal studies [36]
regarding the role of cholesterol-lowering agents and antioxidants in improving male fertility
has not been validated to-date by prospective clinical trials.
Hypertension (HT) & Infertility
HT is a well-established risk factor for erectile dysfunction with effect on male fertility [5]
though testicular end-organ injury has not been clearly outlined. The observed relationship
between increased blood pressure and decreased androgens [38] which could be linked with
impaired reproductive potential [39] is not straightforward. The tested hypothesis, so far, has
been that androgen deficiency may be the root cause of HT by inducing increased arterial
stiffness [40]. Indeed, male patients treated with androgen suppression were found to have
increased aortic and arterial stiffness compared with age matched controls [41]. Studies on
antihypertensive drugs and their effect on testosterone levels demonstrated either no effect or
decrease in testosterone levels depending the agent used [39, 42, 43].
68
Other Related Pathologies & Infertility
Inflammation
An association between inflammation and MetS is well established [10]. Adipose tissue in
males expresses higher levels of inflammatory molecules such as tumour necrosis factor alpha
(TNF-a), interleukin-6 and CRP as well as an increased number of macrophages [44]. Further
an upregulation of inflammatory genes found prior to the development of InR lead to the
hypothesis that increased macrophages in adipose tissues may contribute to the development of
InR [45]. TNF-a and interleukin-1 have been implicated as main mediators of the inflammatory
process [46] which in turn increases levels of ROS negatively impacting normal reproductive
pathways [47] and more specifically sperm morphology and DNA of spermatozoa [15,48].
Inflammation is implicated both in MetS and male infertility and may play a significant role
in linking them [3].
Sleep Apnoea
Sleep apnoea is characterised by fragmented sleep caused by repeated episodes of upper
airway obstruction and hypoxia and is often diagnosed in obese and diabetic males [3]. A study
concluded that the condition is associated with HPG dysfunction as a result to obesity
accompanied by a decline in lower mean testosterone and luteinising hormones and to a lesser
extent to sleep fragmentation and hypoxia [49].
Metabolic Syndrome & Infertility
Individual components of MetS – as mentioned above – have been associated with male
infertility. More recently, there has been an increase in studies examining male infertility and
MetS as a whole. Also, there could be a correlation between MetS and first trimester
biochemical markers in pregnancy and with postpartum pain, but no correlation with
myelofibrosis [50-52]. In these it was demonstrated that decreasing tT and SHBG were
associated with increasing risk factors for MetS and its individual components [53]. Further an
increase prevalence of secondary hypogonadism in subjects with MetS was identified [53, 54].
In addition, in patients that met 3 of the 5 IDF criteria decreased percentage of sperm with
normal morphology was found as well as an increased testicular homogeneity which might be
the radiographic signs of atrophy and fibrosis secondary to the pro-inflammatory state [53].
Conclusion
MetS is an important global medical entity originally described to identify patients with
increased risk for cardiovascular disease. Today it is demonstrated that its effects extend to
other organ systems, including the male reproductive system. Obesity disrupts the HPG axis,
increases scrotal temperatures, impairs spermatogenesis, decreases sperm concentration and
negatively impacts on sperm morphology and function. InR also alters semen parameters,
disrupts normal endocrine function as well as leads to erectile dysfunction hence resulting or
compounding male infertility. Dyslipidaemia with increased oxidative stress may lead to a
further decrease in fertility. Each component having an effect of its own on male infertility but
also when put together having an additive, detrimental effect on reproductive functions such as
endocrine status or semen parameters may occur [54-58]. Oxidative stress and pro-
inflammatory state seen in MetS may provide a pathophysiological pathway to explain how
MetS and its components are associated with male infertility. However, further research is
required to further elucidate this link.
69
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72
Uncontrolled Diabetes Mellitus – Negative Predictive Factor for

Colorectal Cancer Recurrence after Curative Surgical Treatment
SUCEVEANU Andra-Iulia1, SUCEVEANU Adrian Paul1, PAREPA Irinel2,

ARDELEANU Valeriu3, MICU Ioan Sergiu1, DUMITRU Andrada1,
CATRINOIU Doina4, STANCIU Adina Elena5, NITIPIR Cornelia6,
MAZILU Laura7
1 Department of Internal Medicine – Gastroenterology, Faculty of Medicine, “Ovidius” University, Constanta (ROMANIA)
2 Department of Cardiology, Faculty of Medicine, “Ovidius” University, Constanta (ROMANIA)
3 Department of Morphological Sciences, Faculty of Medicine and Pharmacy, University “Dunarea de Jos” Galati
(ROMANIA)
4 Department of Diabetes Mellitus and Nutritional Diseases, Faculty of Medicine, “Ovidius” University, Constanta,
(ROMANIA)
5 Institute of Oncology Bucharest, Department of Carcinogenesis and Molecular Biology, Bucharest, (ROMANIA)
6 Department of Oncology, “Carol Davila” University of Medicine and Pharmacy, Bucharest, (ROMANIA)
7 Department of Oncology, Faculty of Medicine, “Ovidius” University, Constanta (ROMANIA)
All authors have an equal scientific contribution

Emails: andrasuceveanu@yahoo.com, asuceveanu@yahoo.com, irinel_parepa@yahoo.com, valeriu.ardeleanu@gmail.com
micuioansergiu@yahoo.com, d.andrada.92@gmail.com, adinaelenastanciu@yahoo.com, nitipir2003@yahoo.com,
dcatrinoiu@gmail.com, lauragrigorov@gmail.com
Abstract
Diabetes mellitus (DM) is one of the main risk factors for colorectal cancer (CRC), many
studies reporting this causative relation. We aimed to study if uncontrolled DM, trough all its
pathogenically mechanisms, can be a negative predictive factor for CRC recurrence during five
years of follow-up in a group of 95 diabetic patients diagnosed with CRC and surgically treated.
In case of recurrence, we noted the moment of recurrence’s diagnosis (during scheduled visit
or accidentally), the presence of physical symptoms and signs, the diagnostic tools used for
recurrence diagnosis, and other additional methods used for the final diagnosis of recurrence
and its staging, comparative in patients with controlled and uncontrolled diabetes mellitus. We
noted the location of recurrence, the time spent from surgery until recurrence appeared, and the
survival rates. Local or distant recurrences were considered. A series of tumour markers,
glucose serum levels and glycated haemoglobin, colonoscopies and CT-scans were done
according to guidelines during the follow-up period. The rate of recurrence was calculated SPSS
20.0 package was used to analyse the results. Sixty-one patients had a local or distant
recurrence. Uncontrolled-DM negatively influenced the rate of recurrence, especially in males
(p=0.03), the meantime to recurrence (p=0.043), the topography of recurrence (right and
transverse – p=0.022, respectively p=0.019), the rate of distant metastases (p=0.023) and the
mortality rates (p=0.022). Uncontrolled diabetes mellitus represents a key link in CRC
recurrence as well as a negative predictive factor cancer recurrence in diabetes patients.
Keywords: Uncontrolled, diabetes mellitus, colorectal cancer, recurrence
Background and Aim
CRC is one of the most spread cancers over the world, despite the enormous efforts made to
screen and early treat it [1]. DM is considered an independent risk factor for many malignancies,
especially for colorectal one [2]. The control of glucose levels is imperative for a good balance
of the human microbiota and of the host gut inflammatory status. During the last decades,
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glucose imbalance was frequently linked with human dysbiosis and could represent an
important negative predictive factor for colorectal recurrence rates after surgical treatments
with curative intent [3, 4]. We aimed to study if uncontrolled DM, trough all it is pathogenically
mechanisms, can be a negative predictive factor for CRC recurrence during five years of follow-
up in a group of 95 diabetic patients diagnosed with CRC and surgically treated. We used the
patients’ medical files registered in the last five years in Emergency Hospital of Constanta
electronic database archive, and we focused on the following data: tumour stage and location
along the colon or rectum at the time of diagnosis, the length of follow-up, and the mortality
occurrence during the study period, between January 2015 and December 2019. The date of the
surgical treatment for the primary CRC was considered the baseline for the survival intervals
analysis. In case of recurrence, we noted the moment of recurrence’s diagnosis (during
scheduled visit or accidentally, with the occasion of investigations for other non-CRC related
reasons), the presence of physical symptoms and signs, the diagnostic tools used for recurrence
diagnosis, and other additional methods used for the final diagnosis of recurrence and its
staging. We noted the location of recurrence, time spent from surgery until recurrence appeared,
and survival rates. Local or distant recurrences were considered. A series of colonoscopies were
done at 12, 72 and 60 months from the surgical moment. Imagistic procedures, as
abdominopelvic CT scans, were done annually.
Tumour markers as CEA and CA 19-9 specific for CRC recurrence were also monitored
annually. The glucose serum balance and glycated haemoglobin (HbA1c) were usually used to
monitor the control of diabetes mellitus. The rate of recurrence was calculated for each
scheduled visit or intervals between visits. SPSS 20.0 package was used to analyse the results.
Chi-square and Fisher’s tests were useful tools to compare means. Kaplan-Meier analysis
was used for calculating the overall survival in all patients developing a recurrence. The log-
rank test was used for comparison with survival rates in patients without recurrence during the
follow-up period.
Patients and Method
The study has a retrospective observational non-interventional design. Sixty-one patients

(64.21%) admitted in Gastroenterology, Oncology and Diabetes Mellitus Departments had a
recurrence of CRC during standard surveillance procedures recommended by oncology
guidelines for colorectal cancers. We used the patients’ medical files registered in the last five
years in Emergency Hospital of Constanta electronic database achieve and we focused on the
following data: tumour stage and location along the colon or rectum at the time of diagnosis,
the length of follow-up, and the mortality occurrence during the study period, between January
2015 and December 2019. The date of the surgical treatment for the primary CRC was
considered the baseline for the survival intervals analysis. In case of recurrence, we noted the
moment of recurrence’s diagnosis (during scheduled visit or accidentally, with the occasion of
investigations for other non-CRC related reasons), the presence of physical symptoms and
signs, the diagnostic tools used for recurrence diagnosis, and other additional methods used for
the final diagnosis of recurrence and its staging. We noted the location of recurrence, time spent
from surgery until recurrence appeared, and survival rates. Local or distant recurrences were
considered. A series of colonoscopies were done at 12, 72 and 60 months from the surgical
moment. Imagistic procedures, as abdominopelvic CT scans, were done annually. Tumour
markers as CEA and CA 19-9 specific for CRC recurrence were also monitored annually. The
glucose serum balance and glycated haemoglobin (HbA1c) were monitored at every 3 months,
too. The rate of recurrence was calculated for each scheduled visit or intervals between visits.
SPSS 20.0 package was used to analyse the results. Chi-square and Fisher’s tests were useful
tools to compare means. Kaplan-Meier analysis was used to calculate the overall survival in all
74
patients developing recurrence. The log-rank test was used for comparison with survival rates
in patients without recurrence during the follow-up period.
Results
During the study period, 215 patients with DM and concomitantly CRC were identified in
the hospital’s electronic database in the clinics mentioned above. From the total of 215 patients,
120 patients were excluded from our analyse due to different reasons, as tumour stage IV, severe
co-morbidities, like recent heart attacks or strokes 12 months before baseline enrolment period,
inherited CRC with positive anamnesis or genetic testing documentation, history of other risk
factors for CRC like acromegaly, IBD’s or other documented cancers.
Exclusion criteria for patient’s selection into the current study can be followed in Table 1.
Table 1. Exclusion Criteria from the current study

Reason No. (%)
Stage IV disease 77 (64.16)
Hereditary CRC 10(8.33%)
Carcinoma in patients with inflammatory bowel diseases 14(11.66%)
Other malignancies 8(6.66%)
Other reasons (stokes, MI, acromegaly, etc.) 10(8.33%)
Ninety-five patients were finally included for the analysis, of which 15 patients (15.78%)
had stage I carcinoma, 33 patients (27.5%) had stage II carcinoma, and the rest of 47 patients
(49.47%) stage III carcinoma. According to location, the majority of patients had tumours
located on the left colon and rectum – 55 patients (57.89%), of whom 14 patients (14.73%) on
the rectal segment, 12 patients (12.63%) on the transverse colon and the rest of 28 patients
(29.47%) had tumours located on the ascending and cecum segments. Thirteen patients
(13.68%) had synchronous tumours along the colon. The median follow-up period was thirty-
nine months, ranging from 5 to 60 months, and 58 patients (61.05%) finished the follow-up
period. The mean time to recurrence occurrence was 26.7 months, ranging from 5 to 44 months.
Sixty-one patients (64.21%), 41 males (67.21%) and 20 females (32.78%) developed
recurrent disease during the follow-up period. During the follow-up period, 40 patients had
uncontrolled DM, and 21 had controlled DM. Patients with at least two consecutive abnormal
values of glycated haemoglobin were considered uncontrolled DM patients. From these
patients, 4 patients (6.55%) were classified as stage I at diagnosis, 15 patients (24.59%) were
classified as stage II at baseline, and the rest of 42 patients (68.85%) were stage III at baseline.
The mean age of patients with recurrence was 65.3, ranging from 51.7 to 79.5 years of age,
a bit younger in uncontrolled DM patients, but without statistical significance (p=0.68).
According to location, 33 tumours (54.09%) were located on the left colon, 5 tumours
(8.19%) were located on the transverse colon, and 23 tumours (34.42%) were located on the
right colon. According to recurrence location, more right and transverse colon cancers recurred
in uncontrolled-DM compared to controlled-DM (p=0.022, respectively p=0.019), while left
and rectum cancers recurred equally (p=0.051). There were 24 patients (39.34%) who
developed distant recurrence, as liver metastasis – 20 patients (32.78%) or lung metastasis – 4
patients (6.55%), their presence is related with the discovery of CRC recurrence, significantly
more in uncontrolled DM. Symptoms did not always accompany recurrences. Still, the presence
of symptoms was the reason for unscheduled medical visits, and they coincided with the time
of recurrence diagnosis, the recurrence diagnosis being significantly more often fixed in
symptomatic cases (p=0.03, ss). There were 38 patients (62.29%) with recurrence-related
75
symptoms as abdominal discomfort, abnormal transit, and anaemia-related symptoms as

fatigue, pallor, loss of appetite, weight loss, tachycardia, related to local recurrence or right
upper/diffuse abdominal pain and caught or dyspnoea, related to distant metastases. Nineteen
patients (31.14%) had both local and distant symptomatic metastases. Twenty-three patients
(37.70%) did not reveal any clinical sign of recurrence. The median time of recurrence was 41
months from the baseline, shorter in uncontrolled-DM compared to controlled-DM patients
(p=0.04). All features of studied patients at baseline and are exposed in Table 3.
Table 2. Main features of patients with CRC enrolled in the study

Characteristics Total – n %
Age, median (range), y 65.3 (51-79.5) -
Gender
Females 35 36.84
Males 60 63.15
Tumour stage at diagnosis, No. (%)
Stage I 4 6.5
Stage II 15 24.59
Stage III 42 68.85
Primary tumour location
No. (%)
Right colon, including hepatic flexure 28 29.47
Transverse colon 15 12.63
Left colon, including splenic flexure and rectosigmoid
colon 55 57.89
Synchronous tumours 17 17.89
Follow-up period 39 mo (5-60) -
Table 3. Main features of patients with CRC recurrence according to DM compensation

Uncontrolled
Characteristics DM Controlled DM Total (n, %) P-Value
Age, median (range), y 67 (51-73) 69 (46-76) 65.3 (51-79.5) 0.68
Gender
Males 30(73.17) 11(26.82) 41(100) 0.030
Females 10(50) 10(50) 20(100) >0.5
Recurrence location, No. (%)
Right colon, including hepatic flexure 15 (24.59) 8 (13.11) 23 (34.42) 0.022
Transverse colon 4 (6.5) 1 (1.6) 5 (8.19) 0.019
Left colon, including splenic flexure and 19(31.14) 14 (22.95) 33 (54.09) 0.051
rectosigmoid colon
Distant metastases 19 (31.14) 5 (8.19) 24(39.35) 0.023
Median time to recurrence detection,
median (range), mo 8.9 (2-48) 19.5 (4-46) 16.7 (5-44) 0.04
Symptoms related recurrence 40(65.57) 21(34.42) 61(100) 0.002
Type of visit where recurrence was detected, No. (%)
Scheduled follow-up 18 (62.29) 13 (37.70) 31 (100) 0.054
Interval visit 25(40.98) 5 (8.19) 30 (10.9) 0.011
Liver 7(11.47) 2(3.27) 9 (14.75) 0.021
76
Peritoneum 7(11.47) 0 7 (21.7) >0.001

Lung 4(6.55) 1(1.63) 5 (19.6) 0.022
Second primary (metachronous) 4 (6.55) 3 (4.91) 7 (11.47) 0.30
colorectal tumour
Mortality rates 36% 15% 28% 0.022
The method of diagnosing CRC recurrence was different along the study period. The
majority of patients had increased tumour markers, CEA and CA 19-9 (49, respectively, 51
patients). Some of them were diagnosed with CRC recurrence during abdominal imagistic
procedures as ultrasound or CT scans, following the scheduled program or imagistic
investigation made for other reasons. Patients with peritoneal, abdominal or liver metastasis
were routinely diagnosed by abdominal ultrasound. Patients with lung metastasis were
diagnosed usually with chest X-rays or thorax CT-scans. Radiographic or PET-CT diagnosed
bone metastasis. Most patients with uncontrolled DM required an unscheduled visit for the CRC
recurrence (p=0.011). The diagnosis method can be followed in Table no 4.
Table 4. Main diagnostic procedures for CRC recurrence
Test No (%)
CEA 49 80. 32
CA 19-9 51 83.00
Abdominal ultrasound 38 62.29
Colonoscopy 59 96.72
Chest x-ray 5 19.60
CT scan 47 77.04
Combined modalities 40 65.57
The mortality rates were calculated during the study period, and results were compared.
The overall mortality rates in 5 years of follow-up were 28%, still, significantly higher in
uncontrolled DM compared to controlled DM (p=0.022).
Discussions
According to GLOBOCAN data from 2018, CRC is the third most common cancer
worldwide [5, 6]. Reviews of literature in the matter conclude that intensive follow-up leads to
an overall 5-year survival benefit, compared with no follow-up. Still, the disease-specific
survival did not improve significantly over the decades, many risk factors related to host and
stage of disease at diagnosis influencing the cancer outcome and secondary disease-specific
survival [7-11]. Still, the intensive follow-up after CRC surgical treatment and DM control
looks to improve disease-free survival rates in diabetes patients positively influencing the CRC
recurrence rates [12-14]. Our study focused on the negative influence that has uncontrolled DM
in CRC outcome and recurrence rates, knowing that DM is one of the independent risk factors
for CRC occurrence. In our study, the mean age of recurrence was similar in both studied
groups, slightly increased in uncontrolled DM group (p=0.68). The control of DM influenced
the occurrence of CRC, especially in males, where uncontrolled diabetes was a risk factor for
cancer recurrence (p=0.030), accordingly with international literature [15]. Opposite, in
females, the serum glucose level did not influence the occurrence of CRC recurrence (p=0.5).
In our series, DM presence influenced the outcome and recurrence rates significantly in
patients with CRC during the follow-up period. Studies are showing that around 42% of
77
monitored patients presented recurrence during interval visits, and 83.87% of these visits were
related to symptoms suggestive of recurrent disease [16]. In our study, the uncontrolled DM
influenced first the moment of recurrence during the follow-up period. Most patients with
uncontrolled DM presented recurrences diagnosed between scheduled follow-up visits
(p=0.031). Furthermore, during the scheduled visits, patients with uncontrolled DM had much
more recurrences (p=0.044). During interval visits, uncontrolled DM patients had more
recurrences than patients with controlled serum glucose level (p=0.011) again. Majority of
recurrences were symptomatic, the symptoms being the alarm signs for patients to request an
unscheduled visit to their doctors. Altered defecation habits, rectal discharge of blood,
abdominal cramps, weight loss, were main symptoms bringing the patients to physicians [13,
17]. On the other hand, abdominal ultrasound and CT scans detected more recurrences in
patients without complaints [18-20]. Left colon recurrences were similarly represented in
controlled and uncontrolled DM, while right and transverse recurrences were more common in
uncontrolled DM. The meantime, the recurrence was significantly shorter in uncontrolled
diabetes (p=0.043) compared to controlled diabetes, this remark strengthening the hypothesis
that diabetes accelerates the replication of the malignant cell [21]. The mortality rates are still
high in CRC, the specific disease survival not being sufficiently influenced by the intensive
follow-up after primary cancer treatment [22]. In our study, the overall mortality rates in 5 years
of follow-up were 28%, significantly higher in uncontrolled DM compared to controlled DM
(p=0.022).
Conclusions
A stronger understanding of CRC evolution looks to be linked with the environmental and
genetic risk factors, the misdiagnosed or uncontrolled DM being a key link in CRC recurrence
in diabetes patients.
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79
Assessment of Physical Activity in the Elderly an Element in

Maintaining Vascular Resilience?
AURELIAN Sorina Maria1,3, MIHALACHE Ruxandra1,3,

ZAMFIRESCU Andreea1,3, AURELIAN Justin2, CAPISIZU Ana1,3
1 Department of Geriatric Rehabilitation, “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
2 Department of Nursing, “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
3 “Sf. Luca” Hospital of Chronic Diseases, Bucharest (ROMANIA)
Emails: sorinamaria.aurelian@gmail.com, justin.aurelian@gmail.com, ruxy_das@yahoo.com,

andreea.zamfirescu@gmail.com, capisizu.ana@gmail.com
Abstract
Introduction
Frailty is considered to be prevalent with age and associated with a high risk on falls,
disability, hospitalization and mortality. Physical activity is a protective factor for the
occurrence of sarcopenia for older adults, however there are limited studies in this area. Aim of
this study was to evaluate the physical performance and its impact on functionality of
hemodynamic parameters in elderly patients.
Material & Methods

We conducted a prospective study for three months (July-September 2019) in which 143
patients were included, with age over 65 years (mean of 72.85±10.07 years), from which 113
were women. The study was conducted in the Geriatrics Clinic from “Sf. Luca” Hospital of
Chronic Diseases. We measured Physical activity through Short Physical Performance Battery
(SPPB) that targets: tandem standing, 4m walking test and up to go test; functional capacity by
standardized scales Activity Daily Living (ADL) and Instrumental Activity Daily Living
(IADL); the muscle strength through hydraulic dynamometer; arterial vascular stiffness (PWV
and PP) using the Tensiomed Arteriograph.
Results
The physical performance if patients changes with age. That is why in the age group 75-84y
we have the highest percentage of patients with low status of physical activity (53%) compared
to only 30% in the elderly (65-74y). Physical performance and impact on daily activity scores
show a direct relationship between the functionality and the SPPB values. The results showed
a significant statistical difference on the pulse pressure (PP) (p=0.001). We consider this as a
new predictor for cardiovascular risk assessment and risk evaluation of disability by walking
speed.
Conclusions
Determining muscle strength to assess physical persistence with clinical implications in
lifestyle optimization and treatment recommendations in elderly patients, seems to be a
necessity.
Keywords: physical activity, vascular resilience, frailty, elderly
80
Introduction
Frailty is considered to be prevalent with age and associated with a high risk on falls,
disability, hospitalization and mortality [1]. Weakness, slower movement, and limitation in the
ability to perform daily routine physical activities often signal the onset of frailty in the elderly
[2]. Chronic malnutrition as well as musculoskeletal changes associated with age, chronic
diseases, micro-inflammation and associated comorbidities contribute to sarcopenia by loss of
muscle mass [3]. Sarcopenia is characterized by muscle weakness, decrease the movement
speed which leads to disabilities. Sarcopenia is a health problem related to the aging of the
geriatric population [4]. Although physical activity isa protective factor for the occurrence of
sarcopenia for older adults, there are limited studies in this area. Recent research has
demonstrated the importance of physical activity in pre arrival premature aging, maintaining
functional independence, increasing quality of life and not least a joint intellectual integrity.
Aim of this study was to evaluate the physical performance and its impact on functionality
of hemodynamic parameters in elderly patients.
Material & Methods
We conducted a prospective study of three months (July-September 2019) in which 143

patients were included, with age over 65 years (mean of 72.85±10.07 years), from which 113
were women. The study was conducted in the Geriatrics Clinic from “Sf. Luca” Chronic
Disease Hospital. We measured Physical activity through Short Physical Performance Battery
(SPPB) that targets: tandem standing, 4m walking test and up to go test [5]; We measured
functional capacity by standardized scales Activity Daily Living (ADL) and Instrumental
Activity Daily Living (IADL) [6, 7]. We measured the muscle strength through hydraulic
dynamometer; we measure also arterial vascular stiffness (PWV and PP) using the Tensiomed
Arteriography [8]. All determinations were performed according to the protocol and the results
were analysed according to standardized scales. Statistics were conducted through SPSS
software.
Results
The physical performance if patients changes with age. That is why in the age group 75-84
we have the highest percentage of patients with low status of physical activity (53%) compared
to only 30% in the elderly (65-74).
The age caused changes in the physical status of patients. Thus, in the age group “very
elderly” (75-84) 53% of those with low physical activity were presented, compared to only 30%
among those with medium and good physical activity (Chi square test, p<0.001). Also, in the
“very very old” age group (over 85 years) we noticed the same statistically significant
difference between the two groups (Fig. 1).
81
Fig. 1. Physical activity by age
Within the group, there is no statistically significant difference in terms of physical

performance and the number of associated diseases (expressed by the comorbidity index) (Table
1).
Table 1. Physical activity and comorbidity index

Low physical activity Average + good physical activity
Characteristic p
(N = 66) (N = 77)
Comorbidity index
Small (0-2 diseases) 1 (2%) 2 (2%)
0.389
Medium (3-5 diseases) 52 (79%) 66 (86%)
Large (6-8 diseases) 13 (19%) 9 (12%)
SM components
Without SM 18 (27%) 14 (18%)
0.353
SM3 23 (35%) 34 (44%)
SM4 and SM5 25 (38%) 29 (38%)
Diabetes
Yes 29 (44%) 40 (52%) 0.339
Not 37 (56%) 37 (48%)
The functional capacity of the elderly in the study group were relatively normal, with not
more than two activities affected, especially in the instrumental activities of living as follows:
the IADL mean is 6.66±1.71 (out of a maximum of 8 points) and the mean of ADL is 5.74±0.92
(out of a maximum of 6 points) (Table 2).
Physical performance and impact on daily activity scores show a direct relationship between
the functionality and the SPPB values. There is a statistically significant difference (Student t-
test, p<0.001) in the case of IADL, within the group of those with low physical activity
(6.06±1.81 versus 7.17±1.43). Also, there is an affliction of basic needs or in the case of the
individual, measured by the ADL (eating, washing) from the group with a reduced physical
performance (Student t-test, p=0.004) (Table 2).
Measurement of muscle strength by dynamometer and adjustment by age, gender and height
showed a significant difference (Student t-test, p<0.001) for the reduced physical activity group,
being only 11.74±2.9 kg versus a close value of the normal 17.08±4.94 kg among the physically
active (Table 2).
Statins existing in the regime of elderly patients with chronic cardiovascular diseases are
often linked to sarcopenia, fatigue and decreased physical activity.
82
The correlation of statin administration with impairment of muscle strength as well as

physical performance does not show statistically significant differences in our patient
population. More than half (51%) of the elderly treated with statins showed good physical
activity (Table 2).
Table 2. Physical performance and functional capacity in the studied group

Low physical activity Average + good physical activity
Characteristic p
(N = 66) (N = 77)
ADL 5.50±0.93 5.95±0.87 0.004**
IADL 6.06±1.81 7.17±1.43 <0.001***
Subjective fatigue
Rare 8 (12%) 17 (22%)
<0.001***
Sometimes 9 (14%) 37 (48%)
Often 49 (74%) 23 (30%)
Muscle strength 11.74±2.9 17.08±4.94 <0.001***
Speed of walking 0.34±0.1 0.69±0.13 <0.001***
Statin
Yes 29 (44%) 39 (51%) 0.423
Not 37 (56%) 38 (49%)
Analysing speed of walking: we analysed the shift rate of the patients, which we correlated
with some hemodynamic parameters (AV, PWVao, PP) and the functional capacity of the
patients. We divided into three groups of patients after speed as follows: first group[home
movement] – consisting of elderly people with an increased risk of disability, mobile only in
the house, with a speed of up to 0.4 m/s; the second group[limited community movement] –
consisting of elderly people moving in the community with a moderate risk of losing their
autonomy with a walking speed between 0.4-0.8 m/s; third group[full mobilization] – those
who are not at risk of disability and who are fully mobile at a walking speed of over 0.8 m/s
(Table 3). The results showed a significant statistical difference on the pulse pressure (PP)
(p=0.001). We consider walking speed a new predictor for cardiovascular risk assessment and
disability risk evaluation (Fig. 2).
Table 3. Distribution of patients according to the speed of movement

Classification of displacement speed
Group 1 = home Group 2 = limited community Group 3 = full p

movement movement mobilization
(N = 48) (N = 71) (N = 24)
ADL 5.46±0.84 5.80±0.95 6.13±0.85 0.011**
IADL 6.19±1.8 6.75±1.71 7.33±1.09 0.021**
PWVao 10.37±1.74 9.94±1.93 9.95±1.63 0.419
PP 66.71 ± 14.23 65.17±15.31 53.42±12.75 0.001***
AV 68.81±12.7 66.61±11.36 65.33±7.02 0.404
83
Fig. 2. PP correlation with walking speed
Discussion
Physical activity is the only way to preserve autonomy but also to maintain good cognition.
It is well known that regular aerobic exercise improves cardio-respiratory mechanism,
lowers arterial stiffness [3, 9], reduces cardiovascular risk factors and reduces cardiovascular
mortality. In contrast, anaerobic exercises increase the force of skeletal muscle and play an
important role in preventing muscle atrophy that occurs with aging [10]. Our tests confirm
previous studies, with a share of 53% (n=66) of those with low physical activity versus only
30% (n=77) among those with medium and good physical activity (t test, p<0.001).
Physical activity is an effective protective strategy for sarcopenia. Most studies on elderly
people exercise showed that participants achieved positive results, whilst maintaining muscle
strength appears to depend on continued implementation of certain types of physical activities.
A conclusion of a meta-analysis on 10 reviewed studies was that there was no coherence in
the measurement of sarcopenia, additional investigations being needed [11]. For elderly people,
regular exercise (at least one hour, at least twice a week) through personalized exercise
programs can have beneficial cardiovascular consequences (lower blood pressure, improve
skeletal muscle irrigation, and stimulate the use of glucose in myocytes), and consequences on
maintaining the number of muscles fibers and supporting skeletal muscle strength [12]. In our
study, the measurement of muscle strength through the dynamometer demonstrated a
significant difference in the group of elderly patients with low physical activity (Student t-test,
p<0.001).
Although other studies debated the issue of administering statins to elderly in the
development of sarcopenia, fatigue and decreased physical activity. Within our group, no
positive correlation was found indicating this relation. Thus, it can support the
recommendations for the administration of statins, even in old age having demonstrated
protection in the prevention of cardiovascular events.
Analysis of walking speed and interpretation is a challenge for the clinician, and recent
specialist studies have brought news about survival and mortality predictors. Normal speed
requires a relatively low level of energy consumption [3, 13]. In our study analysis on speed of
walking revealed statistically significant results on the value of pulse pressure (PP), considered
a predictor in the assessment of disability risk in the elderly population studied [14].
84
Conclusions
Determining muscle strength to assess physical persistence with clinical implications in

lifestyle optimization and treatment recommendations in elderly patients, seems to be a
necessity. Maintaining physical performance especially in the elderly with associated chronic
diseases has an important role in maintaining autonomy, quality of life, especially in the very
old and very very old age groups that become frailer. Exercise delays and prevents arterial
stiffness, cardiovascular diseases, playing an important role in promoting a healthy aging.
From a practical and clinical point of view, it is necessary to draw up a constant program of
adapted, personalized, individualized physical activities with positive effects on functional
parameters and with considerable impact on the quality of life of the elderly.
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85
Chronic Kidney Disease and Related Conditions in a Roma

Population with Diabetes Mellitus
RUSU Emilia1,2, COSOREANU Andrada2, BALEANU Maria3,

MARINESCU Mihai3, ENACHE Georgiana4, RUSU Florin5,
BEJINARIU Catalina2, JINGA Mariana1,5, RADULIAN Gabriela1
2 “Nicolae Malaxa” Clinical Hospital, Bucharest (ROMANIA)
3 “C.I. Parhon” National Institute of Endocrinology, Bucharest (ROMANIA)
4 “Dr Pompei Samarian” County Emergency Hospital, Calarasi (ROMANIA)
5 “Carol Davila” Clinical Emergency Military Hospital, Bucharest (ROMANIA)
Email: emiliarusumd@yahoo.com
* All authors contributed equally to this work.
Abstract
Introduction
Roma minority is one of the largest ethnic group in Romania with some particularities
regarding their health status and their metabolic profile.
AIM: The aim of the study was to evaluate the prevalence and the risk factors associated
with CKD in Roma patients with diabetes mellitus.

The study population consisted of 258 Roma adults’ participants, 134 (51.9%) males and
124 (48.1%) females, aged between 18 and 86 years. We assed anthropometric parameters and
performed biochemical analyses, including: fasting serum lipids fasting plasma glucose (FPG),
glycated haemoglobin (HbA1c), liver function tests. Renal function (creatinine, urea, eRFG)
was evaluated in all participants. Urine albumin to creatinine ratio (UACR) was determined in
a random urine specimen.
Results
CKD was found in 46.5% patients (n=120), 48.6% with T1DM (n=17), and 46.2% with
T2DM (n=103). Patients with T1DM and CKD had a statistic significantly longer duration of
diabetes, without differences regarding the area of residence, smoking status, the presence of
hypertension, the severity of obesity and concurrent diabetic neuropathy or retinopathy.
Patients with T2DM and CKD were older, greater proportion of hypertension, diabetic
neuropathy and retinopathy, significantly longer duration of diabetes.
Conclusions
Our study showed a high prevalence of CKD and cardiovascular risk factors among Roma
people with diabetes mellitus. Most of the patients fail to achieve the treatment goals regarding
BP, lipid profile and HbA1c, emphasizing the need of stronger interventions.
Keywords: ethnicity, diabetes mellitus, chronic kidney disease, cardiovascular risk factors
86
1. Introduction
Chronic kidney disease represents a worldwide public health problem, nowadays registering
a continuously increase regarding its prevalence and incidence. According to KDIGO
guidelines, chronic kidney disease (CKD) is defined as the presence of kidney damage,
structural or functional, or as estimated glomerular filtration rate (eGFR) less than 60
ml/min/1.73 m2 for more than 3 months [1].
Roma minority represents one of the largest ethnic group in Romania. According to the 2011
nationwide census, their number was 621.573 Roma, representing 3.08% of the population [2],
but the real percentage could vary between 4.6 and 10% of the general population because a lot
of Romani descent do not declare themselves Roma [3].
AIM
The aim of the study was to evaluate the prevalence and the risk factors associated with CKD
in Roma patients with type 1 and type 2 diabetes mellitus.
2. Material and Method
Trial design
A cross-sectional case-control study was conducted in department of Diabetes, Nutrition and
Metabolic Diseases at Nicolae Malaxa Clinical Hospital, Bucharest within January – December
2019.
Participants
The study population consisted of 258 Roma adults’ participants, 35 with type 1 (T1DM),
51.4% males (n=18) and 223 with type 2 diabetes T2DM), 52.0% males (n=116), aged between
18 and 86 years. The inclusion criteria were: Roma patients aged over 18 years with type 1 or
type 2 diabetes. The exclusion criteria were: Caucasians patients, people who refuse to sign the
informed consent, pregnant or nursing women. Written informed consent was obtained from all
participants. The study was approved by the local Ethics Committee.
Assessments
Anthropometric parameters (height, weight, abdominal circumference (WC)) were
evaluated in standard condition. BMI was calculated and classified, according to WHO [4].
Metabolic waist was defined as waist circumference over 94 cm in men, over 80 cm in
women [5].
Laboratory assays
The biochemical analyses, including fasting serum lipids (total cholesterol (TC), triglyceride
(TG), high-density lipoprotein-cholesterol (HDL-C)), fasting plasma glucose (FPG), glycated
haemoglobin (HbA1c), liver function tests, total protein, were performed. Low-density
lipoprotein cholesterol (LDL-C) was calculated using the Friedwald formula [6].
Renal function (creatinine, urea) was evaluated in all participants. Urine albumin to
creatinine ratio (UACR) was determined in a random urine specimen and stratified in normal
(A1), modestly elevated (A2), and severely increased (A3) according with American Diabetes
Association [7]. Estimated glomerular filtration rate (eGFR) was made according to CKD-EPI
equation [8].
87
Definition
According with KDIGO the CKD was classified using estimated glomerular filtration rate
and albuminuria [9] Diabetic retinopathy was assessed through eye fundus examination
according with The Early Treatment for Diabetic Retinopathy Study [10]. Hypertension was
defined as blood pressure <140/90 mmHg in previously diagnosed hypertensive having taken
antihypertensive medication during the previous two weeks.
Statistics
The statistical analysis was performed using Statistical Package for the Social Sciences
(SPSS) 19. Data are presented as number (percentage %), mean (± standard deviation) or
median (with interquartile range). Comparisons among groups were made by use of ANOVA
for quantitative variables and the χ2 test for categorical variables. Simple binary logistic
regression and backward stepwise multiple logistic analyses were performed to identify factors
influencing CKD. Statistical significance was set at 95% confidence interval.
3. Results
CKD was found in 46.5% patients (n=120), 48.6% with T1DM (n=17), and 46.2% with
T2DM (n=103). General characteristics of patients classified by type 1 or 2 diabetes and by the
presence or absence of CKD are detailed in Table 1. Patients with T1DM and CKD had a
statistic significantly longer duration of diabetes, without differences regarding the area of
residence, smoking status, the presence of hypertension, the severity of obesity and concurrent
diabetic neuropathy or retinopathy (Table 1). Patients with T2DM and CKD were older, greater
proportion of hypertension, diabetic neuropathy and retinopathy, significantly longer duration
of diabetes (Table 1). There were no statistically significant differences between patients with
or without chronic kidney disease concerning HbA1c, lipid profile (cholesterol, triglycerides,
HDL-cholesterol, LDL-cholesterol), liver test.
Table 1. General characteristics of patients classified by type 1 or 2 diabetes and by the presence or absence of
CKD
T1DM (n=35) T2DM (n=223)
CKD CKD
Parameters No Yes Total p* No Yes Total p* p#
Age (years) 46.44 48.53 47.46 NS 53.73 58.96 56.14 <0.001 <0.001
±13.85 ±14.31 ±13.91 ±10.98 ±10.16 ±10.90
Diabetes duration (years) 12.5 (19) 10 (1.5) 12 (18) 0.04 5 (9) 8.5 (9.5) 6 (9) <0.001 <0.001
&
Women 8 (47.1%) 9 (52.9%) 17 (48.6%) NS 67 (55.8%) 49 (47.6%) 116 (52%) NS NS
Men 10(55.6%) 8(44.4%) 18(51.4%) 53(44.2%) 54(52.4%) 107 (48%)
Hypertension 3 (16.7%) 7 (47.1%) 10(28.6%) NS 89(74.2%) 89(86.4%) 178(79.8%) 0.023 0.016
BP (mmHg) 126.27 129.07 127.62 NS 145.85 154.00 149.75 <0.001 <0.001
±13.55 ±23.35 ±18.62 ±22.70 ±22.01 ±22.69
Obesity 1 (5.6%) 1 (5.9%) 2 (5.7%) NS 81 67.5%) 70 (68%) 151 67.7%) NS NS
BMI (kg/m ) 2
24.06 ± 22.48 23.30 NS 34.51 33.96 34.25 ±6.47 NS <0.001
4.09 ±5.39 ±4.75 ±6.22 ±6.74
IHD 5 (27.8%) 13 18 (51.4%) 0.005 64 (53.3%) 87 (84.5%) 151 (67.7%) <0.001 <0.001
(76.5%)
DN 15 (83.3%) 12 28 (80%) NS 86 (71.7%) 93 (90.3%) 179 (80.3%) 0,.01 0.003
(76.5%)
DR 7 (38.9%) 10 17 (48.6%) NS 28 (23.3%) 52 (50.5%) 80 (35.9%) <0.001 <0.001
(58.8%)
Laboratory findings
A1c 10.17 11.56 10.90 NS 9.32 ±2.07 9.69 ±2.24 9.50 ±2.15 NS NS
±2.51 ±2.46 ±2.53
88
Creatinine (mg/dL) 0.91 ±0.16 1.48 1.19 ±0.62 0.005 0.87 ±0.16 1.33 ±0.57 1.08 ±0.47 <0.001 NS
±0.69
eGFR (ml/min/1.73m2) 85.28 54.86 70.51 <0.001 88.28 57.51 74.07 <0.001 NS
±14.04 ±23.48 ±24.41 ±15.03 ±22.26 ±24.19
ACR (mg/g) & 18 (1) 22 (25) 22 (23) 0.040 21 (4) 42 (32) 28 (24) <0.001 NS
Abbreviations SBP= Systolic Blood Pressure BMI= Body Mass Index IHD= Ischaemic Heart Diseases DN= Diabetic Neuropathy DR=
Diabetic Retinopathy A1c= Haemoglobin A1c eGFR= estimated Glomerular Filtration Rate ACR= Albumine: Creatinine Ratio, & - Data are presented
median with interquartile range
The proportion of patients with T1DM and eRFG <60 ml/min/1.73 mp (stages G3a-G5) was
34.3% (n=12). In patients with T2DM and eRFG <60 ml/min/1.73 mp the percentage was
31.9% (n=71) (Table 2). Urinary albumin/creatinine ratio (UACR) >30 mg/g was present in
17.1% of patients with T1DM (n=6) and in 29.6 % of those with T2DM (n=66).
Table 2. GFR categories (ml/min/1.73 m2) in all patients

T1DM T2DM Total
(n=35) (n=223) (n=258)
G1 7 (20%) 68 (30.5%) 75 (29.1%)
G2 16 (45.7%) 84 (37.7%) 100 (38.8%)
G3a 7 (20%) 41 (18.4%) 48 (18.6%)
G3b 2 (5.7%) 24 (10.8%) 26 (10.1%)
G4 3 (8.6%) 4 (1.8%) 7 (2.7%)
G5 0 2 (0.9%) 2 (0.8%)
Glycaemic imbalance and CKD

For patients with diabetes mellitus and CKD, glycaemic control is essential to prevent or
delay the progression of microvascular complications. Our data suggest that a large number of
patients with CKD is not meeting treatment goal, 75% (n=90) had HbA1c >8% (70.6% (n=12)
in T1DM and 75.7% (n=78) in T2DM). Regarding the glucose-lowering medication, among
T2DM patients, 78.6% (n=81) are receiving metformin, 33% (n=34) sulfonylureas, 6.8% (n=7)
a GLP-1 receptor agonist and 64.1% (n=66) are on an insulin regimen.
Management of Cardiovascular Risk Factors
Hypertension
Antihypertensive drugs essentials in the management of CKD in diabetes are angiotensin-
converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARB). In our study
group, 45.8% (n=55) of patients were treated with ACE inhibitors and 6.7% (n=20) with ARB.
A large proportion of patients did not achieve the target blood pressure, 68.3% (n=82) with
BP>140/90 mmHg, most of them being patients with T2DM (75.7%, n=78).
Lipid Profile
According to European Society of Cardiology (ESC), and European Association for the
study of Diabetes (EASD), patients with diabetes mellitus in the presence of risk factors for
atherosclerosis (hypertension, dyslipidaemia, smoking, obesity), cardiovascular disease or with
target organ damage are at very high risk and the goal for them is the reduction of baseline
value of LDL-C with ≥50% and to obtain a target value of LDL-C <55 mg/dl. In the study
group, 58.8% (n=10) of patients with T1DM and CKD and 79.6% with T2DM and CKD were
on statin treatment, but only few had LDL-C under 70 mg/dl (14.2%, n=17) and fewer under
55 mg/dl (7.5%, n=9).
89
Risk factors for CKD

In all participants, in multivariate adjustment for all covariates in a backward stepwise
elimination procedure, significant predictors of CKD were IHD and hypertension; similar
results were obtained separately for T1DM and T2DM.
4. Discussion
In Romania, according to the PREDATORR (PREvalence of DiAbeTes mellitus,

prediabetes, overweight, Obesity, dyslipidaemia, hyperuricemia and chronic kidney disease in
Romania) study, about 7% of people are affected by chronic kidney disease, representing 1.3
millions of adults, most of them over 60 years old [11]. There are several studies in the literature
showing that the prevalence of CKD among T2DM patients is around 30% [12-14], with some
variations across population samples in Europe, and similar prevalence number was found in
few small sized studies from Romania [15]. In one study, the prevalence of CKD in T1DM
Romanian patients, it was around 41.5 % [16]. About Roma minority it’s been showed to have
a much higher prevalence of T2DM, metabolic syndrome and cardiovascular disease [17]. Our
study is constant with the available data from the literature, showing that in the Roma study
group there is a high prevalence of BP, dyslipidaemia and other cardiovascular factors. The
data focusing on CKD in the Roma ethnicity are limited, but some reports have shown higher
risk of CKD in this population and faster progression to end stage renal disease [18] that can be
related to the hereditary gene polymorphism, nutritional status, some psycho-social, cultural
and economic particularities [19, 20].
5. Conclusions
Our study found that in Roma ethnic group there is a high prevalence of CKD (46.5%)
among T1DM and T2DM patients, highlighting the possible intervention of other factors in the
evolution of diabetes mellitus and its complication in this ethnic population. A high prevalence
of cardiovascular risk factors was also observed. Most of the patients fail to achieve the
treatment goals regarding BP, lipid profile and HbA1c, emphasizing the need of stronger
interventions.
REFERENCES
1. Eknoyan G, Lameire N, Eckardt K, Kasiske B, Wheeler D, Levin A, et al., KDIGO 2012 clinical practice
guideline for the evaluation and management of chronic kidney disease. Kidney Int. 2013; 3(1): pp. 5-14.
2. de Statistică IN. Rezultate definitive ale Recensământului Populaţiei şi al Locuinţelor – 2011
(caracteristici demografice ale populaţiei). Bucureşti. Retrieved from
http://www.recensamantromania.ro/wp-content …; 2011.
3. Cartner H, Watch HR. Destroying Ethnic Identity: The Persecution of Gypsies in Romania, a Helsinki
Watch Report: Human Rights Watch; 1991.
4. Organization WH. World Health Organization BMI Classification. 2006.
5. Federation ID. The IDF consensus worldwide definition of the metabolic syndrome. IDF
Communications. 2006: pp. 1-24.
6. Friedewald WT, Levy RI, Fredrickson DS. Estimation of the concentration of low-density lipoprotein
cholesterol in plasma, without use of the preparative ultracentrifuge. Clinical chemistry. 1972; 18(6): pp.
499-502.
7. Association AD. 11. Microvascular Complications and Foot Care: Standards of Medical Care in Diabetes
− 2020. Diabetes Care. 2020; 43(Supplement 1): pp. S135-S51.
8. Levey AS, Stevens LA, Schmid CH, Zhang YL, Castro AF, Feldman HI, et al., A new equation to
estimate glomerular filtration rate. Annals of internal medicine. 2009; 150(9): pp. 604-12.
9. Kdoqi. KDOQI Clinical Practice Guidelines and Clinical Practice Recommendations for Diabetes and
Chronic Kidney Disease. Am J Kidney Dis. 2007; 49(2 Suppl 2): pp. S12-154.
90
10. Akduman L, Olk R. The early treatment for diabetic retinopathy study. Clinical trials in ophthalmology:
a summary and practice guide, Kertes, PS, Conway, MD, eds Baltimore: Williams & Wilkins. 1998: pp.
15-35.
11. Mota E, Popa SG, Mota M, Mitrea A, Penescu M, Tuta L, et al., Prevalence of chronic kidney disease
and its association with cardio-metabolic risk factors in the adult Romanian population: the
PREDATORR study. Int Urol Nephrol. 2015;47(11): pp. 1831-8.
12. Mata-Cases M, Franch-Nadal J, Real J, Cedenilla M, Mauricio D. Prevalence and co-prevalence of
chronic comorbid conditions in patients with type 2 diabetes in Catalonia: a population-based cross-
sectional study. BMJ Open. 2019; 9(10): pp. e031281-e.
13. Rodriguez-Poncelas A, Garre-Olmo J, Franch-Nadal J, Diez-Espino J, Mundet-Tuduri X, Barrot-De la
Puente J, et al., Prevalence of chronic kidney disease in patients with type 2 diabetes in Spain:
PERCEDIME2 study. BMC Nephrology. 2013; 14(1): p. 46.
14. Fraser SDS, Aitken G, Taal MW, Mindell JS, Moon G, Day J, et al., Exploration of chronic kidney disease
prevalence estimates using new measures of kidney function in the health survey for England. PLoS One.
2015; 10(2): p. e0118676-e
15. Munteanu M, Bob F, Schiller O, Mihaescu A, Munteanu L, Gadalean F, et al., Diabetic kidney disease
in type 2 diabetes mellitus – a prospective observational study. Acta Diabetologica Romana. 2019; 45(OP
36)
16. Vladu M, Clenciu D, Efrem IC, Forțofoiu M-C, Amzolini A, Micu ST, et al., Insulin Resistance and
Chronic Kidney Disease in Patients with Type 1 Diabetes Mellitus. J Nutr Metab. 2017; 2017: p. 6425359.
17. de Courten BV, de Courten M, Hanson RL, Zahorakova A, Egyenes HP, Tataranni PA, et al., Higher
prevalence of type 2 diabetes, metabolic syndrome and cardiovascular diseases in gypsies than in non-
gypsies in Slovakia. Diabetes research and clinical practice. 2003; 62(2): pp. 95-103.
18. Gadalean F, Lighezan D, Stoian D, Schiller O, Timar R, Timar B, et al., The Survival of Roma Minority
Patients on Chronic Hemodialysis Therapy – A Romanian Multicenter Survey. PLoS One. 2016; 11(5):
pp. e0155271-e
19. Nicholas SB, Kalantar-Zadeh K, Norris KC. Racial disparities in kidney disease outcomes. Semin
Nephrol. 2013; 33(5): pp. 409-15.
20. Lertdumrongluk P, Kovesdy CP, Norris KC, Kalantar-Zadeh K. Nutritional and inflammatory axis of
racial survival disparities. Semin Dial. 2013; 26(1): pp. 36-9.
91
The Metabolic Syndrome – Clinical and Biochemical Correlations
PASARICA Ilinca1, PERTEA Leonard-Iosif1, VLASE Alexandru1,

BARBU Roxana-Mihaela1, MUNTEANU Dragos1, CERNOMAZ Andrei1
1“Grigore T. Popa” University of Medicine and Pharmacy, Iasi (ROMANIA)
Emails: doctor_munteanu@yahoo.com, leo_pertea@yahoo.com
Abstract
The metabolic syndrome is a pathologic constellation which brings negative effects on the
heart, mainly by an atherosclerosis mechanism. Obesity, as a first rank component of the
metabolic syndrome, leads to many imbalances of the cardiac function, respectively
neurohormonal activation with hypervolemia, apoptosis, fibrosis, and hypertrophy, triggered
by the proinflammatory adipocytokines, associated with the lypotoxic effect of the fats of
myocardial cells. There have been attempts to define the relation between obesity,
cardiovascular diseases and standard biochemical investigations, observing the direct relation
between these clinical – pathological variables and the changes of preclinically parameters,
frequently used and studied in medical practice [11]. The retrospective randomized double-
blind study was conducted on a 2 years’ period of time, between 2017 and 2018, and included
two individualized groups, one that included patients with metabolic syndrome, clinically and
functionally investigated, and a control group, subject to the same evaluations.
Keywords: metabolic syndrome, obesity, atherosclerosis, fibrosis
1. Introduction
In this study we was analysed the possibility to reintroduce a minimal basic of biochemical
investigation into general population, together with the mandatory clinical objective
examination, what it can determine economic and social advantages in future (low costs and
with the involvement of general medical staff, already present), thus increasing the possibility
of easier diagnosis of cardiovascular and cardiometabolic risk. Currently, cardiometabolic
disorders are causing real havoc in the general population, with the decrease in the last years
and the age of onset, so it is necessary to develop a clinical-biochemical algorithm that will lead
to an effective primary and secondary prevention.
2. Materials and Methods
The clinical trial was conducted in the 5th Medical Geriatrics and Gerontology Clinic, C.F.
Hospital, between 2017 and 2018, with the approval of the ethics commission of hospital, and
with patients informed consent (we complied the Law 46/2003 on personal data protection and
free circulation). By this procedure, we have not studied an individual with precise identity, but
a subject with certain demographic features, who accepted to be part of our study, by signing a
free and informed consent. 285 patients with metabolic syndrome (MS) have been selected,
who have been clinically and preclinically evaluated. MS diagnosis was set based on the
association of hypertension with diabetes [13], dyslipidaemia, respectively, enlargement of the
abdominal girth [1].
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These patients were subject to the following biochemical investigations: glycaemia, total
cholesterol, HDL cholesterol, LDL cholesterol, GOT, ALT, WBC, and C-reactive protein.
The need for each analysis was justified in the context of associated cardiovascular risk
factors. We should mention the fact that conditions that grant the accuracy of determining the
values of lipid profile have been obeyed. Thus, all subjects that have been enrolled in this study
were clinically stable, and the evaluation of lipid status took place prior to individual
instructions on conditions that must be met on blood sampling. Sampling was done on
peripheral blood à jeun, after at least 10-12 h from the last mean and no alcohol consumption
during 24-48 h, in order to prevent unconcludent results [3]. Both the values of total cholesterol,
and LDL cholesterol were valid only after two determination performed at one-week interval.
If the value of the two parameters varied with more 30 mg/dl, the mean of two values was
considered. Taking into consideration the particularities of the risk factors depending on gender
and age, we separated the patients into two subgroups: subgroup I – adults, and subgroup II –
elderly (table I, fig. 1).
Table 1. Distribution of the patients with MS, on age group

Age Group I – adults Group II – elderly
18-29 years 46 (26.3%) 0
30-55 years 139 (45.6%) 0
56-75 years 0 68 (17.6%)
76-89 years 0 26 (9.3%)
> 90 years 0 6 (2.2%)
Total 165 (71.9%) 100 (28.1%)
Fig. 1. Distribution of the patients with MS, depending on gender
3. Results and Discussions
This study was focused on the analysis of the 265 patients with metabolic syndrome,
hospitalized for a period of two years in an Internal Medicine Clinic, whom we analysed
depending on two study groups, adults and elderly, on: gender, environmental origin, level of
education, smoking, alcohol consumption, and weight. Further on, we studied the incidence of
essential arterial hypertension and of sugar diabetes, completing our research with
determinations of lipid imbalance, proinflammatory state [5], hepatic and renal damage [6].
The patients of both study groups have been studied depending on age groups, namely: Study
Group I – adults: 18-30 years old, 31-54 years old, 55-64 years old; Study Group II – elderly:
65-74 years old, 75-89 years old and over 90 years old (table 1, fig 1).
The analysis of the 265 patients with Metabolic Syndrome showed a similar incidence
between the two genders (73.5% female vs. 26.5% male, fig. 2), also visible by the statistical
93
study realized on the total number of hospitalizations (13.60% vs. 14.50%). Our results did not
point out a significant statistical difference (p=0.552) on glycaemia values in patients of the
two-study group. Dyslipidaemia is a notifiable risk factor with an important role in development
of atherosclerosis and cardiovascular diseases. The disorders on the lipid metabolism can be
genetically determined or progressively installed, depending on an inappropriate lifestyle, with
excessive animal fats, stress or overstress [2, 16].
To set the diagnosis of dyslipidaemia, the following biochemical constants were determined:
serum cholesterol, HDL cholesterol, LDL cholesterol, and serum triglycerides.
Literature data indicate the fact that most of the patients with ischemic cardiopathy show
moderate increases of the total cholesterol. So far, enough evidence has been collected to testify
the relation between the anomalies of lipid metabolism and risk for coronary, cerebral or
peripheral atherosclerosis. Although high cholesterol levels in subjects with cardiovascular
diseases have been noticed long before, only in 1984 results have been published, showing the
reduction of risk for myocardial infarction as a result of lowering the value of serum cholesterol
and the body weight [24, 25]. Initially, most of the studies documented a close relation between
the level of total cholesterol and ischemic cardiac disease, but afterward there has been noticed
that in fact LDL cholesterol is the dominant causing agent with atherogenic properties,
correlated with the increase of cardiovascular risk (fig. 2).
Fig. 2. Distribution of patients with MS on mean values of LDL - cholesterol
Mean value of LDL cholesterol in Metabolic Syndrome patients: study Group I – adults –
mean value of LDL cholesterol: 116.84 mg/dL (DS=35.82); study Group II – elderly – mean
value of LDL cholesterol: 150.04 mg/dL (DS=37.82) (fig. 2). The biological reference interval
for serum triglycerides is 36mg/dL-165mg/dL. The mean value of triglycerides in MS patients:
Study Group I – adults – mean value of triglycerides: 172.02 mg/dl (DS=111.01); Study Group
II – elderly – mean value of triglycerides: 165.62 mg/dl (DS=45.98). Literature data indicate
the fact that the cardiovascular risk is more associated with mild hyperglyceridaemia than with
severe hyperglyceridaemia, probably due to the fact that mild hyperglyceridaemia is caused by
accumulation of IDL and small particles of VLDL, in plasma whereas the severe form is owed
to the accumulation of chylomicrons and large particles of VLDL, which are not atherogenic
[15].
Related to proinflammatory status, we analysed the number of leukocytes and VSH,
monitoring our patients with MS for any signs of proinflammatory status [4].
The mean value of leukocytes in SM patients was: – Study Group I – adults – mean value of
leukocytes: 6668 mm3 (DS=1660); – Study Group II – elderly – mean value of leukocytes:
6510/ mm3 (DS=2079). Mean value of VSH in Metabolic Syndrome: – Study Group I – adults
– mean value of VSH: 11.84 mm/1h (DS=8.68); – Study Group II – elderly – mean value of
94
VSH 13.77 mm/1h (DS=12.83). Out of the total of all patients with metabolic syndrome whom
we studied, we noticed the proinflammatory status, objectivized by the increase of levels of
VSH and leukocytes to 9.3% (34 patients) (34 patients) (fig. 3).
Fig. 3. Distribution of patients on means values of leukocyte
Statistic research showed changes of the proinflammatory status depending on the patients’
gender; thus, in Study Group I – adults, the percentage of women with proinflammatory status
was of 2.4% (6 patients), while in men the percentage was higher, respectively of 4.1% (10
patients). Analysing Study Group II – elderly, we discovered that 9.2% of women (11 patients)
presented proinflammatory status, while the percentage of men was lower, respectively 5.8%
(7 patients), (fig. 4).
Fig. 4. Distribution of patients with inflammatory status reported to all patients
To study the liver damage, we determined the presence of hepatic cytolysis syndrome,
measuring the level of serum transaminases: serum glutamate-oxaloacetate transaminase
(SGOT), serum glutamate-pyruvate transaminase (SGPT), and gamma-glutamic trans-
peptidase (GGT) [23]. We also added the echographia study of the liver, which revealed hepatic
steatosis or signs of chronic hepatitis.
In this context, it is necessary to undergo a population evaluation with prophylactic purpose,
by means of sanitary education and adequate treatments. A few years ago, a national program
of investigating the general health of patients was carried out thorough family doctors’ practice
[17, 18]. Using statistically significant groups of patients revealed the importance of the need
to evaluate patients every year, as well as the role of the family doctor in the intervention plan
of cardiovascular diseases, by detection and treatment of cardiovascular and metabolic risk
factors [12].
Since the national program was carried out for 1 year only, we believed it was useful to re-
introduce a basic package (clinic and biochemical) regarding the necessary investigations for a
routine evaluation [9]. They would aim at Metabolic Syndrome prophylaxis, meaning its 4
components, namely arterial hypertension [20], dyslipidaemia syndrome, mellitus diabetes and
95
obesity. The clinical evaluations, associated with the basic package of biochemical and
laboratory investigations, correlated with clinical and earlier examinations [22], with no
additional costs, could make the patients aware of the need to change their lifestyle,
supplemented by adequate and individualized treatments.
4. Conclusions
Conclusions of this research, as well of the international studies, prove the explosive
incidence of Metabolic Syndrome (27.6% versus 29.3%, adults/elderly) and contribute to a
responsible approach, aiming at prevention and adequate treatment of cardiovascular diseases
and depression disorders [10, 19]. Age remains an unchangeable major risk factor, also for
Metabolic Syndrome as our study proves, with a higher risk of onset in patients over 65 years
(RR=5.8) [14]. Increased frequency of dyslipidaemia, showed in our study, confirms that fact
that it is a treatable cardiovascular risk factor, both in adults and elderly, with a higher incidence
in older women (p=0.002).
Considering numerous current studies on the implication of proinflammatory status and the
role of immunity in etiopathology of atherosclerosis [21], our study focused on determining
these parameters in metabolic syndrome, showing a higher incidence in elderly, comparing to
adults, with statistically significant differences (p=0.001), older women being more prone to
develop it (p=0.001) [8]. A final conclusion is the idea that the metabolic syndrome represents
a very important cardiovascular risk factor in the actually society, both through the component
disorders and through the related pathologies (endocrine, neurological, psychiatric) [7], thus a
population accessible clinical and biochemical algorithm is required.
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1. Palaniappan LP, Wong EC, Shin JJ, et al., (2017). Asian Americans have greater prevalence of metabolic
syndrome despite lower body mass index. In J Obe (35), pp. 393-400.
2. 2015 Obesity collaborators GBD. (2017). Health effects of overweight and obesity in 195 countries over
25 years. N Engl J Med.
3. Ogurtsova K, Fernandes JD, Huang Y, et al., (2017). IDF Diabetes Atlas: global estimates for the
prevalence of diabetes for 2015 and 2040. Diabetes research and clinical practice 128, pp. 40-50.
4. Benrick A, Chanclon B, Micallef P, et al., (2017). Adiponectin protects against development of metabolic
disturbances in a PCOS mouse model. Proc Natl Acad Sci U S A, pp. E7187-96.
5. Jing Y, Wu F, Li D et al., (2017). Metformin improves obesity associated inflammation by altering
macrophage polarization. Mol cell Endocrinol. epub PMID 28935544.
6. Miao Y, Warner M, Gustafsson JK. (2016). Liver X receptor beta – a new player in the regulatory network
of thyroid hormone and browning of white fat. Adipocyte 5(2), pp. 238-42.
7. Stout MB, Justice JN, Nicklas BJ, et al., (2017). Physiological aging: links among adipose tissue
dysfunction, diabetes and frailty. Physiology 32(1), pp. 9-19.
8. Cheong KC, Ghazali SM, Hock LK, Subenthiran S, Huey TC, Kuay LK, et al., (2015). The discriminative
ability of waist circumference, body mass index and waist-to-hip ratio in identifying metabolic syndrome:
variations by age, sex and race. Diabetes Metab Syndr 9(2), pp. 74-8.
9. Rodica Ghiuru, Ana Minodora Grozdan, D Munteanu. (2013). Geriatrics-multidisciplinary collaboration-
an imperative. International Journal of Medical Dentistry 17 (2), pp. 117-119.
10. Salas-salvado J, Bullo M, Estruch R, et al., (2014). Prevention of diabetes with Mediterranean diet – a
subgroup analysis of a randomized diet. Ann Intern Med 160, pp. 1-10.
11. Saklayen, M.G. The Global Epidemic of the Metabolic Syndrome. (2018). Curr Hypertens Rep 20, p. 12.
12. Mikolajczyk, T.P., Guzik, T.J. (2019). Adaptive Immunity in Hypertension. Curr Hypertens Rep 21, p.
68.
13. Loperena R, Van Beusecum JP, Itani HA, et al., (2018). Hypertension and increased endothelial
mechanical stretch promote monocyte differentiation and activation: roles of STAT3, interleukin 6 and
hydrogen peroxide. Cardiovasc Res 114(11), pp. 1547-63.
14. Kachur, S., Morera, R., De Schutter, A. et al., (2018). Cardiovascular Risk in Patients with
Prehypertension and the Metabolic Syndrome. Curr Hypertens Rep 20, p. 15.
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15. Norris AL, Steinberger J, Steffen LM, Metzig AM, Schwarzenberg SJ, Kelly AS. (2011). Circulating
oxidized LDL and inflammation in extreme pediatric obesity. Obesity 19(7). pp. 1415-9.
16. Lakshmi SVV, Padmaja G, Kuppusamy P, Kutala VK. (2009). Oxidative stress in cardiovascular disease.
Indian J Biochem Biophys 46(6), pp. 421-40.
17. Jiménez MC, Rexrode KM, Kotler G, Everett BM, Glynn RJ, Lee I-M, et al., (2016). Association between
markers of inflammation and total stroke by hypertensive status among women. Am J Hypertens 29: pp.
1117-24.
18. Burrage E, Marshall KL, Santanam N, Chantler PD. (2018). Cerebrovascular dysfunction with stress and
depression. Brain Circ 4(2), pp. 43-53.
19. Catharina AS, Modolo R, Ritter AMV, Sabbatini AR, Lopes HF, Moreno Junior H, Faria AP. (2018).
Metabolic Syndrome-Related Features in Controlled and Resistant Hypertensive Subjects. Arq. Bras.
Cardiol 110(6), pp. 514-521.
20. Cӑtoi AF, Pârvu AE, Andreicuț AD, Mironiuc A, Crӑciun A, Cӑtoi C, Pop ID. (2018). Metabolically
Healthy versus Unhealthy Morbidly Obese: Chronic Inflammation, Nitro-Oxidative Stress, and Insulin
Resistance. Nutrients 01, p. 10(9).
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Health 1(2), pp. 86-88.
22. Ana Minodora Grozdan, Rodica Ghiuru, Corneliu Botez, Cristina Maria Gavrilescu, Odetta Duma,
Catalin Buzduga, Costinela Georgescu, Liliana Strat, Dragos Munteanu. (2016). Correlations Between
Cardiovascular/Cardiometabolic Risk and Standard Biochemical Investigations. Rev. Chim. (Bucharest)
67, pp. 1804-1808.
23. Hohenester S, Christiansen S, Nagel J, Wimmer R, Artmann R, Denk G, Bischoff M, Bischoff G, Rust
C. (2018). Lifestyle intervention for morbid obesity: effects on liver steatosis, inflammation, and fibrosis.
Am. J. Physiol. Gastrointest. Liver Physiol 01, 315(3), pp. G329-G338.
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97
Computational Approach for Small Data in Animal Models with

Induced Metabolic Disorders
GHICA Manuela1, BĂNCESCU Irina2, UDEANU Denisa Ioana1,

TĂEREL Adriana1, ARSENE Andreea Letiția1, ANUȚA Valentina1,
VELESCU Bruno Ștefan1, GERGHICEANU Florentina1,
MITITELU Magdalena1, IONIȚĂ Corina Ana1
1“Carol Davila” University of Medicine and Pharmacy, Faculty of Pharmacy, Bucharest (ROMANIA)
2“Costin C. Kiriţescu” National Institute for Economic Research Romanian Academy, Bucharest (ROMANIA)
Emails: manuela.ghica@umfcd.ro, irina_bancescu@yahoo.com, denisa.udeanu@umfcd.ro, adriana.taerel@umfcd.ro,
andreeanitulescu@hotmail.com, valentina.anuta@umfcd.ro, bruno.velescu@umfcd.ro florentina.gherghiceanu@umfcd.ro,
magdalena.mititelelu@umfcd.ro, corina.ionita@umfcd.ro
Abstract
In this article, we present a modern statistical method based on computational simulations

in order to characterize and compare as efficiently as possible clinical trial data having few
observations. The bootstrap technique is a statistical method that does not depend on any data
distribution hypothesis and can be considered adequate to estimate some essential parameters
in data analysis by different types of bootstrap confidence intervals. Relying on the bootstrap
method, we analyse the different influence on haematological parameters between three
substances used in experimental animal models and one negative control with the help of
confidence intervals. Statistically different in clinical research is a statement that can be
expressed through several statistical techniques. To classify and give the best interpretation of
the differences between treatments we deal with effect size and overlapping of confidence
intervals.
Keywords: simulation, bootstrap method, confidence intervals, effect size, overlapping intervals, non-normal data
1. Introduction
Most biomedical publications use descriptive and inferential statistics to obtain essential
quality and quantity conclusions for planning, financing studies and reports. For a great variety
of biomedical studies, the statistical approach is made preferential using confidence intervals
which provide the advantage of a quantitative evaluation for measuring the effect of studies,
unlike the method of hypothesis testing which provides a simple statistic signification
evaluation. Some statistical tests assume normality of data, logarithmic or not, also requiring,
at least theoretically, a volume of minimum 30 values. This last rule is in substantial conflict
with ethical and economic rules applied to various experiments, especially animal experiments
studies [1]. Even though mathematically there are various evaluation methods for obtaining the
optimal number of animals necessary for studies, for a reasonable and feasible scientific data
estimation, the classical inferential statistics have difficulties in having an entirely valid
predefined hypothesis [2, 3]. For situations where data distribution is uncertain, or the sample
number is too small for confirming a specific almost normal distribution, the study should be
evaluated with non-parametric statistical methods. These procedures do not assume over the
distribution of data. The classic non-parametrical approach is based on rank testing such as
Kruskal-Wallis, Wilcoxon or Friedman tests, but these are to be avoided because the loss of
information is essential, especially when reduced volume samples are evaluated. Another non-
98
parametric approach is computational, namely, statistical simulation [4]. Simulation using

different statistical software can reveal a specific behaviour of the biological process, and with
the help of scientific evaluation, we can associate a known statistical distribution to the data.
Simulation techniques, particularly the bootstrap method [5], enlarge standard non-
parametrical statistics and it is recommended by the international medical comities in research
including small data studies [6]. Bootstrapping technique, also called resampling, is based on
repeated random sampling with return. We obtain a dataset having values form the initial
dataset under analysis, having the same size (e.g., the same individual can be encountered
multiple times in the bootstrap set and also in any bootstrap sampling). This method is based
on the hypothesis that the initial set of data is randomly extracted from an infinite population
and is representative of the same population. The probabilistic information contained in the
random sample from the original dataset reflects all the information corresponding to the
original population from which it was extracted. Also, the number of bootstrap re-allocations,
denoted with r, is variable, but it is generally at least 1,000. Each computer-generated
resampling mimics the distribution of the entire population, and so we can conclude the
probabilistic behaviour of the entire population [7]. In clinical pharmacology research, the non-
parametric bootstrap approach was used in individual or population bio-equivalence to
determine a confidence interval for the mean difference of two treatments [7]. However, values
contained in samples may represent both measurements of the AUC or maximum
concentrations [8]. Research based on bio-equivalence bootstrap simulations has been done in
line with international guidelines approved by the FDA [9].
The bootstrap method is a modern statistical technique with essential preclinical and clinical
applications on studies based on small data like those restricted by general regulations of Ethical
Committee and guidelines [10-14]. Bootstrap method is used both in constructing confidence
intervals for any parameter that occurs in the statistical model: mean, truncated mean, bias,
median, standard deviation, dispersion, standard error [15, 16] and for estimating regression
coefficients when errors are not distributed normally [17] or for proportionally hazards models
[18]. Also, the bootstrap method can be applied to discover those atypical, inconsistent or
merely different values in a dataset. The influence of aberrant values in parameter estimation
increases as the data set is restricted, involving making erroneous decisions and inaccurate
predictions. For example, a bootstrap tool bootlier can be used for detecting outliers. If this
instrument, based on the construction of an empirical bootstrap histogram distribution
associated with a truncated number of values is multimodal, the presence of an aberrant value
in the dataset is signalled [19].
The study aims to highlight the importance and utility of the modern statistic in case of small
data experiments according to Ethical Committee recommendations using the example of three
different treatments (phenylhydrazine, deferoxamine and alloxan) with variable effect on some
haematological parameters.
Nowadays, phenylhydrazine is frequently used in animal models for inducing haemolytic
anaemia in preclinical studies [20, 21]. The mechanism of action is based on inducing an
oxidative stress in red blood cells by stimulating the lipoperoxidation process with toxic effects
on cell membranes [20, 21]. The effect is correlated with variations on haematological analyses
like fast decreasing of the red blood cells number (RBC) and haematocrit (HCT), increasing of
the haemoglobin (HGB) level and fluctuation of the erythrocyte indices: mean cell volume
(MCV) and mean cell haemoglobin (MCH).
Deferoxamine is a drug frequently used in chelation therapy for binding iron and aluminium
[22-26]. In experimental animal models, deferoxamine is used for inducing iron deficiency
anaemia characterized by a decrease of the RBC level, haematocrit, haemoglobin, MCV and
MCH. Still, the results are controversial because of the interindividual response variations.
99
Alloxan is a pyrimidine derivate used for inducing diabetes mellitus in animal models [27-
29]. In short term administration, the substance has toxic effect on pancreatic β-cells with direct
effect on glucose metabolism and less influence on haematological parameters [28-30].
The study considers the haematological analyses results obtained after the treatment with the
three substances used in different animal models to construct the bootstrap confidence intervals
for the mean parameter using the R simulation program and to determine a measure of the
effects to highlight the differences between treatments.
Experimental animal model

The animals used for the experiment were 25 Wistar rats weighing 180±10g purchased from
the Animal Biobase of the University of Medicine and Pharmacy “Carol Davila”, Bucharest.
The animals were kept in standard laboratory conditions, received food twice a day and water
ad libitum. The experiment was performed in compliance with the European Communities
Council Directive 2010/63/UE and Law No. 43 of the Romanian Parliament from 11th April
2014.
Animals were randomly distributed in four groups as it follows:
Group 1 (n=6) was treated daily for 3 days with phenylhydrazine administrated
intraperitoneal in a dose 40mg/Kg bw for 3 days. Blood samples were collected on day 4 of the
study.
Group 2 (n=6) was treated daily for 7 days with deferoxamine administrated intraperitoneal
in a dose 200mg/Kg bw. The treatment was associated with a low iron diet for the whole period
of the experiment. Blood samples were collected on day 8 of the experiment.
Group 3 (n=6) was treated with alloxan in a single dose of 120mg/Kg bw, administrated
intraperitoneal. Blood samples were collected after 5 days from the administration.
Group 4 (n=7) was considered the negative control group. The blood samples were collected
in the first day of the experiment.
For blood collection the animals were anesthetized with ether ethylic and the blood samples
were collected on K3-EDTA tubes generally used for haematological analyses.
The haematological parameters were determined using Abacus Junior haematological
analyser (Diatron GmbH, Austria). The following parameters were considered for the study:
red blood cell count (RBC), haemoglobin, haematocrit, red blood cell indices (mean
corpuscular volume – MCV, mean corpuscular haemoglobin – MCH).
Statistical analyses
Statistical analyses were performed using GraphPad Prism version 7.00 for Windows, and
the R Project for statistical computing version 3.5.1. for Windows.
3. Results and Discussion
The blood specimen collected from the animals were used to test the error margins and the
differences between 3 treatments that are supposed to induce significant variation of some
haematological parameters. Thus, the study is based on the results obtained from a total of 4
animal groups corresponding to a control group and three treated groups with testing drugs
alloxan, phenylhydrazine, and deferoxamine and for each group was determined 5
haematological parameters. The first step of the study was to verify the data normality
hypothesis. This analysis was performed using the GraphPad Prism statistical program.
100
Surprisingly, out of the 20 samples, only one sample did not respect the hypothesis of
normality, and apparently, none was appropriate in terms of the minimum number of
individuals imposed in the hypotheses of inferential statistics.
In order to illustrate the role of the number of individuals necessary for a correct
interpretation of the normality of a sample under research we have chosen to exemplify some
distributions commonly found in medical sciences: normal, exponential and Weibull (Fig. 1).
The two-parameter Weibull distribution is frequently used in biomedical sciences for
modelling survival data. The exponential distribution is a left-skewed model, having a
decreasing density function, while Weibull distribution even though it resembles a normal
model for some values of the parameters, it is not asymmetric model, as opposed to the latter.
Fig. 1. Normal, exponential and Weibull density functions for different values of the parameters
Using the simulation software R, 10,000 samples, each of size n=6, were generated for each
of the probability law shown above. Applying the Shapiro test of normality, the percentage of
samples normally distributed was obtained. We notice that for n=6, this number is very high in
every situation, surprising for non-normal and asymmetric distributions. Thus, in the case of
exponential distribution, we obtained 78% of samples of size n=6 that proved to be normal and
94% in the case of Weibull distribution.
If the sample volume increases and exceeds the threshold of 30 individuals admitted in
statistical assumptions, it is evident that the tendency is in the correct estimation of the
population distribution from which each sample originates (Fig. 2). Thus, a nonparametric
bootstrap method, based on random reassignment that overrides the initial sample, is required.
Fig. 2. Percentage of normality of data as the number of sample sizes n increases, obtained for normal,
exponential and Weibull distributions considering different values of the parameters
101
Confidence Intervals
Confidence intervals (CIs) are tools frequently used in medicine, biology, and pharmacy,
being used since 1980. In the specialized literature, these are recommended to be reported
alongside the statistical estimators considered, for a better interpretation of results [31, 32].
CIs respond to questions such as “how big is the effect of a treatment”, “how intense it is...”.
Confidence intervals represent a way of estimating the parameters of a population and are
extremely useful in finding the magnitude of values relative to the mean corresponding to a
certain level of significance, for example, 5%. Confidence intervals, more than statistical
hypotheses testing that offer dichotomous responses, are useful in comparing the means of two
treatment groups on a particular scale, whereby we can also issue estimates of the resulting
differences. In a graphical representation of confidence intervals relative to the same parameter,
the aim is to observe whether these intervals overlap. We can conclude that the means of the
two treatment groups are or not statistically different at a level of significance chosen of 0.05.
If the two confidence intervals do not overlap, we can say that the means are significantly
different.
The problem that arises is how we interpret confidence intervals which partially overlap.
The statistical theory asserts that a null hypothesis on the equality of two sample means is
rejected at a level of 0.05 if the z-test is greater than 1.96. This assertion implies that if the two
95% confidence intervals overlap by less than 29%, then we will reject the null hypothesis of
the equality of two means at a 0.05 level. The 29% level was calculated in a situation where the
two selections have the same size and equal, known variances. For any other configuration, the
percentage is visibly sensitive, mainly depending on the size of the sample.
The second aspect of confidence intervals is the discussion of differences in a sample mean
of two treatment groups. We can issue reasoning similar to a null statistical hypothesis testing,
i.e., the means are significantly equal with a significance level of 0.05 if zero is one of the
plausible value of the confidence interval, otherwise if zero is outside the range, then the sample
means are significantly different at the significance level of 5%.
Even though CIs can be utilized as null hypothesis tests, the best interpretation of them
derives from their definition. For an estimator 𝜃̂, a symmetric 100(1-α) % CI can be written as
equation (1):
(𝜃̂ − 𝑡1−𝛼 𝜎𝜃̂ , 𝜃̂ + 𝑡1−𝛼 𝜎𝜃̂ ) (1)
2 2
where 𝜎𝜃̂ is the standard error of 𝜃̂, and α is the risk we are willing to take, meaning
𝛼 𝛼
𝑃 (𝑡 ≤ 𝑡1−𝛼 ) = 1 − 2 and 𝑃 (𝑡 ≤ 𝑡𝛼 ) = 2 . Given a dataset, a CI provides us with a range of
2 2
values which are plausible to be the true value of the parameter. The values outside the interval
are unlikely, but not impossible to be the true value of the parameter. As a general rule: the
shorter the interval, the better. We can also interpret the CI given risk α=5%, as being 95% sure
that the real value of the parameter lies within this interval. For small datasets, bootstrap CIs
are constructed.
Many bootstrap confidence intervals depend on efficiency and length, having different
characteristics. A proper bootstrap interval, if an exact one can be obtained, should be close to
that one. If this is not possible, a good bootstrap interval should have high probabilities of
coverage. Taking account of all these, in the specialized literature, various bootstrap confidence
intervals have been introduced: percentile, expanded percentile, bootstrap t tables, and bias-
corrected and accelerated. Out of all these CIs the most utilized, are the last ones, which we
present next.
In this paper, we apply the bootstrap CIs for testing the statistically significant advantage of
a treatment in anaemia problems. We apply the 𝐵𝐶𝑎 intervals, meaning bias-corrected and
102
accelerated CIs. These intervals depend on the acceleration parameter 𝑎̂ and on the bias-
correction factor 𝑧̂0 , and are given by equation
∗ ∗
[𝜃(𝛼1)
, 𝜃(𝛼2)
] (2)
𝑧̂+𝑧 𝑧̂+𝑧
Where 𝛼1 = Ф(𝑧̂0 + 1−𝑎̂(𝑧
0 𝛼
̂+𝑧 )
) and 𝛼2 = Ф(𝑧̂0 + 1−𝑎̂(𝑧
0 1−𝛼
) and Ф(∙) is the CDF of
0 𝛼 ̂+𝑧 0 1−𝛼 )
standard normal distribution N(0,1), while 𝑧𝛼 is 100αth percentile of standard normal. The bias-
correction factor 𝑎̂ rectifies the assumption that the standard error of the estimator is the same
for each θ, estimator 𝜃̂ being normally distributed.
Interpretation
For the present study, we constructed 𝐵𝐶𝑎 CIs for means considering five blood parameters,
namely, RBC, HGB, HCT, MCV and MCH. In Fig. 3A-3F these CIs are displayed. In these
figures, the number one is associated with the control group, while number two is for each drug
in particular. For treatments alloxan we notice a considerable overlap of the CIs for all the five
blood tests. This finding suggests that between this drug and the control one there is no
statistical difference. For treatment with deferoxamine we have overlapping of the CIs only in
the cases of RBC, HGB, HCT. However, between phenylhydrazine and control, there is a
statistical difference for some blood tests. This graphical analysis of the drug results suggests
that only phenylhydrazine is statistically different from the control group.
Fig. 3A-3F. Overlapping confidence intervals
Fig. 3A. Control vs Alloxan – HGB, MCH, RBC Fig. 3B. Control vs Alloxan – HCT, MCV
103
Fig. 3C. Control vs Deferoxamine – HGB, MCH, RBC Fig. 3D. Control vs Deferoxamine – HCT, MCV
Fig. 3E. Control vs Phenilhydrazine – HGB, MCH, Fig. 3F. Control vs Phenilhydrazine – HCT, MCV
Effect size
Testing the null statistical hypothesis at a certain level of significance is one of the most
common methods of statistical inference. Unfortunately, it is a too simplistic approach, in terms
of yes or no, generating great uncertainty. Statistical results must necessarily describe the
magnitude of the expected effect for each treatment and the accuracy of the estimate of the
effect. Using the confidence intervals and calculating of the effect sizes between different
treatments complete the poor interpretation of null statistical hypothesis testing. The
combination of the two statistical tools highlights the link between the measured values in
various treatments much more useful than the significance level of a statistical hypothesis and
also allows the comparison of the test results with other studies to facilitate their incorporation
into a meta-analysis [33].
Calculation of effect size can be achieved by several approaches depending on the type of
data (quantitative or qualitative) and the number of treatments used in the study (2 or more).
In this paper, we apply 𝑑𝑟 measure being a robust version of Cohen’s index [34]. A Cohen
size effect greater than zero indicates the degree to which treatment is more effective than the
other, and as a general rule 0.2 is considered to be small, 0.5 as medium and 0.8 as high. A
104
negative Cohen effect size indicates the effect decreases the mean, and a positive effect size
indicates that the effect increases the mean. The effect size (ES) is an inferential statistical
measure that implies a degree of uncertainty, which is why it is also recommended to be
estimated by a confidence interval. An effect size tells us its magnitude, but only if it exists.
The magnitude of this effect is displayed in Table 1.
Table 1. 𝑑𝑟 Effect size for the mean difference between treatment and control groups
𝑑𝑟 index RBC HGB HCT MCV MCH
Alloxan -0.81 -0.20 -0.66 -0.13 0.48
Deferoxamine 0.93 0.11 -1.33 -3.36 -0.85
Phenylhydrazine -15.65 4.23 -15.58 -1.73 9.98
The effect is negative for RBC meaning it decreases the mean. Phenylhydrazine has an effect
of -15.65 for RBC blood test which suggests a very large effect. After the treatment with
Alloxan, we concluded that there is no effect on haematological parameters, in case of
deferoxamine partially and only phenylhydrazine has an effect.
4. Conclusions
In this article, we used a highly useful computational statistical method to build confidence
intervals for common statistical parameters when assumptions for applying the parametric
estimation are not met. Our goal was to introduce examples of the bootstrap method and also
to provide a modern interpretation of confidence intervals with statistical concepts overlapping
and effect size for a deeper understanding of the estimation phenomenon. The method is useful
in clinical and preclinical studies with small or limited access to data for the evaluation of
pharmacological response or identification and estimation of potential significant secondary
effects. These approaches complement the statistical methods already used in practice and lead
to the integration of several comparative studies into a current meta-analysis.
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106
The Relationship between Novel Haematological Markers and

Microvascular Complications among Patients with Diabetes in
Sibiu County
SITTERLI-NATEA Carmen-Narcisa1,2, CALUTIU Nicoleta-Georgiana2

1“Victor Papilian” Faculty of Medicine, Sibiu (ROMANIA)
2Center of Diabetes, Academic Emergency Hospital, Sibiu (ROMANIA)
Emails: narcisa_20@yahoo.com, minea_n@yahoo.com
Abstract
Introduction
The assessment of novel inflammatory markers on diabetic complications between admitted
in Academic Emergency Hospital Sibiu.
Method
We evaluated the association between neutrophil/lymphocyte ratio (NLR),
platelet/lymphocyte ratio and mean platelet volume with diabetes microvascular complications
among patients hospitalized in the Academic Emergency Hospital Sibiu between 01 January
2018-31 July 2018. 100 of the patients admitted in hospital during this period were included in
the study. 13 of these patients had type 1 diabetes mellitus.
Results
Neuropathy showed a significant association with PLR (p=0.021) and retinopathy was found
to be significantly correlated with MPV (p=0.010). In contrast, with the two other complications
(nephropathy and peripheral artery disease), no significant association could be detected.
Conclusions
Knowing that diabetes is a chronic condition with a high impact on the health system, it is
necessary to find new and inexpensive markers that can help predict and manage diabetic
complications from the onset of the disease.
Keywords: neutrophil/lymphocyte ratio (NLR), platelet/lymphocyte ratio, mean platelet volume (MPV),
diabetes microvascular complications
1. Introduction
Diabetes mellitus (DM), the most common systemic metabolic disorder, is characterized by
chronic hyperglycaemia due to inadequate secretion and/or use of insulin. Over time diabetes
can lead to complications (chronic injury and disfunction affecting particularly the heart, blood
vessels, eyes, kidneys, and nerves) that increase the overall risk of dying prematurely. In
pregnancy, poorly controlled diabetes increases the risk of fetal death and other complications
[1]. Compared to type 1 DM, in which complications appear after 5 or more years of evolution,
dependent of glycaemic control, in type 2 DM complications can be present at the moment of
diagnosis.
Inflammatory processes have a major role in the appearance and progression of both diabetes
mellitus and it’s micro- and macro-vascular complications [2]. The mechanisms underlying
107
these changes involve the interaction between inflammation and oxidative stress [3]. One of the
findings of numerous studies was an increase in white blood cell, this leading to the study of
novel inflammatory markers derived from standard blood count test, such as neutrophil to
lymphocyte ratio (NLR) [3, 4]. Another important marker for the worse outcome of diabetes
that can be easily accessible from a complete blood count was found to be the increased mean
platelet volume (MPV), with a recommended value of 9.55 fl as an indicator of poor
glucoregulation [5, 6].
Many studies have shown a significant association between MPV and the evolution of type
2 diabetes mellitus, a higher MPV being associated with a more severe outcome. Kadić et al.,
consider type 2 DM “the most common acquired thrombophilia” because of its prothrombotic
state [6]. In their study Papanas et al., Found a positive correlation between elevated MPV and
retinopathy and nephropathy [7]. Since 2014 Ulutas et al., found an association between MPV
and HbA1c, proposing MPV as prognostic marker for atherosclerosis in patients with type 2
DM [8]. Numerous factors can influence platelet and MPV values, but some researchers
indicate that MPV should always be assessed together with platelet count [5].
NLR represents a well-established marker of inflammation, present in many chronic
conditions, with a significant role in predicting the short- and long-term mortality for
cardiovascular diseases. In diabetic patients was found to be significantly elevated when
compared to healthy controls [4]. In addition, Xu et al., [9] reported higher NLR values in
patients with diabetic peripheral neuropathy (DPN) and that NLR was an independent risk
factor for DPN. Similarly, Fawwad et al., showed that NLR was significantly higher in diabetic
subjects with retinopathy and neuropathy [10].
AIM
The aim of this study was to evaluate the relationship between these parameters obtained
from complete blood count and microvascular complications in diabetic patients hospitalized
in the Academic Emergency Hospital Sibiu.
2. Method
The study included 100 patients with type 1 and type 2 diabetes mellitus hospitalized in the
Academic Emergency Hospital Sibiu, between 01 January 2018-31 July 2018. Data were
retrospectively collected from medical records for metabolic characteristics and laboratory
results. Exclusion criteria were the presence of coronary artery disease, haematological
conditions, malignancy, severe hepatic disorder, severe renal disorder and current smoking.
Venous blood sample was collected on fasting conditions during hospitalisation. For the
patients admitted through the Emergency room, laboratory results at discharge were used in the
analysis. Serum levels of haemoglobin (Hb), haematocrit (Hct), lymphocytes, neutrophils,
platelets, MPV, serum iron, alanine aminotransferase (ALT), aspartate aminotransferase (AST),
γ-glutamyl transpeptidase (GGT) and uric acid levels were judged based on the upper limit of
the reference normal range according to the used kits and gender. High lipid profile was defined
as >202 mg/dl for cholesterol, >65 mg/dl for HDL-cholesterol, >100 for LDL-cholesterol, >140
mg/dl for triglycerides; and low lipid profile was considered at <109 mg/dl for cholesterol, <45
mg/dl for HDL-cholesterol, <100 for LDL-cholesterol, <45 mg/dl for triglycerides. Renal
function was assessed using glomerular filtration rate obtained by MDRD Equation, based on
creatinine value and patient characteristics [11]. Fasting blood glucose and HbA1c were
measured at admittion. Glycaemic control was classified into uncontrolled if HbA1c >7% and
controlled if HbA1c ≤7%. Body mass index (BMI) was calculated, then classified as: grade I
obesity (30 – <34.9 kg/m2), grade II obesity (35 – <39.9 kg/m2) and grade III obesity (≥40
108
kg/m2). BMI ≥25-29.9 was defined as overweight, BMI ≥19-25 was defined as normal and
BMI <19 was defined as underweight.
Patients with glomerular filtration rate <90 ml/min/1.73 m2 were classified as having diabetic
nephropathy, diabetic retinopathy was diagnosed in the Department of Ophthalmology, while
diabetic neuropathy was considered in patients who had either neurologic alterations at foot
exam or findings of polyneuropathy or neuropathy in EMG.
Data about the presence or absence of peripheral artery disease were extracted from patient
file.
Statistical analysis (average values, p values, independent samples t-tests or χ 2 tests) were
calculated using IBM SPSS Statistics 25. A p value of less than 0.05 was considered significant.
3. Results
General characteristics of patients

13 patients had type 1 diabetes (9 men and 4 women), and 87 had type 2 diabetes mellitus
(39 men and 48 women). Of the type 2 diabetics, 17 patients were on diet, 26 received oral
therapy and/or GLP-1 agonists, 20 received oral therapy and insulin and 24 received only
insulin therapy (Table 1).
Table 1. Distribution of diabetes type and treatment between men and women
Type 1 DM Type 2 DM
Men Women Total Men Women Total
Number of patients 9 4 13 39 48 87
Diet - - - 9 8 17
Oral therapy - - - 14 12 26
Oral therapy+insulin - - - 7 13 20
Insulin 9 4 13 9 15 24
The mean age of the patients was 56.26±14.79 years (min = 19; max = 89). When reported
to gender, mean age for women was 60.69±13.81 years (min = 28; max = 89) and for men
51.46±14.32 years (min = 19; max = 79) (Table 2).
There were 48 men, most of them in the decades 40 -60 (27, 15 respectively 12 patients).
Of the 52 women, 40 were aged between 50-80 years. The largest proportion (67%) of
patients were >50 years old. When comparing the patients according to the place of residence
we found 62 patients (62%) living in urban areas, and 38 patients in rural areas (Table 2). Of
the people living in urban areas, 32 were men and 30 were women. 16 patients living in rural
areas were men and 22 were women.
Table 2. Gender distribution of demographic characteristics

Men Women Total
Gender (no. of patients) 48 52 100
Age 51.46±14.32 60.69±13.81 56.26±14.79
Urban 32 30 62
Place of residence
Rural 16 22 38
BMI was 29.54±7.04. We found 71 patients with overweight and obesity, and only 2 patients
underweight. Even though we found no association between BMI and gender, in these patients
we observed that women had a higher rate of obesity (61.5% compared to 35.4%), while
overweight was more frequent in men (27% versus 17.3%) (Table 3). 3 patients had severe
obesity, with a BMI >50 (kg/m2), 2 of them were men.
109
Table 3. Distribution of patients according to BMI

Number
BMI (kg/m2)
Men Women
<19 2 - 2
19-24.9 16 11 27
25-29.9 13 9 22
30-34.9 11 16 27
35-39.9 3 12 15
>40 3 4 7
Clinical laboratory findings and statistical analysis

Of all 100, only one patient had all laboratory results within the normal range. Number of
patients who presented either lower or higher values for the investigated parameters, as well as
mean value and standard deviation are presented in Table 4.
Table 4. Descriptive statistics of laboratory results

Number of patients
Minimum Maximum Mean±SD
abnormal values
Fasting glucose 64 50 633 197.92±113.17
Hb A1c >7 82 4.5 16 9.80±2.52
Haemoglobin 16 8.3 17.1 14.18±1.58
Haematocrit 15 26.9 51.1 41.62±4.21
Neutrophils 11 1.26x103 11.73 4.78±1.63
Lymphocytes 9 0.78 x103 4.99 x103 2.45±0.80
Platelets 6 74x103 523x103 250.46±66.90
GFR 50 32.3 245.7 91.46±30.57
AST 10 7 133 24.17±16.92
ALT 18 6 156 31.76±20.70
GGT 54 10 403 50.35±51.72
Uric acid 21 1.4 13.8 5.19±2.06
Cholesterol 43 91 573 207.19±60.36
HDL-cholesterol 47 8 79 45.65±13.43
LDL-cholesterol 56 18.8 215.8 113.85±41.18
Triglycerides 66 48 1377 243.41±237.53
Serum iron 25 20.1 204.3 83.9±34.96
Median (IQR)
NLR 1.90 (5.59) 0.65 6.24 2.14±1.01
PLR 106.45 (281.86) 37.37 319.23 112.75±46.97
MPV 11.00 (4.1) 9.1 13.2 11.06±0.84
The mean NLR in diabetics was 2.14, having a positive and significant correlation with PLR
(p<0.000), neutrophils (p<0.000) and fasting glucose (p=0.007)), while with lymphocytes the
correlation was negative (p<0.000). Mean PLR was 112.75 and correlated positively with
NLRm (p<0.000), platelets (p<0.000), BMI (p=0.047) and HDL-cholesterol (p=0.002). The
correlation with lymphocytes was strong, but negative, same as NLR (p<0.000). Mean MPV
was 11.06, but when the cut-off range 9.55 fl suggested by Kadić et al., was applied in patients
with Hb A1c >7, mean value raised to 11.15±0.76. MPV only correlated negatively with
platelets (p=0.022).
Multivariate ANOVA was used to investigate the associations of diabetic complications
(neuropathy, retinopathy, nephropathy and peripheral artery disease) with NLR, PLR and MPV.
Statistical significance was considered at p<0.05. Neuropathy showed a significant
association with PLR (p=0.021) and retinopathy was found to be significantly associated with
MPV (p=0.010). In contrast with the two other complications (nephropathy and peripheral
artery disease), no significant association could be detected (Table 5).
110
Table 5. The association of diabetic complications with MPV, NLR, and PLR
Neuropathy Number Mean Standard deviation P value
MPV (fl) Present 61 11.02 0.8486 0.199
Absent 39 11.12 0.8009
NLR (%) Present 61 2.15 0.9008 0.294
Absent 39 2.12 1.1588
PLR (%) Present 61 106.23 122.95 0.021
Absent 39 37.945 56.3775
Retinopathy
MPV (fl) Present 24 11.31 0.7235 0.010
Absent 76 10.98 0.8475
NLR (%) Present 24 2.15 0.9039 0.528
Absent 76 2.14 1.0404
PLR (%) Present 24 104.49 44.7605 0.236
Absent 76 115.36 47.0442
Nephropathy
MPV (fl) Present 51 11.16 0.7730 0.440
Absent 49 10.96 0.8759
NLR (%) Present 51 2.19 0.9519 0.594
Absent 49 2.10 1.0638
PLR (%) Present 51 112.49 40.3154 0.869
Absent 49 113.03 52.5937
Peripheral artery
disease
MPV (fl) Present 15 10.78 0.9873 0.658
Absent 85 11.11 0.7906
NLR (%) Present 15 2.49 1.1670 0.934
Absent 85 2.08 0.9658
PLR (%) Present 15 114.67 44.4423 0.176
Absent 85 112.42 47.1227
4. Discussion
The association between neutrophil/lymphocyte ratio (NLR), platelet/lymphocyte ratio and

mean platelet volume with many chronic conditions, diabetes mellitus included, and their role
in disease progression has been clearly established [2, 4, 5, 6]. They can also be easily measured
by a simple peripheral blood count, which is simple and less expensive than calculating other
inflammatory cytokines [10].
In our study we included also insulin treated patients, on the basis of their increasing number
among diabetic patients. We found a slightly lower MPV in these patients (10.87 compared to
11.06), probably due to anti-inflammatory and protective effects of insulin.
Type 1 diabetic patients with a duration of diabetes longer than 5 years were not excluded.
Similarly, their mean MPV was lower than in type 2 diabetic patients (10.59 versus 11.13).
We found a positive correlation between PLR and neuropathy, and between MPV and
retinopathy, consistent with the data reported in literature [2]. No other correlation with diabetic
complications was found.
5. Conclusions
Knowing that diabetes is a chronic condition with a high impact on the health system, it is
necessary to find new and inexpensive markers that can help predict and manage diabetic
complications from the onset of the disease.
111
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112
The Oral Antidiabetic Treatment in Patients with Type 2 Diabetes

Mellitus and Peripheral Artery Disease
DIACONU Camelia Cristina1,2, HORODINSCHI Ruxandra-Nicoleta1,2,

BRATU Ovidiu Gabriel1,3, BACALBASA Nicolae1,4, ILIESCU Laura1,5,
STANESCU Ana Maria Alexandra1
2 Clinical Emergency Hospital, Bucharest (ROMANIA)
3 Emergency University Central Military Hospital, Academy of Romanian Scientists, Bucharest (ROMANIA)
4 “I. Cantacuzino” Clinical Hospital, Bucharest (ROMANIA)
5 Fundeni Clinical Institute, Bucharest (ROMANIA)
Abstract
Diabetes mellitus (DM) represents a major risk factor for all types of cardiovascular disease,
including peripheral artery disease (PAD). Cardiovascular events are the most important cause
of mortality in patients with diabetes mellitus. About one-third of patients with PAD have also
diabetes. The prevalence of PAD can be underestimated in patients with DM, because they are
often asymptomatic until they progress to advanced disease and because of the simultaneous
diabetic neuropathy. In patients with diabetes, the distal arteries, below the knee, such as
popliteal, anterior or posterior tibial, peroneal arteries, are usually affected. Typical claudication
is less frequent in patients with DM, so many diabetic patients are asymptomatic a long time,
they are diagnosed in more advanced stages and have a worse prognosis. The control of glucose
blood level to maintain HbA1c less than 7% is necessary in patients with DM and PAD to
reduce the complications. The main treatment in type 2 DM is represented by oral antidiabetic
drugs, so 56.9% of the patients with type 2 diabetes receive oral antidiabetic agents. Sodium-
glucose co-transporter 2 inhibitors (SGLT2-i) can decrease the rate of cardiovascular events,
including mortality, in patients with DM and cardiovascular disease. At the moment, SGLT2-i
are the most effective and promising oral antidiabetic drugs for patients with DM and PAD.
Glucagon-like peptide-1 receptor agonist (GLP-1), liraglutide and semaglutide, also reduce
the rate of cardiovascular events. Dipeptidyl peptidase-4 inhibitors (DPP-4) have pleiotropic
effects, such as improving endothelial dysfunction, reducing blood pressure and inflammation
and may have a protective effect against cardiovascular disease, to delay the progress of
atherosclerosis and decrease the risk of PAD.
DPP-4 are particularly useful in association with metformin. Thiazolidinediones have an
important effect in preventing cardiovascular disease by improving insulin sensitivity in
peripheral tissues. Pioglitazone may reduce atherosclerosis by improving insulin resistance and
decreasing systemic inflammation which are involved in atherosclerotic plaque formation.
Pioglitazones reduce the risk of developing cardiovascular events – myocardial infarction,
stroke, compared to placebo in patients with clinically manifest vascular disease. In conclusion,
good control of glucose blood levels in patients with DM can reduce significantly the risk of
developing PAD.
Keywords: type 2 diabetes mellitus, peripheral artery disease, oral antidiabetic agents
113
Introduction
Diabetes mellitus (DM) represents a major risk factor for all types of cardiovascular disease,
including peripheral artery disease (PAD) [1-3]. Cardiovascular events are the most important
cause of mortality in patients with diabetes mellitus [4]. Patients with both DM and PAD have
a higher risk to develop acute cardiac events and acute limb ischemia and amputation [5]. PAD
can lead to severe disability and affects the quality of life in patients with DM [6, 7].
Prevalence of PAD in Patients with DM
Diabetes is a highly prevalent disease. Worldwide, about 170 million people have diabetes
and its prevalence is increasing [8]. Approximately one-third of patients with PAD have also
DM [6, 7]. The prevalence of PAD can be underestimated in patients with DM, because they
are often asymptomatic until they progress to advanced stages of the disease and because of the
simultaneous diabetic neuropathy [9-11]. Even patients with dysglycemia have an impaired
ankle-brachial index – less than 0.9 compared to those with normal glucose blood levels [12].
Patients with DM and PAD often complain about rest pain and they present ulceration or
gangrene in the lower limb. Another interesting point is that about half of the patients with
chronic limb-threatening ischemia of one limb, develop limb-threatening ischemia also in the
other lower limb during the next 5 years [13].
In diabetic patients, the distal arteries below the knee are most commonly affected [14, 15].
Diabetic patients have a 4-fold higher risk of limb-threatening ischemia [16].
Risk Factors for PAD
DM is an important risk factor for PAD. Other risk factors are smoking, dyslipidaemia,
hypertension, coronary artery disease.
In patients with DM, there are specific risk factors for PAD. The duration of DM is related
to the incidence and extent of PAD, so patients with a longer duration DM have an increased
prevalence of PAD and the illness may extend to several arteries [17]. The severity of DM is
another risk factor, a 1% increase in HbA1c is related to a 30% higher risk of developing PAD
[17]. Women with DM develop claudication more frequently than men with DM [18, 19]. In a
review of the Framingham study, DM with glycosuria increased the risk of claudication 9-fold
in females and 3.5-fold in males [20]. Regarding the race, diabetic African Americans and
Hispanics white people have a higher risk to develop PAD than non-Hispanic whites [5, 7].
Peripheral neuropathy is often associated with PAD and patients may perceive the symptoms
of neuropathy as vascular pain [6].
Clinical Manifestations
Patients with PAD may be asymptomatic or may present symptoms as intermittent

claudication, which appears with physical exercise, rest pain, limb ulceration, and gangrene.
The symptoms usually appear due to the stenosis or occlusion of the arteries, but can also be
caused by thrombosis or embolism of unstable atherosclerotic plaque.
Typical claudication is less frequent in patients with DM, so many diabetic patients are
asymptomatic a long time, they are diagnosed in more advanced stages and have a worse
prognosis [21, 22]. DM is related to more severe distal PAD, below the knee, located at
popliteal, anterior or posterior tibial, peroneal arteries compared to smoker patients, who have
proximal PAD in aorto-ilio-femoral arteries [14].
114
The Effectiveness of Oral Antidiabetic Agents in Patients with Type 2 DM and PAD
The control of glucose blood level to maintain HbA1c less than 7% is necessary for patients
with DM and PAD to reduce the occurrence of the complications [23].
The majority of patients with DM receive oral antidiabetic agents – 56.9%, in comparison
with those who receive oral drugs and insulin – 14.7% and those treated only with insulin –
14% [24].
The therapy with metformin may decrease the rate of cardiovascular morbidity or mortality,
but the effect regarding PAD is controversial [25]. According to several studies, metformin
reduces the risk of developing PAD, but the reduction is not significant [25].
Sodium-glucose co-transporter 2 inhibitors (SGLT2-i) can decrease the rate of
cardiovascular events, including mortality, in patients with DM and cardiovascular disease.
SGLT2-i block the sodium-glucose cotransporter in the kidney tubules, leading to glucose
excretion. Several trials showed the effectiveness and safety of SGLT2-i in patients with DM
and PAD. The CANVAS trial (CANagliflozin cardioVascular Assessment Study) analysed the
effectiveness of canagliflozin. It revealed that in patients who received canagliflozin the
amputations were below the ankle – toe, trans-metatarsal and occurred more often in patients
with a history of PAD or with a prior amputation [26-28]. The amputations occurred 1.5 years
after the initiation of the treatment with canagliflozin [24].
The EMPA-REG Study included 1461 people with PAD who were treated with
empagliflozin, which proved to reduce cardiovascular and all-cause mortality rates [27].
Regarding amputations, there was no important difference between patients treated with
empagliflozin compared with placebo [29]. The prevalence of PAD of about 20% was similar
in both trials, CANVAS and EMPA-REG [24]. Canagliflozin and empagliflozin have proved
similar rates in reducing cardiovascular events, but in patients with treated with canagliflozin it
was noticed an increased risk of lower limb amputation. The majority of the amputations were
below the ankle, as mentioned before [24].
DECLARE-TIMI 58 trial included 17160 patients with type 2 DM who had or were at risk
to develop atherosclerotic cardiovascular disease, who received dapagliflozin versus placebo.
Patients who received dapagliflozin had the same rate of major adverse cardiovascular
events (myocardial infarction, cardiovascular death, ischemic stroke) as the placebo, but had a
lower rate of cardiovascular death or hospitalization for heart failure [30]. At the moment,
SGLT2-i are the most effective oral antidiabetic drugs for patients with DM and PAD.
Glucagon-like peptide-1 receptor agonist (GLP-1), liraglutide and semaglutide, lower the
rate of cardiovascular events. Liraglutide decreases the risk of stroke and myocardial infarction
by about 13% in patients with established high cardiovascular risk [24].
Semaglutide reduces the risk of non-fatal stroke, non-fatal myocardial infarction,
cardiovascular death with about 26% [24, 31]. Liraglutide and semaglutide decrease the risk of
developing kidney disease in patients with DM [24, 32]
Dipeptidyl peptidase-4 inhibitors (DPP-4) are oral antidiabetic agents that increase incretin
levels by inhibiting the decrease of the incretin hormones glucagon-like peptide-1 and glucose-
dependent insulinotropic polypeptide. Thus DPP-4 reduce glucagon production and increase
glucose-dependent insulin secretion. Apart from reducing glucose blood level, DPP-4 have
pleiotropic effects, such as improving endothelial dysfunction, reducing blood pressure and
inflammation [33]. This is why DDP-4 may have a protective effect against cardiovascular
disease, delay the progress of atherosclerosis and decrease the risk of PAD [33]. DPP-4 are
particularly useful in association with metformin [33, 34].
Thiazolidinediones have an important effect in preventing cardiovascular disease by
improving insulin sensitivity in peripheral tissues. Pioglitazone is a peroxisome proliferator-
activated receptor γ (PPAR γ) agonist prescribed in type 2 DM. PPAR γ are expressed by
115
macrophages and adipocytes. Pioglitazone may reduce atherosclerosis by improving insulin

resistance and decreasing systemic inflammation, which are involved in atherosclerotic plaque
formation [33, 35]. Pioglitazone reduces the risk of developing cardiovascular events –
myocardial infarction, stroke, compared to placebo in patients with clinically manifest vascular
disease [33]. The drug also decreases the risk of recurrent cardiovascular events in patients with
already established cardiovascular events [36, 37]. The disadvantage of this drug is that it
increases the risk of heart failure by 33% in susceptible patients with cardiovascular disease
[33]. Pioglitazone can lead to the development or worsening of heart failure.
Sulfonylurea antidiabetic drugs are commonly used in patients with type 2 DM, but seem to
have no advantage in patients with PAD and type 2 DM [24].
Conclusions
DM and PAD are commonly associated. Diabetes mellitus represents one of the risk factors
for cardiovascular disease. Peripheral artery disease is more frequent in patients with DM than
in the general population. The clinical manifestation of PAD in patients with DM are similar,
with the difference that patients with diabetes may be asymptomatic until advanced stages. This
is why PAD is diagnosed later and the prognosis may be worse. Oral antidiabetic drugs can
improve the prognosis of these patients by decreasing the risk of PAD in patients with diabetes.
The most effective agents seem to be sodium-glucose co-transportor 2 inhibitors and
glucagon-like peptide-1 receptor agonist which decrease the rate of cardiovascular events and
mortality. Concluding, good control of glucose blood levels in patients with DM can reduce
significantly the risk of developing PAD.
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117
Vascular Calcification in Diabetes Mellitus Type2 – Molecular

Mechanisms and Clinical Implications
CASOINIC Florin1, BUZOIANU Anca2, HANCU Nicolae3

1 Department of Internal Medicine, “Iuliu Hatieganu” University of Medicine and Pharmacy, Cluj-Napoca (ROMANIA)
2 Department of Pharmacology, Toxicology and Clinical Pharmacology, “Iuliu Hatieganu” University of Medicine and
Pharmacy, Cluj-Napoca (ROMANIA)
3 Department of Diabetology, “Iuliu Hatieganu” University of Medicine and Pharmacy, Cluj-Napoca (ROMANIA)
Emails: fcassoinic@yahoo.com, abuzoianu@umfcluj.ro, nhancu@umfcluj.ro
Abstract
In patients with diabetes mellitus type 2 (DMT2) prevalence of atherosclerotic vascular

disease and vascular calcification is increased along with higher incidence in and cardiovascular
disease. Although the mechanisms of diabetes associated progression of avascular calcification
are not fully understood, recent evidence suggests that vascular calcification is an active high
regulated process, were vascular smooth muscle cells (VSMCs) are the main players and are
able to express bone matrix proteins that and by transform these cells VSMCs osteoblast-like
cells, able facilitate or regulate the calcification process.
Intense research in this area of vascular calcification and several pathogenic pathways have
being describe along with identifying risk factors were high glucose and in oxidative stress are
important players. The increase prevalence of VC warrants further research in to the mechanism
by which diabetes induces this complication and design new effective therapeutic strategies.
Our review will focus on molecular mechanism and clinical implications of various type of
vascular calcification in patients with DMT2.
Keywords: vascular calcification, diabetes mellitus type 2, coronary artery disease, vascular smooth muscle cells,
intimal calcification, medial calcification
Introduction
Diabetes mellitus type 2 (DMT2) is a major health problem worldwide and is developing in
parallel with the sharp rise in obesity prevalence, sedentary and diabetogenic and pro-
atherogenic risk lifestyle. The prevalence of chronic complications and comorbidities
associated with diabetes is very high. Cardiovascular disease is the leading cause of mortality
in patients with diabetes, especially by coronary heart disease and stroke [1]. Diabetic patients
have twice as much risk of developing various vascular diseases (from coronary heart disease
to ischemic and haemorrhagic stroke and vascular deaths) compared with non-diabetic patients
with similar risk factors and independently from other conventional risk factors. In patients
without diabetes, fasting blood glucose concentration is modest and non-linearly associated
with risk of vascular disease [2]. In people with diabetes, vascular calcification (VC) can be
found in both coronary arteries and peripheral arteries and represent an independent predictor
of cardiovascular and overall mortalities [3, 4]. VC is a complex, active and highly regulated
biological process associated with the deposition of calcium crystals within the extracellular
matrix and various type of cells in media or intima of the arterial wall.
Molecular mechanisms leading to vascular calcifications are highly regulated by a variety of
molecular signalling pathways but this complex process is still not completely understood.
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There are two major types of vascular calcifications in patients with DMT2 described: VC
of the intima of the vessel wall is associated with atherosclerotic (ATZ) plaques and Medial VC
(Mönckeberg medial sclerosis) were calcium deposition occur in the arterial wall at the level of
the elastic lamina and extracellular matrix.
Intimal VC is systemic and is associated with ATZ plaques is associated by the classical
cardiovascular risk factors (age, sex, obesity, hypertension, smoking, dyslipidaemia, diabetes)
and may be triggered by oxidative stress or inflammatory pathways. ATZ arterial calcification
is the most common type of VC and can lead to proliferation of vascular smooth muscle cells
and determine the luminal narrowing. This type of VC can lead to increase in arterial stiffness,
systolic blood pressure and can lead to increase the vulnerability and ATZ plaque rupture. The
importance of calcification in ATZ plaque instability is influenced by the size and the location
of calcification [5]. Coronary artery calcium (CAC) evaluate the extend of the ATZ plaque
based on the calcium deposits is detected on cardiac CT imaging.
Medial VC (also called Mönckeberg medial sclerosis) is generally associated with diabetes
and chronic kidney diseases and present deposition of calcium hydroxyapatite along the elastic
lamina and the extracellular matrix. There are several risk factors for medial calcification:
hyperphosphatemia, reduction of glomerular filtration rate, hypercalcemia, parathyroid
hormone abnormalities, and duration of dialysis (for patients with end stage kidney disease)
[6]. Medial calcification can be present along with intimal VC, mostly in patients with DMT2
and chronic kidney disease, but may occur independently of atherosclerosis. Medial arterial
calcification represents an independent risk factor for future cardiovascular events and increase
cardiovascular mortality in patients. Clinical manifestation in patients with severe medial VC
are: arterial stiffness and increased cardiac post load and impaired vasodilation during ischemia
[7].
In a prospective study publish by Niskanen et al., in 133 patients new diagnosed with DMT2
17% present with medial arterial calcification and 23% of these patients present with intimal
calcification at baseline. During the 10 years of follow-up 21% of these patients died secondary
of cardiovascular disease and the odds ratios for cardiovascular mortality were 4.2 for patients
with median VC and 1.6 for patients present with intimal VC, after multiple logistic regression
analysis, medial VC was a stronger predictor of cardiovascular mortality compared with other
risk factors (including age, sex, hypertension, dyslipidaemia, smoking, hyperglycaemia, body
mass index, fasting serum insulin, urinary albumin, and ischemic ECG changes) [8].
In diabetes, VC is present both in the arteries of the lower extremities, where it is associated
with neuropathy and not always associated with coronary artery calcifications [9].
Our goal is to present several molecular mechanisms involved in vascular calcification
involving both intimal and medial arterial wall.
Risk Factors for Vascular Calcification in DMT2
All majors’ risks factors involved in development of CVD are also considered to be risk
factors for VC. For examples: age, dyslipidaemia, smoking, uncontrolled hypertension,
hyperglycaemia, obesity, stress, physical inactivity is associated with increase in VC, mostly
on intimal VC. In addition, with these classical risk factors, a genetic component is also,
important in several genetic disease VC was described at the very young age groups or since
birth, or multiple genetic variants were shown to be linked to aortic or coronary calcifications
[10, 11].
Although the role of lipids in VC pathology is not understood, dyslipidaemia can increase
osteogenic differentiation of vascular smooth muscle cells. Both acetylated and oxidized LDL
and triglycerides can increase calcification in vitro by calcifying vascular cells and HDL was
able to inhibit osteogenic differentiation pathways [12, 13]. Is unclear if presence of obesity is
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a risk factor for VC, but is a major risk factor for the development of insulin resistance,
inflammation and metabolic syndrome. In several reports increase in epicardial fat was found
to be associated with worsening in coronary calcification, in patients [14].
The role of hypertension in VC development remain to be elucidated, but renin-angiotensin-
aldosterone system seems to play an important role in pathogenesis of VC by increase VSMC
apoptosis, growth and differentiation and promoting osteogenic cell differentiation, also
treatment with angiotensin-converting enzyme inhibitors or mineralocorticoid antagonists can
reduce VC [15].
Inflammation present in patients with DMT2, but also in patients with metabolic syndrome
and insulin resistant state is associated with ATZ and increase in coronary artery calcifications.
Several pro-inflammatory cytokines including TNF-α (Timor necrosis factor-α) can induce
calcification of vascular cells in vivo [16].
Smoking can be also associated with increase in VC possible by increased oxidative stress,
by high levels of nitrogen oxide and other free radicals and impaired antioxidative mechanisms
[17].
High levels of oxidative stress can promote VC and ATZ by increase production of reactive
oxygen species, free radical nitric oxide and H2O2 by multiple cells including VSMC,
endothelial cells and macrophages and switch the VSMC to an osteogenic phenotype [18].
Hyperglycaemia and poor diabetic control are a well-known risk factors both for
microvascular and macrovascular complications in patients with DMT2. Coronary artery
calcification was found to be associated with high HbA1c levels and tight glycaemic control
can reduce cardiovascular events in patients with coronary ATZ in a patient with CAC score
<100, but not in patients with a high CAC score, suggesting that diabetic control before advance
ATZ develop can decrease intimal VC [19].
Presence of renal disease in patients with DMT2 increase the prevalence and severity both
for intimal and medial VC compared with patients with only renal disease, who already have a
higher risk in developing VC compared with general populations [20].
In Chronic Renal Insufficiency Cohort) Study were almost 2000 patients with impaired
kidney were included and almost half of whom have DMT2 (900 patients) a strong and linear
relationship was found between estimated glomerular filtration rate (eGFR) and levels of CAC
was found. This study proved that relationship between severity of CKD and CAC is
independent of traditional risk factors and VC should be considered as a complementary
evaluation in patients with DMT2 and chronic kidney disease [20].
Molecular Mechanism of VC
In patients with DMT2 secondary to chronic hyperglycaemia leads to accumulation of the

free radicals and increase in oxidative stress which are capable to modify multiple cellular
components including proteins, lipids and carbohydrates and activate multiple cellular
pathways including nuclear factor-κB-mediated vascular inflammation [21].
Advanced glycation end products (AGEs) are form by interaction between glucose,
galactose and fructose with various amino groups from protein structure and by interaction with
receptors for advanced glycation end production (RAGEs) can increase reactive oxygen species
and bone matrix proteins through multiple pro-calcific signalling pathways (PKC, p38 mitogen-
activated protein kinase MAPK, transforming growth factor-β TGF-β, nuclear factor-κB NFκB,
and Extracellular signal-regulated protein kinases 1 and 2 ERK1/2). [22]
AGE/RAGE signalling also increase oxidative stress to promote diabetes-mediated vascular
calcification through various mechanism and switch the VSMC to a osteoblast-like cells by
increasing Runt-related transcription factor 2 (RUNX2) he first transcription factor required for
determination of the osteoblast lineage, able to induces the differentiation of multipotent
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mesenchymal cells into immature osteoblasts [22]. More studies need to elucidate furthermore
the role of AGE/RAGE signalling pathways in diabetes-mediated vascular calcification in order
to develop new therapeutic target to decrease the severity of this diabetic complication.
The transforming growth factor-β(TGF-β) family members are essential in regulation of
angiogenesis and vasculogenesis, cell development embryogenesis and various genetic
mutations in this family often resulting in vascular pathologies such as atherosclerosis and
cardiovascular disease. TGF-b plays an important role in most cell types known to be present
in ATZ lesions: VSMC, platelets, endothelial cells, myofibroblasts, monocytes, macrophages
and can be released by these cells and leads to promotion of calcium accumulation in a matrix
that is prone for mineralization [23].
Bone morphogenetic proteins (BMPs) is a factor involved in osteoblastic differentiation
were the members of the TGF-b family described in literature as important players in the
induction of VC in vivo [24]. In 1993, Bostrom et al., described for the first-time expression of
BMP-2 in calcified human atherosclerotic plaques rising to the hypothesis that VC is a regulated
process like bone formation. Later on, BMP subfamily was found to have more than 20
members, with several members BMP 2 and 4 being important players in VC in diabetes both
in human and in animal models were hyperglycaemia can induce the level of this protein
expression in arterial wall [25].
Another important system in involved in VC and relationship with bone homeostasis
represent receptor activator of nuclear factor kappa-B ligand (RANKL)/receptor activator of
nuclear factor k B (RANK)/osteoprotegerin (OPG) system. OPG/RANK/RANKL is a cytokine
network involved in osteoclast differentiation and activation as a main regulator of the critical
balance between bone formation (osteoblasts) and bone resorption (osteoclasts).
RANKL-RANK interaction will initiate activation of the NF-kB pathway, essential in
osteoclast differentiation and activity. The OPG cytokine, a member of the tumour necrosis
factor receptor (TNFR-) superfamily, functioning as a decoy receptor to prevent
RANKL/RANK interaction in this way inhibits differentiation, activation, and osteoclast
survival. RANKL/RANK signalling increases bone turnover by enhancing resorption and bone
loss, whereas OPG promotes bone formation by blocking RANKL/RANK interaction [26].
High OPG proteins levels has been identified in ATS plaques in mouse models as well as
humans with was reported at low levels being protective for ATS plaque calcification but at the
high levels were observed in unstable plaques [27]. Clinical studies in different types of patients
have shown that high OPG serum levels are associated with vascular calcification, advanced
atherosclerosis, coronary artery disease, heart failure, and an increase in cardiovascular overall
mortality but other studies are showing a protective effect on calcification with high levels of
OPG [28, 29, 30, 31]. It seems that in early-phase ATZ, OPG release inhibit vascular
calcification but, latter on after ATZ became more mature, higher levels of OPG are having an
opposite effect and increasing VC by promoting inflammation and fibrosis.
Fetuin-A is a liver produced glycoprotein and represents an important inhibitor of VC by
stabilizing calciprotein particles (phosphate, calcium phosphate crystals) and inhibiting calcium
uptake by VSMCs and also by decreased inflammatory response after these calcium complexes
are ingested by local macrophages [32, 33]. Patients with end stage renal disease and low levels
of fetuin-A have a high risk of cardiovascular mortality. Multiples studies in various
populations reports an indirect correlation between fetuin-A serum levels and or arterial
stiffness, calcification score, and cardiovascular mortality.
Another important inhibitor of VC is Matrix Gla protein (MGP), a vitamin-K-dependent
activated protein expressed by multiple cells involved in vascular calcification including
VSMCs, chondrocytes, ECs, and fibroblasts. MGP acts by binding calcium ions excess and
inhibit crystal growth. MGP require vitamin K in order to become active (as a carboxylated
MGP) and protein expression is downregulated by vitamin D deficiency. VSMCs are capable
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of synthesizing active MGP, which inhibits the BMP-2 pathways and prevents VMSCs
differentiation toward an osteochondrogenic phenotype. [34] Vitamin K antagonists (warfarin)
use in patients on haemodialysis is associated with increase VC both in via an MGP-dependent
mechanism [35].
Vascular Calcification in DMT2 – Clinical Implications
DMT2 and metabolic syndrome represent important causes of VC and presence of renal
disease in diabetic patients increase the prevalence of both intimal and medial calcifications.
VC increase risk for developing of multiple comorbidities in patients with DMT2:
myocardial infarction, stroke, vascular dementia, renal insufficiency, peripheral vascular
disease, limb ischemia and amputation. CAC is considered to be one of the important
cardiovascular risk factors prediction factors in for coronary vascular disease and helps to
classify patients in either higher or lower risk categories based on the CAC score. CAC is
detected on cardiac CT imaging and it is the most common quantified with Agatston score,
introduced by doctor Agatston in 1990 [36]. In a study which followed more than 10.000
asymptomatic individuals over a 5 years period characterized by CAC, Shaw et al., and
compared classical risk factors (presence of hypercholesterolemia, hypertension, smoking and
diabetes mellitus or a family history of coronary artery disease) and found that CAC is an
independent predictor of mortality and was superior to a Framingham risk score [37]. CAC can
be used as a prognostic factor for an increase risk in mortality and morbidity secondary to
cardiovascular events and patients with low CAC score have a good prognostic over time [38].
Calcium scores outside of coronary artery – thoracic aorta calcification, abdominal aorta
calcifications or peripheral artery disease calcifications have been less well studied, but can be
used as markers for an increased mortality and morbidity. In diabetic patients, CAC score is
higher than general population, both in patients with CAD and in patients with subclinical ATZ
[39]. In a study published by Carson et al., higher HbA1c levels were associated with advanced
CAC progression, with an increase >100 Agatston units between examinations at the baseline,
independent by other risk factors, in patients with higher HbA1c levels, but without DMT2
[40].
In Multi-Ethnic Study of Atherosclerosis (MESA) almost 7000 patients were follow for
CAC progressions over 10 years period, and demonstrate a mean CAC score progression was
23.9 Agatston units/year in the general population, with a higher progression for patients with
DMT 2 of 31.3 Agatston units/year, demonstrating an increase levels of VC in patients with
diabetes[41]. In Heinz Nixdorf Recall Study, more than 3000 patients with uncontrolled DMT2
(HbA1c>7) were followed over five years period are rep greater CAC progression between the
baseline and the follow-up [42]. In Diabetes Prevention Program Outcome Study, more than
2000 patients with prediabetes were treated with metformin or placebo and followed over a 14
years period for men but not for woman. CAC severity and presence were significantly lower
in metformin versus the placebo group and was not influenced by demographic, anthropometric,
or metabolic parameters or statin treatment [43]. In the study by Anand et al., 510 patients with
uncomplicated DMT2 without known cardiovascular disease were evaluated for CAC and
myocardial perfusion scintigraphy if the patients Agatston score was more than 100 units. Over
two years period high CAC score and abnormal myocardial perfusion were significant predictor
factors for significant cardiovascular events [44]. Presence of VC indicate the increase risk for
further cardiovascular events but the extend of calcification is not relate d with the vulnerability
of ATZ plaque and is extremely important for this patients to be evaluated by additional tests
(angiography, myocardial perfusions scintigraphy, intravascular ultrasound) in order to
determine presence of stenosis and establish plaque vulnerability. CAC can be used in diabetic
patients for stratification in various risks and can be followed by various tests to better
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determine clinical course. Compared with intimal VC, there are fewer studies to evaluate the
relationship of aortic, thoracic or peripheral arterial calcification with mortality and morbidity
cardiovascular causes in patients with DMT2. Everhart et al., have showed that risk factors for
arterial calcification in patients with DM over a 20 years period in Pima Indians were
hyperglycaemia, duration of diabetes and neuropathy. Among diabetic patients, risk factors for
medial arterial calcification were impaired vibration perception, long duration of diabetes, and
high plasma glucose concentration. Over time these patients have a higher risk for lower leg
amputations, proteinuria, retinopathy, CAD, and mortality rate compare with patients without
medial VC [45]. Lehto et al., investigate the predictive value of medial VC in over 400 patients
with DMT2 compared with 600 without medial VC over a seven year period and conclude that
medial VC is a strong predictor of total CV events, including fatal or nonfatal myocardial
infarction, stroke, and amputation and CV mortality independent of duration of DM and
glycaemic control [46].
Medial VC is also known to be associated with Charcot neuroarthropathy (from 80-90% of
the patients and is also associated with diabetic neuropathy and foot ulceration in this population
[47, 48, 49].
The clinical significance of medial VC and how to follow the extension of medial VC in
patients with DMT2 is still not understood and cause of the increase mortality and morbidity
secondary to CV events remains speculative.
Conclusions
Despite substantial advance have been made in the molecular mechanism underlying
vascular calcification the significance of a VC in diabetes is not fully known and many
questions that remain without answer. It is clear that both intimal and medial VC are increased
in patients with DMT2, and the prevalence increase even more if these patients also present
renal impairment.
Multiple risk factors present on diabetic patient (increase in oxidative stress, increase in pro-
inflammatory cytokine productions, endothelial dysfunction) can increase the pro-calcific
media and prone to vascular calcification. Measurement of CAC in patients with DMT2 is a
predictor of mortality or morbidity mostly from cardiovascular causes and can be beneficial for
risk stratifications of patients. Using the CAC score in patients with less extensive calcification
seems to benefit from better glycaemic control, comparison with patients with more extensive
calcification were better control of diabetes doesn’t have a significant improvement in
progression of VC. More research is needed to improve our understanding in the VC
pathophysiology in order to better define the molecular mechanism and create therapeutic
potential in order to inhibit or attenuate progression of VC. By creating new pharmacological
strategies to target VC we can decrease cardiovascular risk in diabetic patients.
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prevention Program and its outcome study on coronary artery calcium. Circulation 2017; 136: pp. 52e64.
44. Anand DV, Lim E, Hopkins D, et al., Risk stratification in uncomplicated type 2 diabetes: prospective
evaluation of the combined use of coronary artery calcium imaging and selective myocardial perfusion
scintigraphy. Eur Heart J 2006; 27: pp. 713e21.
45. Everhart JE, Pettitt DJ, Knowler WC, Rose FA, Bennett PH (1988) Medial arterial calcification and its
association with mortality and complications of diabetes. Diabetologia 31: pp. 16-33 2.
46. Lehto S, Niskanen L, Suhonen M, Ronnemaa T, Laakso M (1996) Medial artery calcification. A neglected
harbinger of cardiovascular complications in non-insulin dependent diabetes mellitus. Arterioscler
Thromb Vasc Biol 16: pp. 978-983.
47. Sinha S, Munichoodappa CS, Kozak GP (1972) Neuroarthropathy (Charcot joints) in diabetes mellitus.
Medicine (Baltimore) 51: pp. 191-210.
48. Clouse ME, Gramm HF, Legg M, Flood T (1974) Diabetic osteoarthropathy. AJR 121: pp. 22-34.
49. Young MJ, Adams JE, Anderson GF, Boulton AJM, Cavanagh PR (1993) Medial arterial calcification in
the feet of diabetic patients and matched non diabetic control subjects. Diabetologia 36: pp. 15-621.
50. Forst T, Pfutzner A, Kann P, Lobmann RR, Schafer H, Beyer J (1995) Association between diabetic
autonomic C fibre neuropathy and medial wall calcification and the significance in the outcome of trophic
foot lesions. Experimental and Clinical Endocrinology and Diabetes 103: pp. 94-98.
125
Diabetes Mellitus: Unforgettable Became Inevitable – a Case

Report with a 2-Years Follow-Up
CHELARU Sorina-Alina1,2, CATRINOIU Doina1,2, NEAMŢU Mirela1,

ION Ileana2
1Unit of Diabetes Mellitus, Nutrition and Metabolic Diseases, Emergency Clinical County Hospital, Constanta (ROMANIA)
2Faculty of Medicine, “Ovidius” University, Constanta (ROMANIA)
Emails: cristeasorinaalina@yahoo.com, dcatrinoiu@gmail.com, neamtumirela82@yahoo.com, ileana_ion58@yahoo.com
Abstract
Diabetes Mellitus is not a single disorder. Regardless its type, it represents a cluster of
pathological modifications in glucose metabolism that leads to chronic hyperglycaemia.
Taking into account that many patients do not fit into a single category, there have been
recent calls to review the classification system.
Our case is one of the many examples that reinforce the need for new and updated
classification.
Keywords: diabetes mellitus, young patient, new classification
Introduction
Diabetes Mellitus (DM) is not a single disorder. It represents a cluster of pathological

modifications in glucose metabolism that leads to chronic hyperglycaemia, the biochemical
hallmark of micro and macrovascular complications.
The impressive dynamics of diabetes prevalence is part of the global burden of diabetes
mellitus with major impact on health expenditure related to the management of the disease and
its complications [1].
Diabetes has been known about since ancient times but World Health Organization (WHO)
published in 1965 its first classification system using age as diagnostic categories but
recognized also other forms [2]. After 15 years, a widely accepted and globally adopted
classification was published including two major types (insulin dependent diabetes mellitus –
IDDM or type 1 and non-insulin-dependent diabetes mellitus or type 2 – NIDDM), other types
and gestational diabetes. In 1999, they recommended that classification should include not only
different aetiological types but also clinical stages [3, 4].
Taking into account that many patients do not fit into a single category, there have been
recent calls to review and update the classification system.
In 2019, WHO published an update of the previous classification of DM as presented below
[2, 5].
126
Table 1. Types of diabetes [2]

Type 1 diabetes
Type 2 diabetes
Hybrid forms of diabetes:
Slowly evolving immune-mediated diabetes of adults
Ketosis prone type 2 diabetes
Other specific types:
Monogenic diabetes
- Monogenic defects of β-cell function
- Monogenic defects in insulin action
Diseases of the exocrine pancreas
Endocrine disorders
Drug- or chemical-induced infections
Uncommon specific forms of immune-mediated diabetes
Other genetic syndromes sometimes associated with diabetes
Unclassified diabetes:
This category should be used temporarily when there is not a clear
diagnostic category especially close to the time of diagnosis of
diabetes
Hyperglycaemia first detected during pregnancy:
Diabetes mellitus in pregnancy
Gestational diabetes mellitus
The revised classification does not include subtypes of type 1 and 2 but includes new types
as hybrid forms or unclassified ones.
One of the hybrid forms of diabetes include slowly evolving immune mediated diabetes of
adults who presents initially as type 2 diabetes mellitus (T1DM) but with evidence of pancreatic
antibodies. We usually refer to as LADA (latent autoimmune diabetes in adults) and this group
does not require insulin therapy at diagnosis, are initially controlled with lifestyle changes and
oral agents but with rapid progression to insulin treatment. There isn’t a universally agreed
criteria, but three of them are often used: presence of anti-glutamic acid decarboxylase antibody
(GADA), age more than 35 years old at diagnosis and no need of insulin for 6-12 months after
diagnosis [6].
Thus, a similar subtype has also been reported in younger patients (10-17 years old) and has
bed referred to as latent autoimmune diabetes in youth [7].
In 2018, a group of researchers from Sweden identified five replicable type of patients with
different characteristics and risk of diabetic complications: severe autoimmune diabetes, severe
insulin-deficient diabetes, severe insulin-resistant diabetes, mild obesity-related diabetes, mild
age-related diabetes.
Data regarding T1DM are unknown but it showed an annual increase of 3-4% of incidence
in childhood and between 70% and 90% of them have evidence of immune-mediated process
[8].
On the other hand, type 2 diabetes mellitus accounts 90% of diabetes patients. Thus, the
reality of 21st century is governed by obesity being the main health disorder and major risk for
type 2 diabetes [3]. It is responsible for 80% of the cases with diabetes, 35% with ischemic
cardiopathy and 55% with arterial hypertension [9].
All the increases lead to globalization phenomenon of diabetes with major impact on the
costs of disease management and associated complications.
Case Presentation
Two years ago, we have reported the case of a 19 years old boy who complained to his
general practitioner for polydipsia, polyuria, weight loss, symptoms that started 6 months before
and increased values of glycaemia proved by a random self-monitoring. The patient had a
positive family history for diabetes mellitus (2nd degree relatives with diabetes mellitus) and
127
not only (mother diagnosed with dermatomyositis, Raynaud syndrome and autoimmune
thyroiditis), normal birth weight but with a major pathological personal history. At the age of
13, he was diagnosed with a systemic form of idiopathic juvenile arthritis remitted after 2 years
of treatment with metotrexat. Initially, he received corticoid treatment for two months which
leaded to unbalanced glycaemic control with necessity of basal bolus insulin therapy until he
stopped taking this drug.
Taking into account the clinical findings and paraclinical investigations developed through
its admission in Diabetes Unit, we’ve come with the following aspects: normal clinical
evaluation, high variability of glycaemic values with severely increased HbA1c but negative
ketones, normal endogenous insulin secretion evaluated by measuring the level of c peptide but
positive marker of autoimmunity against beta cells. At discharge, we’ve recommended physical
activity, restriction of carbohydrates and treatment with sulphonylureas (SU). Shortly after
discharge, he came with good results by reaching the glycaemic targets so we concluded that
we were in front of an unclear diagnosis of Diabetes Mellitus regarding its type.
Now, we know that after 3 months, he presented to its diabetologist with hyperglycaemia
which decided to stop SU and initiate basal insulin therapy associated to dipeptidyl peptidase-
4 inhibitors (DPP4I).
And everything went quite well for 2 years until the first signs of major insulin deficiency.
The evaluation showed major unbalanced glycaemic control with lack of endogenous insulin
secretion and a new marker of autoimmune process.
Table 2. Paraclinical investigations

Glucose 272 mg/dl HDL 54 mg/dl
cholesterol
HbA1c 13.7% LDL 89 mg/dl
cholesterol
Serum creatinine 0.85 mg/dl Tryglycerides 33 mg/dl
Serum Urea 25 mg/dl Urine sample glucosuria
1000, negative
ketones
GADA 47 IE/ml Islet cells <1/10
Antibodies
Insulin Antibodies 5.48 U/ml Basal C 0.321ng/ml
peptide
Our goal was to establish the glycaemic control so we’ve initiated basal bolus insulin therapy
after stopping the oral drugs. After 3 months, the patient reached its glycaemic targets.
Discussions
Initially, the clinical aspects and biological evolution leads us to diagnosis of maturity onset
diabetes of the young (MODY) but further genetic evaluation were needed taking into account
the presence our autoimmune process. Thus, his family and personal history for immune
mediated diseases and the following evolution of the case lead us to a positive diagnosis of type
1 Diabetes Mellitus.
Conclusion
It seems that a new classification is needed taking into account that the phenotypes of T1DM
and T2DM are less distinctive with high proportion of incident cases of T1DM in adulthood
and the occurrence of T2DM in young people.
128
REFERENCES
1. Şerban V., Tratat Român De Boli Metabolice 1, Brumar, Timişoara, 2010.

2. Classification of diabetes mellitus, Geneva: World Health Organization, 2019.
3. Leslie RD, Palmer J, Schloot NC, Lernmark A. Diabetes at The Crossroads: Relevance of Disease
Classification to Pathophysiology and Treatment. Diabetologia. 2016; 59: pp. 13-20.
4. Karamanou M, Protogerou A, Tsoucalas G, Androutsos G, Poulakou-Rebelakou E. Milestones in the
history of diabetes mellitus: the main contributors. World J Diabetes. 2016; 7: pp. 1-7.
5. Diabetes mellitus. Report of a WHO Expert Committee. Geneva: World Health Organization, 1965.
6. Tuomi T, Groop L, Zimmet P, Rowley M, Mackay I. Antibodies to glutamic acid decarboxylase (GAD)
reveal latent autoimmune diabetes mellitus in adults with a non-insulin dependent onset of disease.
Diabetes. 1993; 42: pp. 359-362.
7. Klingensmith GJ, Pyle L, Arslanian S, et al., and the TODAY Study Group. The presence of GAD and
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8. https://www.thelancet.com/journals/landia/article/PIIS2213-8587(18)30051-2/fulltext?elsca1=tlpr;
9. Graur M., Obezitatea, Junimea, Iaşi, 2004.
129
Spectrum of Bacteria Associated with Diabetic Foot Syndrome

and Their Susceptibility to Antibiotics
ZAHA Dana Carmen1,2, POPA Amorin Remus1,2, JURCA Alexandru1,

POPA Loredana2, POPOVICIU Mihaela1,2, FERICIAN Anca2,
DAINA Cristian Marius1,2, MAGHIAR Octavian1, VESA Cosmin Mihai1,2
1Faculty of Medicine and Pharmacy, University of Oradea, Oradea (ROMANIA)
2Clinical County Emergency Hospital, Oradea (ROMANIA)
Email: danaczaha@gmail.com
Abstract
Lower limb amputations, appearing as a consequence of diabetic foot ulcers (DFU) and
diabetic foot infections (DFI), are common devastating complications of diabetes mellitus.
DFIs could be polymicrobial and multidrug-resistant (MDR) because of the ability of some
strains to form biofilm, which is an important virulence factor and results in treatment failure.
The main objective of the study is to investigate the microbial distribution and drug
susceptibility among diabetic foot infections.
In our study were enrolled 115 patients aged between 38-93 having chronic diabetic foot
infection and attending the Diabetes, Dermatology and Surgery department of Emergency
Clinical County Hospital Oradea for one year. Deep secretion was collected for microbial
culture and testing of drug susceptibility was done before antibiotic therapy and the results were
statistically analysed. There were processed 191 samples and 159 yielded positive cultures. The
159 positive samples included 141 cases (88.67%) of single-pathogen infections and 18 cases
(11.32%) of double-pathogen infections. Staphylococcus aureus was the predominant
organism, followed by Escherichia coli and Enterococcus faecalis. Even if the isolates had
relatively good antibiotic sensitivity rates, the pathogens and drug resistance in patients with
diabetic foot syndrome should be monitored to reduce the incidence of related complications
and improve the prognosis of patients.
Keywords: diabetes mellitus, foot ulcers, bacteria, antibiotics
Introduction
Diabetes prevalence has been rising more rapidly in many countries, the global prevalence
among adults over 18 years of age has risen from 4.7% in 1980 to 8.5% in 2014 [1]. There were
1.785.300 cases of diabetes in Romania in 2017, with a national prevalence of 12.4% [2].
Complications of diabetes (reduced blood flow, ischemia, and peripheral neuropathy) in the
feet range from simple to highly complex, including limb amputations and life-threatening
infections. Even though preventive strategies have been proper applied, diabetic foot ulcers still
occur frequently and are a challenge for the patient and physician as well. The global prevalence
of foot ulcers was 6.3%, higher in males (4.5% versus 3.5%) than in females, and higher in type
2 diabetic patients (6.4% versus 5.5%) than in type 1 diabetics [3].
Diabetics with infections commonly involved gram-positive organisms such as
Staphylococcus aureus, Enterococcus spp, and gram-negative organisms like Escherichia coli,
Pseudomonas aeruginosa, Klebsiella spp, Proteus spp, etc., and anaerobes, all these can display
multi-drug resistance. The microorganism that colonises the surface of the wound provides an
130
ideal opportunity for further invasion resulting infections. They also can exist independently or
combine together to form micro-communities within a matrix of extracellular polymeric
substances (biofilm), an important virulence factor establishing a protective environment for
the organisms to survive and resist to antibiotics. In the same time, biofilms are responsible of
chronic infections such as diabetic foot ulcers, emergence of multidrug-resistant strains and
treatment failure. The main objective of the study is to investigate the microbial distribution
and drug susceptibility among diabetic foot infections.
This retrospective study was conducted at the Clinical County Emergency Hospital of
Oradea in patients attending the Diabetes, Dermatology and Surgery departments for one year,
between November 2018-2019. One hundred fifteen patients were included in the study after
institutional ethical approval and informed consent obtained from the subjects. All patients
having chronic diabetic foot infections or/and ulcers were included in the study.
These patients had not received antibiotics before admission. Material for culture was
collected with a cotton-tipped sterile swab from the deeper parts of the foot infection/ulcer.
The swabs were transported immediately to the Laboratory of Microbiology for culture and
test for sensitivity to antibiotics. Samples were cultured on blood agar Columbia and Levin agar
and the plates were incubated overnight at 37 °C. Colonies obtained were identified by using
standard techniques and MALDI TOF system. Antibiotic sensitivity was performed using Kirby
Bauer’s disc diffusion technique according the Clinical Laboratory Standard Institute (CLSI)
guidelines 2018 and Vitek-2 [4]. Multidrug-resistant organisms for gram-positive and gram-
negative bacteria are resistant to three or more antimicrobial classes as are defined in the same
standard.
Results
A total of 115 adult diabetic patients who had diabetic follow-up (especially type 2) were
involved in our study. From the total number of patients involved in the study, 69 (60%) were
males and 46 (40%) were females. The patients were aged between 38 and 93 years, with an
average age of 64.8±0.82 years, the number of cases increasing with age. Also, the number of
cases is higher in patients with poor and very poor diabetes mellitus control as expected
otherwise (Table 1).
Table 1. Clinical and demographical data of the patients

Parameter n(%) or range
Gender
Male 69(60)
Female 46(40)
Age
<40 years 2(1.73)
40–50 years 11(9.56)
50–60 years 21(18.25)
60–70 years 33(28.69)
>70 years 48(41.73)
Diabetes
type 1 7(4.40)
type 2 152(95.59)
HbA1c (%)
≤7% (good control) 14(12.17)
7.1–8% (fair control) 24(20.86)
8.1–10% (poor control) 31(26.95)
>10% (very poor control) 46(40)
131
From these patients, 159 isolates were obtained including 89 (55.97%) gram-positive
organisms, 69 (43.39%) gram-negative bacilli, and a single fungal strain. There were 141
(88.67%) cases (56.8%) with single-pathogen infections and 18 cases (11.3%) with double-
pathogen infections.
Fig. 1. The distribution of pathogens
Staphylococcus aureus was the most commonly isolated pathogen in the ulcers followed by
Escherichia coli, Proteus spp, Enterococcus faecalis. Streptococcus spp, Pseudomonas
aeruginosa and Klebsiella spp were isolated in a small number, less than 10% for each (Fig. 1).
Fig. 2. The susceptibility of S. aureus Fig. 3. The susceptibility of E. coli
AMC-Amoxicillin/clavulanic acid, AZM-Azithromycin, CXM-Cefuroxime, GEN-Gentamicin

CIP-Ciprofloxacin, CLI-Clindamycin, ERY-Erythromycin, LNZ-Linezolid, MXF-Moxifloxacin,
OXA-Oxacillin, PEN-Penicillin, TEC-Teicoplanin, VAN-Vancomycin, AMK-Amikacin, FEP-
Cefepime, CTX-Cefotaxime, CAZ-Ceftazidime, CRO-Ceftriaxone, COL-Colistin, ETP-
Ertapenem, LVX-Levofloxacin MEM-Meropenem TZP-Piperacillin/Tazobactam
Staphylococcus aureus was the only pathogen that was consistently isolated (34.05%).
These strains showed good sensitivity rates to all antibiotics tested except for Penicillin,
Erythromycin, Oxacillin (Fig. 2), but unfortunately a quarter were methicillin resistant with
inducible resistance to Clindamycin.
Escherichia coli, the second etiological agent of foot infections (15.94%) showed good
sensitivity rates except to Ciprofloxacin and Ceftriaxone (Fig. 3). Isolated Proteus spp strains
represented 13.76% of all strains and the third etiological agent of foot syndrome. These strains
have also exhibited low resistance rates, but low sensitivity rates to Ceftazidime and
Ciprofloxacin explained by their frequent prescription (Fig. 4).
132
Fig. 4. The susceptibility of Proteus spp Fig. 4. The susceptibility of Enterococcus faecalis
There were identified among gram negative isolates seven extended-spectrum beta-
lactamases (11.11%), a single strain producing carbapenems and all were Escherichia coli.
Although a small number of multidrug resistant strains have been identified (only four, two
were Acinetobacter baumannii), they are significant because of their resistance to antibiotics
[5].
In our study, the fourth etiological agent of the foot diabetic infectious disease was
Enterococcus faecalis showing low level resistance to the tested antibiotics. These strains
exhibited relatively low rates of sensitivity to Penicillin, but no strain showed vancomycin
resistance (Fig. 4).
Discussion and Conclusion
The annual incidence of diabetic foot ulcers is estimated to be 1.0-4.1% and a common
complication of these ulcers is infection, which if left untreated or poorly treated leads to distal
limb amputation [6, 7]. Difficulties of treatment may be done by chronic bacterial infections
and/or associated with biofilms which are present in more than 80% [8].
In the present study, almost all the samples displayed monomicrobial isolates the most
frequent were found to be gram-positive. This is significantly different from other studies
results in which diabetic foot infections are polymicrobial and the predominance is of gram-
negative germs [9]. In this study, Staphylococcus aureus and Escherichia coli were the most
commonly isolated organisms followed by Proteus spp. The prevalence of methicillin-resistant
Staphylococcus aureus is higher probably due to hospitalization. The MRSA displayed a high
level of resistance to clindamycin, erythromycin, and penicillin. All of the organisms were
sensitive to linezolid, vancomycin, and quinolones similar to other studies [10].
The gram-negative organisms in our study showed resistance to ceftazidime, ciprofloxacin
and ceftriaxone. Contrary other studies in which one half (or more) strains are ESBL producers
[11, 12], in our study this percent is lower (11.11%) with the highest production by E. coli and
Piperacillin + tazobactam are the most effective drug against them.
A good glycaemic control is mandatory in diabetic patients. As expected, very poor control
of diabetes was associated with an increase rate of diabetic foot syndrome showing a high rate
of complications [13]. The inappropriate use of third-generation cephalosporins and heavy use
of some antibacterial drugs will be followed by a significant increase in drug-resistant
pathogens and difficulties in eradicating a chronic diabetic foot infection. These are the reasons
for monitoring the pathogens and drug resistance in patients with diabetic foot syndrome, in
order to improve the prognosis of patients. For this purpose, implementation of a quality
management in the hospital and adoption of effective measures implies the existence of
diagnostic and therapeutic protocols [14, 15].
133
REFERENCES
1. Sarwar N, Gao P, Seshasai SR, et al., (2010). Diabetes mellitus, fasting blood glucose concentration, and
risk of vascular disease: a collaborative meta-analysis of 102 prospective studies. Emerging Risk Factors
Collaboration. Lancet.; 26; 375: pp. 2215-2222.
2. https://www.idf.org/our-network/regions-members/europe/members/154-romania.html
3. Zhang P, Lu J, Jing Y, Tang S, Zhu D, Bi Y. (2017). Global epidemiology of diabetic foot ulceration: a
systematic review and meta-analysis, Ann Med. 49(2): pp. 106-116. doi:
10.1080/07853890.2016.1231932.
4. Clinical and Laboratory Standards Institute (CLSI) (2018). Performance Standards for Antimicrobial
Disk Susceptibility Tests
5. Zaha DC, Bungau S, Aleya S, et al., (2019). What antibiotics for what pathogens? The sensitivity
spectrum of isolated strains in an intensive care unit, Science of The Total Environment Volume 687, 15
October 2019, pp. 118-127.
6. Wu SC, Driver VR, Wrobel JS. et al., (2007). Foot ulcers in the diabetic patient, prevention and treatment.
Vasc Health Risk Manag. 3(1): pp. 65-76.
7. Popa AR, Vesa CM, Uivarosan D et al., (2019). Cross sectional study regarding the association between
sweetened beverages intake, fast-food products, body mass index, fasting blood glucose and blood
pressure in the young adults from North-western Romania, Revista de Chimie;70(1)
8. Ashok D. (2011). Why Diabetic Foot Ulcers do not heal? JIMSA; 24(4): p. 205.
9. Ling L, Zhihui L, Xinxin L, et al., (2018). Bacterial distribution, changes of drug susceptibility and
clinical characteristics in patients with diabetic foot infection, Experimental and Therapeutic Medicine,
Volume 16 Issue 4, pp. 3094-3098.
10. Mathangi T, Prabhakaran P. Prevalence of Bacteria Isolated from Type 2 Diabetic Foot Ulcers and the
Antibiotic Susceptibility Pattern Int. J. Curr. Microbiol. App. Sci. 2013; 2: p. 10.
11. Mamdouh ME, Ahmed Saif AI. (2012). Diabetic foot infection: Bacteriological causes and antimicrobial
therapy. Journal of American Science; 8(10).
12. Rani V, Nithya Lakshmi J. (2014). A comparative study of Diabetic and Non-diabetic wound infections
with special reference to MRSA and ESBL. Int. J. Curr. Microbiol. App. Sci. 3(12): pp. 546-554.
13. Popa AR, Judea Pusta CT, Vesa CM, et al., (2019). Prediction Models of Albumin Renal Excretion in
Type 2 Diabetes Mellitus Patients, REV.CHIM. (Bucharest), 70, No. 11.
14. Gabor-Harosa, FM, Stan, OP, Daina, L, Mocean F. (2016). Proposed model for a Romanian register of
chronic diseases in children. Computer methods and programs in biomedicine, 130, pp. 198-204.
15. Daina L, Sabău M, Daina CM, Neamțu C. (2019). Improving performance of a pharmacy in a Romanian
hospital through implementation of an internal management control system, Science of the Total
Environment, 675, pp. 51-61.
134
Cardiovascular Risk Assessment in a Group of Non-alcoholic

Fatty Liver Patients from Transylvania
DASCĂLU Daciana Nicoleta1,2

1Clinical Coounty Hospital, Sibiu (ROMANIA)
2“Lucian Blaga” University, Sibiu (ROMANIA)
Email: daci.nicoleta@gmail.com
Abstract
The clinical diagnosis of metabolic syndrome is not sufficient to assess the risk of
cardiovascular disease. In order to appropriate assessment and management of overall
cardiovascular risk in clinical practice, is important to take into account the traditional risk
factors and the additional contribution brought by obesity/insulin resistance and their related
complications. The overall risk resulting from this traditional risk factors along with intra-
abdominal obesity is known as global cardiometabolic risk.
The aim of the paper is to evaluate the cardiovascular risk of patients with non-alcoholic
fatty liver, and to assess the cardiovascular risk in all the 125 fatty liver patients and all subjects
in the control group using two forms of clinical practice assessment – Framingham score and
SCORE-HeartScore® formula.
Patients were also evaluated in terms of two new combinations of clinical factors and
laboratory predictors of cardiovascular risk: hypertriglyceridemia waist and hypertensive waist.
Keywords: Non-alcoholic fatty liver disease, cardiovascular risk, Framingham risk, SCORE risk
Introduction
Non-alcoholic fatty liver disease (NAFLD) falls into a spectrum of liver diseases
characterized mainly by macrovesicular fatty degeneration that occurs in the absence of
significant alcohol consumption (20-30 g pure alcohol per day or under 200g pure
alcohol/week) [1].
The spectrum of disease is composed of three clinical-pathological entities that are in fact
evolutionary stages of the disease:
1. Fatty liver disease: is characterized by predominant presence in hepatocytes of fatty
acids and triglycerides macrovezicules.
2. Steatohepatitis: fatty liver disease associated with a necro-inflammatory process,
Mallory bodies and incipient fibrosis.
3. Cirrhosis: characterized by architectural changes in liver fibrosis and inflammatory
infiltration associated steatosis [2, 3].
Both liver steatosis and steatohepatitis are associated with insulin resistance. Oxidative
status of a cell is determined by the balance between pro-and anti-oxidant processes. Thus, the
oxidative stress may occur when reactive oxygen species (ROS) are generated in hepatocytes
but also when there is a breakdown of antioxidants. This singularity of steatosis is also
associated with increased levels of 3-nitrotyrozin, a product of lipid peroxidation. This indicates
the presence of oxidative stress in non-alcoholic fatty liver hepatocytes. Fatty liver disease is
keeping up the appearance until the second “hits” which consists in appearance of one risk
factors like surgery on the intestine, drugs, development of type 2 diabetes. An important role
135
in this scenario appears to be owned by the sudden release of free fatty acids in circulation. So
non-alcoholic steatohepatitis occurs in patients already suffering from diabetes, central obesity,
weight loss, unexpected factors which mobilizes free fatty acids and precipitate their storage in
the liver [3].
Methodology
This is a prospective observational investigation on patients with ultrasound appearance of

hepatic steatosis without significant alcohol consumption (<200gr/week) and without chronic
hepatitis B or C infection. The paper tries to identify any features of this non-alcoholic fatty
liver patients in geographical area of Transylvania, and aimed to evaluate the cardiovascular
risk of patients with NAFLD, in order to evaluate the importance of fatty liver presence,
between other comorbidities, in the life prognosis of this patients. The data were analysed
versus a control group.
Study inclusion criteria:
- Group A: 125 patients with ultrasound appearance of fatty liver without serological
markers of HVB or HVC viral infection, without significant alcohol consumption (less
than 200 g pure alcohol/week), without clinical or biological aspect of hemochromatosis
or other liver pathology.
- Control group: 34 individuals with normal ultrasound appearance of liver without
serological markers of infection with hepatitis B or C, without significant alcohol
consumption (less than 200 g pure alcohol/week).
Exclusion criteria: Patients with ultrasound appearance of fatty liver with alcohol use over
200g/week or known with serological markers of infection with HVB/HVC, with the
clinical/biological aspect of hemochromatosis, autoimmune hepatitis or other liver pathology.
Results
Hypertriglyceridemia waist is defined as the simultaneous presence of abdominal

circumference above the normal range associated with serum triglycerides above 150 mg%.
Because metabolic syndrome increases the risk of type 2 diabetes and cardiovascular disease,
several organizations have proposed a screening approach to identify patients with metabolic
syndrome features. Based on the consideration that waist circumference and triglyceride levels
may be as important as other more demanding approaches, such as ATP III criteria,
hypertriglyceridemia waist may be the simplest tool available for rapid initial screening
assessment of the metabolic syndrome in clinical practice.
In patients of group A, the hypertriglyceridemia waist prevalence was 43.2% in men (n=16)
and 46.6% in women (n=41), with a total prevalence of 45.5% (n=57), unlike the control group,
where this condition was present in 2 women (5.8%) and in no man.
Hypertensive waist is defined as simultaneous presence of abdominal circumference above
the normal range associated with systemic hypertension (SBP>140mmHg or hypotensive
treatment). Many studies consider this being a method with a high sensitivity and specificity,
very useful in screening of metabolic syndrome due to ease of measuring the two para-meters.
Subsequently, in patient’s hypertensive waist there will be the need to determine the other
elements necessary to define the metabolic syndrome (blood glucose, triglycerides, HDL-
cholesterol) [4].
In patients from the lot A this condition was detected in a total of 48 (54.5%) women and 19
(51.3%) men, with an overall prevalence of 67 patients (53.6%).
136
Framingham cardiac risk score – The absolute risk of developing a cardiovascular disease
is defined as the probability of a clinical event (in the case of cardiovascular death) occurring
in a particular person in a period of time. In this case, the prediction interval is set at 10 years.
Framingham risk score is used today in clinical evaluation and consider a number of personal
factors – age, sex, cholesterol, smoking, hypertension, diabetes – indicating which is the risk of
developing a heart attack or dying of cardiovascular disease in the next 10 years.
A risk of 10% means that 10 of the 100 people who have those characteristics will develop
heart disease or will die of cardiovascular cause in the next 10 years. [5]
The evaluation of risk in patients aged 30-74 years in the 2 groups and has the next results:
a total of 113 patients in NAFLD lot were framed in terms of age with an average risk of
12.21239%, and statistically significative compared to the population of the same age and sex
(p=0.000515) (Fig. 1).
Fig. 1. Framingham risk of NAFLD patients in comparation with control lot
The risk in the control group was 3.473684%, calculated for the 19 subjects who fell under
the criteria of age.
The analysis found weak positive linear correlation between waist circumference (r=0.137),
respectively average blood pressure (r=0.238) and Framingham risk (Fig. 2), meaning that
patients with increased waist circumference and blood pressure have a high average 10 years
risk to present a major cardiovascular pathology.
SCORE cardiovascular risk – European Society of Cardiology has initiated the

development of a risk assessment system (SCORE) using data from 12 European cohort studies
137
(N=205.178) covering a wide geographic area at different levels of cardiovascular risk. To

calculate the cardiovascular risk according to SCORE system we used the formula HeartScore®,
which is a web program of risk prediction and management in order to assist clinicians in
optimizing individual cardiovascular risk reduction [6].
In patients of group A which were included in the age criteria for calculating cardiovascular
HeartScore® risk (n=101), we obtained an average of 3.03% statistically significant higher risk
than the people the same age and sex (1.88%) – p=0.001739. A total of 20 NAFLD patients in
group A showed an increased cardiovascular risk (≥5) quantified into the SCORE system,
mostly of them being men (n=13). Estimation of Framingham and SCORE cardiovascular risk
proved an increased risk with age (Spearman coefficient r=0.64, respectively r=0.47).
The risk was lower in female sex and higher in those presenting obesity, hypertensive waist
or metabolic syndrome.
Conclusions
This is a prospective observational investigation study of 125 patients with ultrasound

appearance of hepatic steatosis without significant consumption of alcohol or infection with
hepatitis B or C and aimed to evaluate the cardiovascular risk of these patients. The data were
analysed versus a control group of 34 subjects. The group of patients with fatty liver disease
has an increased prevalence of subjects with elevated waist/hip ratio above the normal range
(78 women with values >0.85 and 33 men with waist/hip >0.9).
A total of 65.6% fall within the definition of obesity (BMI ≥30kg/m2 presenting values),
most have a low level of physical activity and a pattern of inappropriately food rich in fat and
carbohydrates, with an unhealthy life style.
There were significant differences between the two groups studied in terms of clinical-
biological abnormalities characteristic metabolic pathology but also in terms of cardiovascular
risk level.
A significant proportion of the subject’s present cardiovascular disease (hypertension,
chronic heart disease)/severe dyslipidaemia and thus are classified as patients with high
cardiovascular risk.
These patients need a multidisciplinary approach (diabetes/cardiology/internal medicine
team) in order to establish the most proper therapeutic algorithm and mostly to decrease their
risk of death.
REFERENCES
1. Ramesh Seela, Arun J. Sanyal (2005). Evaluation and management of non-alcoholic steatohepatitis;
Journal of Hepatology 42: pp. S2-S12
2. Oproiu Carmen, Oproiu I, Alexandru C, Manuc M. (2003). Ficatul gras non-alcoholic. Date la zi; Revista
pentru educatie medicala continuă Gastroenterologie, 2003. 2: pp. 125-131.
3. Leuscher U. (2006) – Non-alcoholic Steatohepatitis (NASH) – Dr. Falk Pharma GmbH: 5th Edition.
4. Nita C., Rusu A, Bala C, Hâncu N. (2008). The Ability of Hypertensive Waist to Predict High
Cardiovascular Risk in General Population – Applied Medical Informatics 2008. 23 (3-4): pp. 37-42.
5. Bedogni G, Miglioli L, Masutti F, Tiribelli C, Marchesini G, Bellentani S. (2005). Prevalence of and risk
factors for non-alcoholic fatty liver disease: the Dionysos nutrition and liver study. Hepatology. Jul;
42(1): pp. 44-52.
6. Conroy R.M. et al., (2003). Estimation of ten-year risk of fatal cardiovascular disease in Europe: the
SCORE project. European Heart Journal 24: pp. 987-1003.
138
Evolution Particularities and Prognostic Significance of New-

Onset Atrial Fibrillation in a Group of Diabetic Female Patients
with Acute ST-Elevation Myocardial Infarction Treated by
Primary Percutaneous Transluminal Coronary Angioplasty
MACOVEI Liviu1,2, POPA Delia Melania1, TRINCA Cristina2,

ADOAMNEI Dumitru Marius2, NASTASA Dan George2, LOVIN Nicusor2,
BAZYANI Amin2, ACHITEI Ionut Silviu2, BURLACU Alexandru1,2,
NEDELCIUC Igor2
2“George I. M. Georgescu” Institute of Cardiovascular Diseases, Iasi (ROMANIA)
Email: deliamelaniapopa@gmail.com
Abstract
Atrial fibrillation (AF), an apparently benign supraventricular tachyarrhythmia, acquires a

special significance when it appears in the evolution of an acute myocardial infarction (AMI),
inducing significant hemodynamic changes, sometimes even life-threatening. When associated
with diabetes, the risk of mortality increases, being more important as it occurs earlier in the
evolution of an AMI. Our study included 106 diabetic female patients with acute ST-segment
elevation myocardial infarction (STEMI), who underwent primary percutaneous transluminal
coronary angioplasty (PTCA). The overall group was divided into two subgroups depending on
the presence or absence of AF: 53 patients with new-onset AF and 53 patients with sinus
rhythm. We concluded that patients with new-onset AF after the acute STEMI were older, with
higher systolic blood pressure and higher admission blood glucose levels when compared to
those without new-onset AF. They also presented an increased risk of potentially life-
threatening complications: acute pulmonary edema (p=0.037), cardiogenic shock (p=0.015) and
death (p=0.015).
Keywords: atrial fibrillation, diabetes mellitus, acute ST- elevation myocardial infarction, PTCA
Introduction
Patients with diabetes mellitus (DM) and acute myocardial infarction (AMI) are
heterogeneous individuals with different clinical status, being burdened with metabolic
disorders and numerous risk factors. Patients associating these two entities have worse
prognosis than subjects with AMI without DM, despite the introduction of drug-eluting stents.
Regardless of DM, hyperglycaemia is an independent risk factor for poor prognosis in
subjects with acute coronary syndromes (ACS) and the presence of hyperglycaemia on
admission in patients with DM and AMI is now undoubtedly associated with a significantly
higher mortality and morbidity [1, 2]. Patients presenting with acute ST-segment elevation
myocardial infarction (STEMI) have significantly higher levels of serum glucose on admission
than patients with non-ST-segment elevation myocardial infarction (NSTEMI) and, in addition,
a higher initial fasting glucose level was an independent predictor of new-onset AF after STEMI
[1, 3].
139
Atrial fibrillation (AF), the most commonly encountered clinical arrhythmia, can often
complicate acute coronary syndromes (ACS), particularly STEMI, with an incidence between
6 and 21% [4]. AF can be secondary to the complications of AMI, but it has a poor prognosis
when occurs independently, these patients being prone to an increased risk of all in-hospital
major cardiac complications and also to an increased risk of mortality [5]. Even if patients with
AMI could have a reduced mortality rate after receiving primary percutaneous transluminal
coronary angioplasty (PTCA), patients with new-onset AF after an acute STEMI have a higher
all cause-mortality after 12 months of follow-up [3].
It is also known that DM makes a significant contribution to the prevalence and incidence
of AF, independent of other established risk factors such as hypertension and congestive heart
failure. AF is 44% more prevalent and 38% more likely to develop when diabetes is present.
Besides, diabetes is not a significant predictor of AF among men but is highly significant
among women [6].
However, the independent contribution of diabetes in the occurrence of AF after STEMI is
not yet fully established. Therefore, the purpose of this work is to identify the particularities
and to quantify the prognostic significance of the new-onset AF in a group of diabetic female
patients with AMI revascularized by primary PTCA.
Methods
Our observational study comprised a total group of 106 diabetic female patients presenting
with acute ST-segment elevation myocardial infarction (STEMI), who underwent primary
percutaneous transluminal coronary angioplasty (PTCA). All of them were hospitalized in the
Coronary Intensive Care Unit of “Prof. Dr. George I.M. Georgescu” Institute of Cardiovascular
Diseases, Iasi and the study was conducted over a period of 5 years. In order to highlight and
quantify the prognostic significance of new-onset atrial fibrillation in diabetic patients after
STEMI, the overall group was divided into two subgroups depending on the presence of either
atrial fibrillation (AF) or sinus rhythm (SR): the AF group (53 diabetic patients with new-onset
atrial fibrillation) and the SR group (53 diabetic patients without atrial fibrillation).
The patients from both groups were analysed comparatively, throughout the entire
hospitalization, from the moment of admission to the time of discharge or death. For each of
them, the following were taken into account: clinical aspects, the presence of comorbidities
(arterial hypertension, dyslipidaemia, smoking) as well as prior medical history of type 1 or
type 2 diabetes, with the analysis of admission blood glucose levels, glycaemic control markers
and the presence or absence of proteinuria. The diagnosis and the location of the acute
myocardial infarction (AMI) were established by electrocardiography (ECG). AF was also
detected by the 12-lead ECGs recorded during the entire hospitalization and by the 24-hour
continuous ECG monitoring in the Coronary Intensive Care Unit. The existence of AF prior to
admission was considered an exclusion criterion. Each patient was also evaluated by
transthoracic echocardiography in order to determine the following: ejection fraction, left atrial
dimensions, left ventricular end-systolic and end-diastolic diameters, tricuspid annular plane
systolic excursion (TAPSE), RV-RA pressure gradient, the presence or absence of ischemic
mitral regurgitation and the presence of left ventricular aneurysm. It also allowed the evaluation
of the pericardium. Every patient included in this study underwent coronary angiography in
order to detect and quantify the lesions on the coronary arteries and also to perform the primary
angioplasty. Finally, all the complications that occurred during the hospitalization (acute
pulmonary edema, cardiogenic shock, early post-infarction pericarditis, thromboembolism,
mechanical complications and death) were analysed and summarized.
140
Statistical Analyses
Statistical analyses were performed using SPSS 18.0. The centralization of the data was done
nominally and numerically. We conducted bivariate comparisons of baseline covariates among
patients with and without atrial fibrillation using t tests for continuous variables and χ2 test tests
for categorical variables. Intergroup statistical data were analysed through the nominal variables
using Fisher’s exact test. A p value of <0.05 was considered statistically significant.
Results
The age of diabetic patients with acute myocardial infarction treated by primary PTCA
complicated with newly-onset AF (71.13±7.80 years) was significantly increased compared to
patients with sinus rhythm (SR) whose mean age was 67.09±9.02 years (p=0.015). All patients
included in the study had type 2 diabetes. The admission blood glucose level was significantly
increased in patients with AF compared to those with SR (252.38 vs 221.42 mg/dl; p=0.031).
The presence of proteinuria was observed in 41.2% of the patients in the FA group and in
28.3% of the patients in the SR group, but the percentage distributions were not statistically
significant (Chi-square=1.89; df=1; p=0.170).
Regarding the risk factors, we found out that 24.5% of the patients from AF group and 15.1%
of the patients from the SR group were smokers (Chi-square= 0.95; df =1; p=0.330).
Being an important cardiovascular risk factor, especially in diabetic persons, the presence of
dyslipidaemia in both study groups was also evaluated. In the AF group dyslipidaemia was
present in 75.5% of the total number of patients, and in the SR group in 77.4% of the patients,
(Chi-square= 0.52; df=1; p=0.820). Arterial hypertension was present in 69.8% of patients with
AF and 64.2% of patients with RS, without statistical significance (p=0.680). Regarding the
admission values, the systolic blood pressure (SBP) was significantly lower in the SR group
compared to patients in the AF group (127.74±33.46 mmHg vs 145.60±26.69 mmHg, p=0.003).
Diastolic blood pressure (DBP) at admission recorded a slightly lower value in the SR group
compared to the AF group (79.15±13.11 mmHg vs 72.92±19.52 mmHg, p=0.057).
In the group of patients who developed AF, 38.7% of them had heart failure (18.9% with
NYHA class II, 17% with NYHA class III and 1.9% with NYHA class IV). In patients
remaining in SR, 25.5% had heart failure (15.1% NYHA class II, 3.8% NYHA class III and
5.6% NYHA class IV). NYHA class III heart failure was found to be significantly more
frequent in the group of patients with AF (p=0.026).The echocardiographic evaluation showed
minor differences between the groups for: ejection fraction (36.66±12.31% vs 39.60±10.61%),
TAPSE (18.66±5.31mm vs 19.1±4.13mm) and RV-RA pressure gradient (29.26±13.90 vs
25.59±10.68). On the other hand, the dimensions of the left atrium showed significant
differences between the two categories of patients (p=0.013).
The primary PTCA localization, that is, the main coronary artery on which stent angioplasty
was performed, was also evaluated. The difference obtained between the two groups of patients
is illustrated in Fig. 1. Thus, in patients with AMI and AF, in 5.7% of cases the angioplasty was
performed on the circumflex coronary artery (Cx), in 34.0% on the right coronary artery (RCA),
while in most of the cases (60.3%) on the left anterior descending artery (LAD) or its branches.
In the group with SR, 13.2% of the PTCA were made on Cx, 37.7% on RCA and 49.1% on
LAD.
141
Fig. 1. The primary PTCA localization
Fig. 2 displays the most important complications encountered in both patient groups. In
patients with AF, the relative risk of ischemic mitral regurgitation is 1.57 times higher than in
patients with sinus rhythm (p=0.151). The highest relative risk in the group of patients with
AMI and PTCA complicated by AF is the risk of acute pulmonary edema (p=0.037). Patients
with new-onset AF also have a relatively increased risk of developing cardiogenic shock
(p=0.015) and thromboembolism (p=0.151). The left ventricular aneurysm (13.2% vs 11.3%)
and pericarditis (18.9% vs 15.1%) are found in the picture of complications in both study groups
in similar proportions (p>0.05).
Complications
25
20 22
15
10 14 13 13
10 10
5 7 8 8
6 1 1 0 4
3 3
0
AF group SR group
Fig. 2. Complications
Within the group of patients with AF, there were a number of 13 cases (24.5%) that evolved
to death, following cardiogenic shock and/or acute pulmonary edema, whereas in the group of
patients with SR only 3 cases (5.7%) progressed to death. This shows that mortality is
significantly higher in patients with AMI complicated by AF than in patients with SR (p=0.015).
Discussions
The present study, based on a group of 106 diabetic female patients hospitalized for STEMI,
underlines the particularly important role played by new-onset AF in a whole constellation of
142
complications encountered in this category of patients. The findings are consistent with various
previous studies. The association between AF and DM has been already proven both in
epidemiology and experimental studies [7] and available evidence indicates that it significantly
increases morbidity and mortality [8]. Moreover, the association of these two entities acquires
particular aspects in the context of an acute coronary syndrome, such as STEMI. Kundu et al.,
have shown that patients who develop AF after STEMI are older, more likely to be women and
have a history of previous cardio-vascular diseases. In addition, they are more prone to have a
hospital course characterized by the development of several life-threatening complications such
as stroke, heart failure, acute renal failure, and cardiogenic shock [9, 10]. Vukmirović et al.,
suggested that the strongest predictors of AF development during the hospital period were older
age, enlarged diameter of LA as well as the presence of moderate to severe mitral regurgitation
(MR) and increased BMI [11].
In our study population, we concluded that the age of patients developing AF after STEMI
was significantly higher than the age of the patients who remained free from AF during their
hospitalization (p=0.015). All patients had type 2 diabetes. Analysing the biomarkers of
glycaemic control, they revealed poorly controlled diabetes in 50.9% of the patients who
developed AF and in 45.3% of the patients without this complication (p=0.562). Besides,
significant data were obtained regarding the admission blood glucose levels (p=0.031), attesting
once again that hyperglycaemia on admission in patients with STEMI is associated with adverse
outcome, as other previous studies have suggested [1, 12, 13]. Ekmekci et al., have shown that
the rate of in-hospital mortality, major adverse cardiovascular events (MACE), congestive heart
failure (CHF), cardio-pulmonary resuscitation (CPR), inotropic usage, and ventricular
tachycardia/ventricular fibrillation (VT/VF) were significantly higher in patients with
hyperglycaemia at admission. In addition, death, re-infarction and MACE are also more prone
to appear in the long-term follow-up [14]. In our study, echocardiographic evaluation at
admission confirmed the increased risk of developing AF in patients with enlarged LA
(p=0.013), while the other parameters did not appear to have an independent prognostic role.
We particularly analysed the evolution of the patients after the primary PTCA was
performed. Numerous previous findings have drawn attention to the multiple complications that
occur in patients with new-onset AF [9, 15, 16]. Patients with AMI complicated by AF have
also a significantly higher risk of dying during their hospitalization [9]. In contrast, ventricular
arrhythmia, ventricular aneurysm, cardiac rupture and mural thrombosis after AMI seldom
occur in patients without atrial fibrillation receiving PTCA [17]. In this context, we concluded
that patients with AF present a particularly higher risk of developing acute pulmonary edema
(p=0.037) and cardiogenic shock (p=0.015), among other possible complications
(thromboembolism, ischemic mitral regurgitation, left ventricle aneurysm).
Conclusions
Today, the negative impact of diabetes on the evolution of acute myocardial infarction is
well established. In addition, the post-STEMI prognosis appears to be less favourable in
diabetic women, even after primary PTCA. The present study concluded that the association of
new-onset AF in diabetic women with STEMI treated with primary PTCA significantly
increases the risk of potentially life-threatening complications, among which the most
significant are acute pulmonary edema, cardiogenic shock and death.
REFERENCES
1. Dąbek, J., Bałys, M., Majewski, M., & Gąsior, Z. (2016). Diabetic Patients with an Acute Myocardial
Infarction in Terms of Risk Factors and Comorbidities Management: Characteristics of the Highest-Risk
143
Individuals. Advances in Clinical and Experimental Medicine, 25(4), pp. 655-663. doi:
10.17219/acem/58785
2. Cosentino, F., Grant, P., Aboyans, V., Bailey, C., Ceriello, A., & Delgado, V. et al., (2019). 2019 ESC
Guidelines on diabetes, pre-diabetes, and cardiovascular diseases developed in collaboration with the
EASD. European Heart Journal, 41(2), pp. 255-323. doi: 10.1093/eurheartj/ehz486
3. Hsu, H.P., Jou, Y.L., Wu, T.C., Chen, Y.H., Huang, S.S., Lin, Y.J., & Chen, S.A. (2012) Hyperglycaemia
increases new-onset atrial fibrillation in patients with acute ST-elevation myocardial infarction. Acta
Cardiologica Sinica, 28(4), pp. 279-285.
4. Schmitt, J., Duray, G., Gersh, B., & Hohnloser, S. (2008). Atrial fibrillation in acute myocardial
infarction: a systematic review of the incidence, clinical features and prognostic implications. European
Heart Journal, 30(9), pp. 1038-1045. doi: 10.1093/eurheartj/ehn579
5. Iqbal, Z., Mengal, M. N., Badini, A., & Karim, M. (2019). New-onset Atrial Fibrillation in Patients
Presenting with Acute Myocardial Infarction. Cureus. doi: 10.7759/cureus.4483
6. Nichols, G.A., Reinier, K., & Chugh, S.S. (2009). Independent Contribution of Diabetes to Increased
Prevalence and Incidence of Atrial Fibrillation. Diabetes Care, 32(10), pp. 1851-1856.
7. Sun, Y., & Hu, D. (2010). The link between diabetes and atrial fibrillation: cause or correlation? Journal
of cardiovascular disease research, 1(1), pp. 10-11. doi:10.4103/0975-3583.59978
8. De Sensi, F., De Potter, T., Cresti, A., Severi, S., & Breithardt, G. (2015). Atrial fibrillation in patients
with diabetes: molecular mechanisms and therapeutic perspectives. Cardiovascular diagnosis and therapy,
5(5), pp. 364-373. doi: 10.3978/j.issn.2223-3652.2015.06.03
9. Kundu, Amartya & O’Day, Kevin & Shaikh, Amir & Lessard, Darleen & Saczynski, Jane & Yarzebski,
Jorge & Darling, Chad & Thabet, Ramses & Akhter, Mohammed & Floyd, Kevin & Goldberg, Robert &
Mcmanus, David. (2016). Relation of Atrial Fibrillation in Acute Myocardial Infarction to In-Hospital
Complications and Early Hospital Readmission. The American Journal of Cardiology. 117.
10.1016/j.amjcard.2016.01.012
10. Ibanez, B., James, S., Agewall, S., Antunes, M., Bucciarelli-Ducci, C., &Bueno, H. et al., (2017). 2017
ESC Guidelines for the management of acute myocardial infarction in patients presenting with ST-
segment elevation. European Heart Journal, 39(2), pp. 119-177. doi: 10.1093/eurheartj/ehx393
11. Vukmirović, M., Bošković, A., Tomašević Vukmirović, I., Vujadinovic, R., Fatić, N., Bukumirić, Z., &
Vukmirović, F. (2017). Predictions and Outcomes of Atrial Fibrillation in the Patients with Acute
Myocardial Infarction. Open medicine (Warsaw, Poland), 12, pp. 115-124. doi:10.1515/med-2017-0018.
12. Lonborg, J., Vejlstrup, N., Kelbaek, H., Nepper-Christensen, L., Jorgensen, E., Helqvist, S., Engstrom,
T. (2014). Impact of Acute Hyperglycaemia on Myocardial Infarct Size, Area at Risk, and Salvage in
Patients with STEMI and the Association with Exenatide Treatment: Results from a Randomized Study.
Diabetes, 63(7), pp. 2474-2485. doi: 10.2337/db13-1849
13. Sanjuán, R., Núñez, J., Blasco, M.L., Miñana, G., Martínez-Maicas, H., Carbonell, N., Sanchis, J. (2011).
Prognostic Implications of Stress Hyperglycaemia in Acute ST Elevation Myocardial Infarction.
Prospective Observational Study. Revista Española De Cardiología (English Edition), 64(3), pp. 201-
207. doi: 10.1016/j.rec.2010.08.005
14. Ekmekci, A., Uluganyan, M., Tufan, F., Uyarel, H., Karaca, G., Kul, S., Eren, M. (2013). Impact of
admission blood glucose levels on prognosis of elderly patients with ST elevation myocardial infarction
treated by primary percutaneous coronary intervention. Journal of geriatric cardiology: JGC, 10(4), pp.
310-316. doi: 10.3969/j.issn.1671-5411.2013.04.002
15. Reinstadler, S.J., Stiermaier, T., Eitel, C., Fuernau, G., Saad, M., Pöss, J., Eitel, I. (2018). Impact of Atrial
Fibrillation During ST-Segment–Elevation Myocardial Infarction on Infarct Characteristics and
Prognosis. Circulation: Cardiovascular Imaging, 11(2). doi: 10.1161/circimaging.117.006955
16. Nambiar, D.S.D. (2017). Cardiac and Extra Cardiac Predictors and Complications of Acute Atrial
Fibrillation Complicating ST Elevation Myocardial Infarction (STEMI) ST Elevation myocardial
infarction Acute Atrial Fibrillation (STAAF) Study. Journal of Medical Science and Clinical Research,
05(05), pp. 22124-22134. doi: 10.18535/jmscr/v5i5.139
17. Zhang, Y., Zhang, L., Zheng, H., & Chen, H. (2018). Effects of atrial fibrillation on complications and
prognosis of patients receiving emergency PCI after acute myocardial infarction. Experimental and
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144
The Importance of Physical Exercise for the Correction of Spinal

Deviations in Children with Type 1 Diabetes
AMARICAI Elena1
1
Department of Rehabilitation, Physical Medicine and Rheumatology, “Louis Turcanu” Emergency Children’s
Hospital, “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
Email: amaricai.elena@umft.ro
Abstract
Thoracic kyphosis and scoliosis represent spinal deviation that affects a high number of
children and adolescents. The objectives of our study were to assess the respiratory function
and functional capacity in children with type 1 diabetes and spinal deviations that followed a
physical therapy programme. The study included 11 children and adolescents with type 1
diabetes and spinal deviations (thoracic kyphosis or/and idiopathic scoliosis, thoracic or
thoraco-lumbar type). The study patients performed a twelve-week supervised exercise
programme in the outpatient Rehabilitation Department. The patients were assessed at the
beginning and at the end of rehabilitation by spirometry and functional capacity testing (6-
minute walk test). In children and adolescents with type 1 diabetes and spinal deviations
respiratory parameters and functional capacity improved after a twelve-week supervised
exercise programme. Still, when compared to healthy controls functional capacity had still
lower values.
Keywords: Type 1 diabetes, kyphosis, scoliosis, physical therapy, functional capacity
1. Introduction
Thoracic kyphosis and scoliosis represent spinal deviation that affects a high number of
children and adolescents. A pronounced thoracic kyphosis and moderate or severe types of
scoliosis have an impact not only on the development of paediatric population, but can also
have consequences on the ventilatory parameters and on the aerobic functional capacity.
A recent study (2019) showed that children with type 1 diabetes present a low bone-turnover
state with reduced bone mineralization and poorer bone microarchitecture [1].
Physical activity and exercise should be encouraged in people with type 1 diabetes.
Regular exercise can decrease risk factors for cardiovascular diseases, offering protection
against all-cause mortality [2] and improvement of the quality of life [3]. The objectives of our
study were to assess the respiratory function and functional capacity in children with type 1
diabetes and spinal deviations that followed a physical therapy programme.
2. Methodology
The study included 11 children and adolescents (aged between 6 and 14 years) with type 1
diabetes and spinal deviations. 5 patients had thoracic kyphosis; 3 had idiopathic scoliosis
(thoracic or thoraco-lumbar type). 3 patients had thoracic kyphosis and thoraco-lumbar
scoliosis. 11 gender and age-matched healthy controls were recruited as controls.
The following demographic characteristics related to patients and controls were collected:
age, gender, weight and height. The study patients performed a twelve-week supervised
145
exercise programme in the outpatient Rehabilitation Department. The aims of the rehabilitation
were: posture improvement, strengthening of the back, stretching the chest muscles, enhancing
exercise capacity and increasing chest expansion. The physical exercise programme was
performed gradually, according to the patients’ exercise capacity.
The patients were assessed at the beginning and at the end of rehabilitation by spirometry
(forced vital capacity-FVC, forced expiratory volume in 1 second-FEV1 and peak expiratory
flow-PEF) and functional capacity testing (6-minute walk test: 6MWT). The 6MWT was
conducted according to the standardized protocol [4]. The subject was instructed to walk up
and down a measured corridor, covering as much ground as possible over a 6-minute period.
3. Statistical Analysis
The statistical analysis was performed using the GraphPad Prism 5.0 for Windows.
Descriptive statistics were calculated for patients and controls characteristics, respiratory
parameters and the 6-minute walk distance (mean and standard deviation).
4. Results
The study and control groups were homogenous in terms of their demographic features
(Table 1).
Table 1. Patients’ and controls’ baseline characteristics
Variables Patients group Control group p
Age (years)
- mean ± SD 9.33±2.2 9.33±2.2 NS
Height (cm)
- mean ± SD 138.5±11.61 139.7±7.21 NS
Weight (kg)
- mean ± SD 37.6±8.25 39.2±9.8 NS
Gender
- Male- N (%) 6 (55%) 6 (55%) NS
- Female- N (%) 5 (45%) 5 (45%)
SD: standard deviation; N: number of patients; NS: Not significant
In patients with type 1 diabetes with spinal deviations FVC, FEV1, PEF and 6-MWT
improved significantly after twelve-week physical exercise programme. After rehabilitation
there were no differences between study patients and controls, except for functional capacity
that had lower values in children and adolescents with type 1 diabetes and spinal deviations
(thoracic kyphosis and/or scoliosis) (Table 2).
Table 2. Pulmonary function and functional capacity of study group and control
Patients initial Patients final p Controls p
evaluation evaluation Patient initial evaluation Patient final
(mean±SD) (mean±SD) vs final (mean±SD) vs Control
FVC (L) 2.16±0.8 2.57±1.37 0.005 2.75±1.33 0.07
PEF (L/min) 4.12±1.1 4.34±1.24 ˂0.0001 4.46±1.06 0.09
FEV1 (L/s) 2.06±0.67 2.34±0.54 0.008 2.47±0.9 0.3
6MWT (m) 298.3±55.2 326.2±29.2 0.008 352.7±50.1 0.024

SD: standard deviation; FVC: forced vital capacity; PEF: peak expiratory flow;
FEV1: forced expiratory volume in 1 second; 6MWT: 6-minute walk test
146
5. Discussions
Pulmonary function and overall functional capacity are extremely important parameters for
a harmonious children’s motor development. Children suffering from type 1 diabetes must have
an adequate functional status taking into account their chronic disease. Besides the respiratory
system, cardiac function should be considered when the children have moderate or severe spinal
deformations [5, 6].
In our study, the children and adolescents with type 1 diabetes had also spinal deviations.
The thoracic kyphosis was low or moderate degree, while the scoliosis was mild type.
Although mild or moderate type of vertebral deformities, these associated disorders must be
detected as early as possible and adequately treated. The impact of spinal deformities on the
development of children in their growth period should be considered when we are referring to
the quality of life [7, 8]. Physical activity is a key factor in the prevention of kyphosis and
scoliosis [9].
6. Conclusions
In children and adolescents with type 1 diabetes and spinal deviations respiratory parameters
and functional capacity improved after a twelve-week supervised exercise programme. Still,
when compared to healthy controls functional capacity had still lower values. This fact implies
the necessity of follow-up of the physical exercise in order to increase the overall functional
status.
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147
Type 2 Diabetes Mellitus and the Risk of Hepatocellular

Carcinoma in Chronic Hepatitis C Patients Treated with Direct
Acting Antivirals
ILIESCU Elena Laura1,2, TOMA Letitia1,2, DIACONU Camelia2,3,

ZGURA Anca4, BACALBASA Nicolae2, MERCAN-STANCIU Adriana1
1 Department of Internal Medicine II, Fundeni Clinical Institute, Bucharest (ROMANIA)
3 Department of Internal Medicine, Clinical Emergency Hospital, Bucharest (ROMANIA)
4 Department of Chemotherapy, OncoFort Hospital, Bucharest (ROMANIA)
Emails: laura_ate@yahoo.com, letitia_toma@yahoo.com, drcameliadiaconu@gmail.com, medicanca@gmail.com,

nicolaebacalbasa@gmail.com, adriana.mercan@yahoo.ro
Abstract
The effect of diabetes mellitus on the development of hepatocellular carcinoma in chronic

hepatitis C virus infected patients is a highly-discussed topic that still remains inconclusive. In
this paper we present the epidemiological data linking diabetes mellitus, hepatocellular
carcinoma and hepatitis C infection, while reporting a cohort study that aims to evaluate the
effect of diabetes mellitus on the development of hepatocellular carcinoma, in chronic hepatitis
C patients treated with direct acting antivirals. Patients were monitored by liver function tests,
blood cell count, coagulation profile and tumoral markers at the beginning of therapy, EOT and
SVR. Imagistic evaluation included Fibroscan® and abdominal ultrasonography and, in selected
cases, computerized tomography/magnetic resonance imaging. HCC prevalence was higher in
diabetic patients than in those with normal glucose metabolism, with an even higher prevalence
in individuals requiring insulin-therapy. Almost half of the HCC cases were discovered during
antiviral treatment, but medication was not stopped, resulting in all patients achieving SVR. In
conclusion, patients diagnosed with DM are at risk of developing HCC, while some oral
antidiabetic drugs are associated with a lower incidence of HCC. The findings in our
retrospective cohort study are concordant with the medical data.
Keywords: diabetes mellitus, hepatocellular carcinoma, hepatitis C infection, direct acting antivirals
1. Introduction
Hepatitis C virus (HCV) is highly prevalent worldwide, affecting approximately 71 million

people and leading to severe health problems, such as chronic hepatitis, cirrhosis, liver failure
and hepatocellular carcinoma [1]. Moreover, in 40-70% of the cases, HCV can induce
extrahepatic manifestations, including metabolic disorders [2]. Growing evidence shows a
direct relationship between HCV and type II diabetes mellitus (DM), suggesting that DM is the
most common extrahepatic implication of HCV [3, 4]. Currently with a global prevalence of
approximately 9%, DM is estimated to affect about 300-400 million individuals worldwide by
2030 [5, 6]. Hepatocellular carcinoma (HCC) is considered the fifth most frequent type of
cancer globally and the second most common cause of cancer-related death [7, 8]. HCV
infection is a well-known cause of HCC (with a 15- to 20- fold increased risk for HCC
development) [9], but current research also suggests that DM and IR are independent risk
factors for HCC development [10, 11]. The use of direct acting antivirals (DAAs), meant to
interfere with specific steps in the HCV replication, currently allows viral eradication and might
148
interfere with HCC prevalence [12]. Antiviral therapy also has an impact on the association
HCV-DM. A particular entity, new-onset diabetes after transplantation is mainly described
during the first 6 months after the transplant, due to the use of immunosuppressive agents [13,
14]. This study aims to assess the effect of diabetes mellitus on the development of
hepatocellular carcinoma in chronic hepatitis C virus infected patients, that underwent treatment
with direct acting antivirals.
We conducted a retrospective cohort study on a number of 797 patients, all of them infected
with HCV genotype 1b, who received treatment with ritonavir-boosted paritaprevir/ombitasvir
and dasabuvir (OBV/PTV/r + DSV) and with ledipasvir/sofosbuvir respectively. The subjects
were admitted to our clinic between December 2015 and July 2019.
Antiviral therapy was given to each eligible patient, according to the National Healthcare
Program. The local Ethical Committee approved the study. An informed written consent was
taken from all the participants and all their records were confidential. The study only included
subjects with active HCV genotype 1b infection, determined by HCV-RNA (real-time PCR
(TaqMan), that described a high viral load in all patients (more than 800 000 IU/L). The subjects
presented various degrees of fibrosis, from F0 to F4, estimated by Fibromax® and Fibroscan®;
no subject underwent liver biopsy for liver evaluation, due to its invasive character and
susceptibility of associated complications. Moreover, patients presenting with the following
conditions were excluded from the study: HBV or HIV co-infection, personal history of HCC,
type 1 DM, DM due to corticoid therapy. Both antiviral regimens were administered according
to the protocol: OBV/PTV/r 12,5mg/75mg/50mg 2 pills once a day and DSV 250mg twice a
day, with a total duration of treatment of 12 weeks; Ledipasvir/sofosbuvir 90mg/40 mg fixed-
dose combination tablet once daily; in cirrhotic patients, therapy duration was 24 weeks, while
non-cirrhotic subjects underwent antiviral treatment for 12 weeks. Virologic response at the
end of treatment (EOT) and at 12 weeks after the end of treatment-sustained virologic response
(SVR) were defined as undetectable HCV-RNA. Patients were monitored by liver function tests
(aspartate aminotransferase AST, alanine aminotransferase ALT, total bilirubin TB, gamma-
glutamyl-transpeptidase GGT, alkaline phosphatase, serum albumin), assessment of renal
function (serum creatinine, urea, proteinuria), blood cell count, coagulation profile and alpha-
fetoprotein (AFP) and the beginning of therapy, EOT and SVR. Standard imagistic evaluation
included Fibroscan® and abdominal ultrasonography. In patients with high AFP levels, as well
as in those presenting ultrasonographic liver abnormalities, further imagistic examinations
(such as computerized tomography or magnetic resonance imaging) were performed, in order
to assess the presence/absence of hepatocellular carcinoma. Fasting glucose blood levels were
determined in all subjects. In patients with established diabetes mellitus and in those with
hyperglycaemia, glycosylated haemoglobin levels were checked at the beginning of therapy,
EOT and SVR. Diabetic patients were classified as: type 2 DM controlled with diet, type 2 DM
controlled with oral antidiabetic drugs (OAD) and insulin-requiring type 2DM. Before initiating
antiviral therapy, oral antidiabetic medication, insulin dosages and all other concomitant
medication suffered adjustments in order to avoid potential drug interactions with the
Interferon-free treatment. Statistical analysis was executed with the SPSS 18.0 statistical
software (SPSS Inc., Chicago, IL, USA). Mean +/- standard deviation were used to express
numerical values, while ANOVA test was performed for in-between groups comparison (with
statistically significant p-values of less than 0.05).
149
3. Results and Discussion
We retrospectively collected and analysed data from 797 patients infected with HCV
genotype 1b. Baseline characteristics are presented in Table 1.
Table 1. Baseline characteristics of the study group

Parameter OBV/PTV/r + DSV Ledipasvir + Sofosbuvir
N=574 pts (72%) N=223 (28%)
No DM DM (diet) DM DM No DM DM (diet) DM (OAD) DM
N=420 pts N=18 pts (OAD) (insulin) N=121 pts N=6 pts N=61 pts (insulin)
(73.17%) (3.13%) N=81 pts N=55 pts (54.26%) (2.7%) (27.35%) N=35 pts
(14.11%) (9.58%) (15.7%)
Mean Age 47.28+/- 50.82+/- 51.28+/- 53.17+/- 48.21+/- 50.77+/- 53.28+/ 55.22+/-
17.52 18.45 22.84 23.45 19.33 17.65 20.55 21.45
Gender-male 202 pts 8 pts 38 pts 39 pts 65 pts 2 pts 38 pts 19 pts
(48.09%) (44.44%) (46.91%) (70.90%) (53.71%) (33.33%) (62.29%) (54.28 %)
ALT (U/ml) 58+/-31 52+/-28 63+/-34 59+/-27 65+/-35 73+/-33 69+/-29 77+/-34
AST (U/ml) 42+/-17 45+/-19 48+/-21 50+/-17 55+/-27 59+/-22 61+/-33 69+/-35
Albumin (g/dl) 4.2+/-1.5 4.3+/-0.7 4.2+/-0.9 4.0+/-0.8 4.1+/-1.1 3.9+/-0.9 3.5+/-0.9 3.2+/-1.1
TB (mg/dl) 1.0+/-0.4 1.1+/-0.3 1.2+/-0.5 1.3+/-0.7 1.2+/-0.4 1.3+/-0.2 1.5+/-0.8 1.6+/-0.7
INR 1.07+/-0.1 1.03 +/-0.3 1.09+/-0.1 1.08+/-0.2 1.21+/-0.1 1.20+/-0.09 1.3+/-0.1 1.4+/-0.12
AFP (ng/ml) 10.6+/-5.3 12.7+/-8.2 14.7+/-6.3 14.3+/-9.4 14.2+/-9.1 15.7+/-6.9 17.7+/-8.3 17.9+/-6.3
Fasting glucose 84+/-18 119+/-17 129+/-27 165+/-39 85+/-19 122+/-18 132+/-29 161+/-29
(mg/dl)
HbA1c (%) 5.2+/-0.4 6.0+/-0.5 6.5+/-0.8 7.9+/-2.1 5.3+/-0.3 6.1+/-0.6 6.6+/-0.9 8.1+/-2.1
F0-F1 fibrosis 19 pts 6 pts 3 pts 1 pt 103 pts 0 pts 15 pts 12 pts
N=29 pts (4.52%) (33.33%) (3.70%) (1.81%) (85.12%) (24.59%) (34.28%)
(5.05%)
F2 fibrosis 55 pts 5 pts 21 pts 10 pts
N=91 pts (13.09%) (27.77%) (25.92%) (18.18%)
(15.85%)
F3 fibrosis 71 pts 4 pts 33 pts 14 pts
N=122 pts (16.90%) (22.22%) (40.74%) (25.45%)
(21.25%)
F4 fibrosis 275 pts 3 pts 24 pts 30 pts 8 pts (6.61 4 pts 14 pts 10 pts
N=332 pts (65.47%) (16.66%) (29.62%) (54.54%) %) (66.66%) (22.95%) (28.57%)
(57.83%) 10 pts 2 pts 32 pts 13 pts
(8.26 %) (33.33%) (52.45%) (37.14%)
HCC 4 pts 2 pts 5 pts 18 pts 0 1 pt 3 pts 16 pts
N=29 pts (0.95%) (11.11%) (6.17%) (32.72%) (16.66%) (4.91%) (45.71%)
(5.05%)
Within the study group, data revealed that diabetic patients were mostly males and slightly
older than those without DM (p=0.04, p=0.05 respectively, CI 95%). Noticeably, diabetic
patients had higher levels of transaminases than those with normal glucose metabolism (p=0.01,
CI 95%). One potential explication may be the association of DM and IR with non-alcoholic
steatohepatitis, which can lead to increased liver enzymes, as well as increased Fibromax and
Fibroscan scores. In our cohort, assessment of fibrosis degree resulted in a predominance of F4
(over 53% of the total population). Most of them had compensated cirrhosis (86.58%). Diabetic
patients represented almost a third of the whole study group (32.12%) and more than a half of
the diabetic patients (51.56%) had advanced fibrosis (F4).
These findings are persistent with data found in several epidemiologic studies, that suggest
an association between diabetes mellitus/insulin resistance and a more rapid progression
towards fibrosis in patients with chronic liver disease [14, 15]. Moreover, in a multivariate
analysis conducted by Ong et al., [16], advanced fibrosis was significantly more likely in
patients with DM. The relationship between diabetes mellitus and advanced hepatic
fibrosis/cirrhosis is bidirectional. On one hand, impaired liver function (caused by cirrhosis) is
responsible for the main metabolic changes (altered insulin sensitivity, hyperinsulinemia and
pancreatic β-cell dysfunction), leading to the so-called “hepatogenous diabetes” [3, 17-18].
150
On the other hand, diabetes is associated with an increase in mitochondrial oxidative stress,
thus promoting fibrosis and cirrhosis [19]. In the literature, estimated prevalence of DM among
cirrhotic subjects is largely variable (from approximately 20% to 70%) [18]. Among cirrhotic
patients included in our study, DM prevalence is 31.05%. 49 patients (6.14%) developed HCC,
with a significantly higher prevalence in diabetic patients (17.56%), compared to those without
diabetes (Table 2).
Table 2. HCC prevalence within the study group and subgroups

797 pts
No DM Diabetic pts P-value DM (diet) P-value DM P-value DM P-value
N=541 N=256 N=24 (OAD) (insulin)
(67.87%) (32.12%) (3.01 %) N=142 N=90
(17.81%) (11.29%)
HCC 4 (0.73%) 45 (17.56%) P<0.001 3(12.5%) P=0.05 8 pts P=0.09 34 P=0.01
N=49 (5.63%) (37.77%)
(6.14%)
The association between DM and HCC has been described, over time, in various cohort and
case-control studies [12, 20-22]. A meta-analysis published in 2006, conducted by El-Serag et
al., [23] observed that DM is associated with an approximately 2.5-fold increased risk of HCC.
Later, Tanaka et al., found a positive association between DM and HCC risk [24]. The exact
mechanisms that are responsible for the connection between these two entities remain poorly
understood, but there are several factors considered to be involved: endogenous/exogenous
hyperinsulinemia, hyperglycaemia, chronic inflammation [6]. A number of 21 patients
developed HCC while on antiviral treatment, all of them having previously been classified as
cirrhotic (F4). Abdominal CT scan was performed, showing single HCC nodules in 14 subjects
and multiple HCC nodules in the other 7. Five patients presented signs of portal vein thrombosis
(PVT). The direct antiviral treatment was not interrupted in any of the cases. The subjects with
HCC and no signs of PVT underwent trans-arterial chemoembolization (TACE) with
doxorubicin, while on antiviral therapy, with good outcome. At the EOT (week 12), 12 more
F4 subjects were diagnosed with single HCC nodules, after performing computerized
tomography, all of which also underwent TACE. The SVR evaluation revealed another 16 cased
of HCC, that also had an indication for performing TACE. Moreover, our study showed that
HCC prevalence was significantly higher in patients with insulin-requiring diabetes (37.77%)
and diet-controlled DM than in those undergoing treatment with OAD (Table 2). This fact
correlates with previous in vitro and in vivo studies, implying that OAD may have an influence
on the development of multiple types of cancers, including hepatocellular carcinoma.
Metformin (a drug that was highly prevalent among our OAD-treated DM patients) and
thiazolidinediones (TZDs) are associated with a reduced incidence of neoplasms [6, 25], while
insulin and insulin secretagogues appear to correlate with a higher cancer incidence [6, 26].
Prolonged therapy with metformin seems to inhibit hepatic cells transformation, diminishing
the risk of HCC, similarly to nondiabetic individuals, although the molecular process remains
unclear [6, 27, 28]. In the era of direct acting antivirals for chronic HCV infection, recent trials
suggest possible better outcomes for diabetic patients after achievement of SVR. A large cohort
study including over 24.000 patients, showed that HCV eradication with DAAs is associated
with better glycaemic control in diabetic patients, demonstrated by lower HbA1c levels and
reduction of insulin doses [29].
Antiviral treatment options in these patients included sofosbuvir/ledipasvir, OBV/PTV/r +
DSV and sofosbuvir/simeprevir, the first two being also used within the cohort we described,
with very good results (all patients included in our study achieved SVR).
151
4. Conclusions
Diabetes mellitus is a global health problem and patients diagnosed with DM are at risk of
developing numerous pathogenic conditions, including HCC. The connection between these
two entities has been a constant subject of investigation for decades and nowadays DM is
considered to be a risk factor for HCC. Metformin and thiazolidinediones are associated with a
lower incidence of HCC. SVR achievement through DAAs is an important step in obtaining a
better glycaemic control in patients with DM. The findings within our retrospective cohort study
align with the data found so far in the medical literature. However, this topic remains
controversial and further research is required, in order to validate the nature of this association.
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Correlated or incidental? A brief review. World J Hepatol 10(9): pp. 595-602.
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Gastroenterol 22(27): pp. 6100-6113.
11. De Lope CR et al., (2012) Management of HCC. J Hepatol 56 (Suppl 1): S
12. Aghemo A et al., (2017) Sustained virologic response prevents the development of insulin resistance in
patients with chronic hepatitis C. Hepatology 56(5): p. 1681, 7.10.1002/hep.25867
13. Iliescu L, Mercan-Stanciu A et al., (2018) A Severe Case of Hyperglycaemia in a Kidney Transplant
Recipient Undergoing Interferon-Free Therapy for Chronic Hepatitis C. Acta Endo (Buc) 2018 14: pp.
533-538.
14. Kita Y, Mizukoshi E, Takamura T et al., (2007) Impact of diabetes mellitus on prognosis of patients
infected with hepatitis C virus. Metabolism – Clinical and Experimental, 56(12), pp. 1682-1688.
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of patients with genotype 1 HCV infection, American Journal of Gastroenterology 103(5), pp. 1136-1144.
16. Ong JP et al., (2001) Chronic hepatitis C and superimposed non-alcoholic fatty liver disease. Journal of
Liver, 21(4), pp. 266-271.
17. Garcia-Compean D et al., (2009) Liver cirrhosis and diabetes: risk factors, pathophysiology, clinical
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153
Novel Therapeutics Aspects in Advanced and Metastatic Non-

small Cell Lung Cancer – Experience of Oncology Department of
Chronic Disease St. Luke’s Hospital Bucharest
RAHNEA-NITA Gabriela1,2, CIUHU Anda-Natalia1,

RAHNEA-NITA Roxana-Andreea1, STOIAN Alexandru-Rares2,3
1 Department of Oncology – Palliative Care, “Sf. Luca” Chronic Disease Hospital, Bucharest (ROMANIA)
3 Department of Surgery, “Bagdasar Arseni” Emergency Hospital, Bucharest (ROMANIA)
* All authors had equal scientific contribution and share first authorship.
Emails: gabriela_rahnea@yahoo.com, andadum@yahoo.com, roxana_rahnea@yahoo.com, dr.raresstoian@yahoo.com
Abstract
Introduction
Bronchopulmonary cancer remains the leading cause of cancer death all over the world.
The molecular analysis and the identification of different genetic mutations/alterations of
tumour cell, discovering new targeted therapies, changed the treatment algorithm, especially
for patients with NSCLC. As well as, lung cancer is characterized by a strongly
immunosuppressive microenvironment, leading to the introduction in the landscape of
treatment management of NSCLC of immune checkpoint inhibitors.

In 2018-2019 periods, 558 patients with lung cancer were admitted to Department of Oncology
from Chronic Disease “St. Luke” Hospital, Bucharest. Only 276 patients were admitted for
specific oncological treatment, most of them with NSCLC. Therapeutic decisions for patients
with NSCLC was based on molecular analysis of the biological samples and, in some cases,
required the administration of novel therapies, according to the reimbursement and the specific
protocols in Romania.
Results and Discussions

Only 8,33 % patients (23/276) with advanced/metastatic lung cancer were eligible for
targeted therapy or immunotherapy. Patients in treatment with targeted therapies had clinical
benefit, partial remission or stationary disease, but with the development of treatment
resistance, mainly after 5-6 months. Step by step, we implemented immunotherapy to NSCLC
patients, and the identification and management of immunological adverse reactions remains a
challenge.
Conclusions
The discovery of EGFR mutations in lung cancer tumour cells and, subsequently, of a
specific treatment – TKIs – revolutionized the treatment of NSCLC patients. EGFR TKIs and
ALK TKIs have shown significant benefit in the treatment of a subset of patients with lung
cancer. Immunotherapy tends to become the standard in the treatment of patients with
bronchopulmonary cancer, either in monotherapy or in combination with conventional
chemotherapy.
Keywords: lung cancer, targeted therapy, immunotherapy
154
1. Introduction
Lung cancer remains the leading cause for death from cancer in Romania. Frequently, the
patients are diagnosed in advanced stages of the disease, when it is already inoperable or
metastatic. Therapeutic strategies in advanced or metastatic stages of lung cancer are as follows:
systemic treatment, radiation therapy for local control of tumour or palliative treatment (local
control of metastatic disease or control of symptomatology), best supportive care, nutrition,
surgery with limitative role [1, 2]. In 2011, studies specified a median survival for advanced
clinical stages of NSCLC (non-small cell lung cancer) IIIA and IIIB of 14 months and 10
months, respectively, with a survival rate up to 5 years of 19% and 7%, respectively.(1) In
clinical stage IV of NSCLC, the median survival was of 6 months, with a survival rate up to 5
years of 2%. (1) At that time, the systemic treatment of patients with NSCLC was the
conventional chemotherapy: Gemcitabine, Vinorelbine, Etoposide, Paclitaxel/Docetaxel,
Cisplatin/Carboplatin. The molecular mutations of proto-oncogenes, such as fi k-ras, EGFR,
erb B2 being into the study to ascertain the clinical relevance [3].
Afterwards, the treatment of NSCLC was revolutionized by “targeted therapies”: tyrosine
kinase inhibitors (TKIs) – erlotinib, gefitinib with action in intracellular tyrosine kinase domain
of epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF)
directed antibody – Bevacizumab [4-7].
Recently, the advent of novel immunotherapy targeting immune checkpoint has
revolutionized the treatment in advanced and metastatic NSCLC, based on the fact that lung
cancer is characterized by a strongly immunosuppressive microenvironment [8, 9]. Currently
used immunotherapy agents in NSCLC are: Pembrolizumab, Nivolumab, Atezolizumab,
Durvalumab [10-12]. Therefore, the treatment algorithm of patients with NSCLC includes
molecular diagnosis with the identification of gene mutations/alterations, at the moment being
examined the following EGFR (epidermal growth factor receptor) gene mutations, ALK
(anaplastic lymphomas kinase) gene rearrangements, ROS1 (ROS proto-oncogene 1) gene
rearrangements, BRAF (B-RAF proto-oncogene) point mutations, KRAS (KRAS proto-
oncogene) point mutations, NTRK (neurotrophin tyrosine receptor kinase) gene fusions, PD-
L1 (programmed death legend 1) (Table 1).
Table 1.
Mutation/alteration gene Targeted therapy/immunotherapy FDA approval
EGFR mutation positive Osimertinib Apr 2018
Afatinib 2013
Erlotinib Nov 2004
Gefitinib Jul 2015
Dacomitinib Sept 2018
EGFR mutation positive – Osimertinib Nov 2015
subsequent therapy p. T790M
mutation positive
ALK rearrangement positive Alectinib Dec 2015
Brigatinib Apr 2017
Ceritinib Apr 2014
Crizotinib Aug 2011
Lorlatinib Nov 2018
ROS 1 Rearrangement positive Ceritinib Mar 2016
Crizotinib Mar 2016
Entrectinib Aug 2019
BRAF V600 mutation positive Dabrafenib/trametinib Apr 2015
NTRK gene fusion positive Larotrectinib Nov 2018
Entrectinib Aug 2019
PD-L1 >1% Pembrolizumab Oct 2015
Atezolizumab Oct 2016
Nivolumab Mar 2015
155
In the period 2018-2019 a number of 558 patients with lung cancer were admitted to the
Department of Oncology of Chronic Disease St. Luke’s Hospital Bucharest. The retrospective
assessment of this patients was intended to implement a personalized treatment algorithm,
taking into account the therapeutic progress from the last decade, mainly for the non-small cell
lung cancer, as well as refunding the targeted therapy and immunotherapy within specific
protocols, from the last 3 years.
Having a long experience of over 10 years in the treatment with Erlotinib and Bevacizumab
for patients with advanced/metastatic NSCLC, according to the specific protocols in force
(without the evaluation of the genetic biomarkers for prescription), there was a challenge to us,
in the last 2 years, to initiate the management of the treatment and adverse reactions of targeted
therapies recently refunded in Romania (afatinib, crizotinib, osimertinib) and immunotherapy
(nivolumab, pembrolizumab), with addressability to patients with NSCLC.[13]Thus, patients
with advanced/metastatic NSCLC have been evaluated as follows: histopathological and
immunohistochemical diagnostic, tumours genomic profile – limited to EGFR, ALK, PD-1,
p.T790M mutations/alterations evaluation, performance status, associated comorbidities which
contraindicate immunotherapy/targeted therapy, adverse reactions of innovative treatments.
In the period 2018-2019, in the Oncology Department of Chronic Diseases “St. Luke”
Hospital, Bucharest, there were hospitalized 558 patients diagnosed with lung cancer – 254
patients in 2018 and 304 patients in 2019. 276 patients of them (49,46%) followed a specific
oncological treatment – chemotherapy, targeted therapy or immunotherapy; that is 51,18% of
the lung cancer patients hospitalized in 2018 and 48,02% of the patients who were hospitalized
in 2019. All the patients undergoing a specific oncologic treatment have histopathological and
immunohistochemical confirmation, 23,18% (64/276) patients diagnosed with SCLC and
76,81% (212/276) patients with NSCLC. We focused our study on the patients with NSCLC
admitted on the Oncology Department who benefited from targeted therapy and
immunotherapy, according to the specific protocols in Romania.
22,16% patients (47/212) from the patients with NSCLC had adjuvant chemotherapy after
thoracic surgery for stages I-IIIA and 77,83% patients (165/212) underwent treatment for
advanced/metastatic cancer disease. The histopathological and immunohistochemical analysis
of the biological specimens of this 212 patients with NSCLC pointed out the following: 0,47%
patients (1/212) had large-cell neuroendocrine carcinoma, 0,47% patients (1/212)
hadsarcomatoid cell carcinoma of the lung, 72,64% patients(154/212) sere diagnosed with
adenocarcinomas (ADKs) and 26,41% patients(56/212) were diagnosed with squamous cell
lung carcinomas (SCCs) (Fig. 1).
156
Fig. 1. Immunobiological sampling of patients with lung cancer from Oncology Department
180
154
160
140
120
100
80 64
56
60
40
20 0
0
SCLC NSCLC
SCLC 64
sarcomatoid carcinomas 0 1
squamous carcinomas 0 56
adenocarcinomas 154
neuroendocrin large cell
1
carcinomas
The patients with lung ad carcinomas and squamous cell lung carcinomas were asked to have
evaluated the molecular markers – EGFR, ALK, PD-L1 – which can be analysed in Romania
for free, at the moment. 89.04% patients (187/210) received a molecular diagnosis where there
were noticed the following mutations/alterations: EGFR mutations – 5,34% patients (10/187),
ALK gene rearrangements – 0,95% patients (2/210), PD-L1 – 2,13% patients (4/210) (Fig. 2).
Fig. 2. Molecular sampling of patients with NSCLC

200
180 172
160
140
120
100
80
60
40 23
20 10
3
0
EGFR mutation positive 10
ALK rearrangements 2
PD-L1 positive >50% 3
EGFR - /ALK- /PD-L1 <50% 172
Not evaluated 23
Somatic mutation in EGFR gene is detected in 10-15% of Caucasian patients with NSCLC
and are associated with responsiveness to EGFR tyrosine kinase inhibitor (TKIs) therapy [5,
157
14, 15]. All evaluated patients with NSCLC, EGFR mutation positive, have adenocarcinomas,
70% patients (7/10) were non-smokers female, 30% patients (3/10) were light-smokers male.
All of them presented deletion in exon 19, only 70% patients (8/10) were eligible for
treatment with TKIs in first line for metastatic disease (the rest of them – 2 patients followed
adjuvant post-surgery chemotherapy and 1 patient did not fit the protocol because of a bad
performance status) [16].
Patients undergoing Erlotinib in first line treatment showed disease progression after 8, 10
and 24 months, respectively, of targeted therapy One of the patients was lost from the record
after 2 months of treatment. 3 patients who followed treatment with Afatinib had disease
progression after 8 months, and 2 patients, after 10 months of treatment. For patients with
mutations of EGFR gene, TKIs in first line is the treatment of choice, because studies have
identified in advanced/metastatic NSCLC a response rate of 67% for Erlotinib and Gefitinib,
with an overall survival of about 24 months and a progression free survival of 11.6 month for
Afatinib treatment vs 6,7 months for chemotherapy treatment [17-19]. 42,85% patients (3/7)
had reversible acneiform rash and no other adverse events were notice to TKIs treatment.
After disease progression, 42,85% patients (3/7) had pT790M mutation positive and
followed treatment with Osimertinib. After 5 months and, respectively, 6 months of treatment,
2 female patients had progressive disease and continued with palliative chemotherapy. One died
after 3 months of chemotherapy and the other patient is still under treatment. The third female
patient with Osimertinib treatment deceased after 3 months of treatment because of extensive
arterial thrombosis condescending.
Only 0,95% patients (2/210) had ALK gene rearrangement positive. One female patient
followed treatment with Crizotinib after disease progression post chemotherapy (at that time
Crizotinib was not refunded in Romania). The main adverse event was the increasing of serum
transaminases what required dose reduction of Crizotinib. On CT imaging evaluation after 6
months of treatment there has been noticed a partial remission, but with an exponential
progression in the next 2 months of treatment. The patient died after 8 months since the
initiation of Crizotinib. The second patient is a male one, followed treatment with Crizotinib
for 12 months without significant adverse reactions and no progressive disease. Treatment with
Crizotinib in the first line at patients with ALK gene rearrangements yields very high response
rate (>60%) and a median time to progression about 7 months to 1 year [20-22].
There has been initiated the immunotherapy with Pembrolizumab for 2 months, for one of
the 3 patients with PD-L1 positive >50% of tumour cells, following the established criteriain
Romania. First line treatment with Pembrolizumab for patients with metastatic non-squamous
or squamous lung cancer improve overall survival compared with chemotherapy (80,2% vs
72,4%) and show higher response rate versus chemotherapy [20-24].
6,19% patients (13/210) with NSCLC, EGFR mutations negative, ALK gene rearrangements
negative, PD-1<50% in tumour cells, followed targeted therapy or immunotherapy [20]. Taking
into account the experience from previous years, it was preferred a treatment with
Bevacizumab/Paclitaxel/Carboplatin for 0,95% of patients (2/210) with advanced or metastatic
lung cancer [25]. ADK and with Erlotinib, after the failure of several lines of chemotherapy at
2,38% patients (5/210) with ADK metastatic lung cancer.
Patients under treatment with Bevacizumab/Paclitaxel/Carboplatin have not progression of
the disease after 5 and 8 months of treatment. One patient on Erlotinib treatment died after 1
month after the beginning of Erlotinib. 3 patients have a favourable clinical evolution, waiting
for a CT evaluation in the next 3 months. 1 patient had minimum disease progression on CT
images after 16 months of treatment and had substantial clinical benefit, with the remission of
respiratory symptoms after the first 3 months of the treatment, without significant adverse
reactions. After the failure of one or several lines of chemotherapy, 2,85% patients (6/210)
followed a treatment with Nivolumab. 2 patients with ADK metastatic lung cancer are under
158
active treatment from 2 months, 2 patients with SCC metastatic lung cancer had progressive
disease after 5 months and 6 months, respectively, of treatment, 1 patient with SCC with partial
remission after 6 months of treatment died because of bronchopneumonia, 1 patient with SCC
metastatic lung cancer had a partial remission of disease after 6 months from the beginning of
Nivolumab, but with the progression of the disease on CT images after other 3 months of
treatment (Fig. 3).
Fig. 3. Targeted Therapy/Immunotherapy in NSCLC

7
6
6
5
5
4
4
3 3
3
2
2
11 1
1
00 0 00 00 0
0
Bevacizu Osimerti Crizotini Nivolum Pembrol
Erlotinib Afatinib
mab nib b ab izumab
First line 2 4 3 0 1 1
Progression after chemotherapy 0 5 0 0 1 6
Progression after EGFR TKIs 0 0 0 3 0
Checkmate -057 trial showed an increased median overall survival at patients who received
Nivolumab compared to Docetaxel (12,2 vs 9,4 months) and a median duration of response
higher for patients with Nivolumab compared to those in treatment with Docetaxel (17,2 vs 5,6
months) [26].
4. Conclusions
Targeted therapies and immunotherapy prove to be a substantial benefit for the treatment of
patients with non-small cell lung cancer, with increased median overall survival, response rate,
and progression free survival. The profile of mutations/alterations of genes in NSCLC
maximizes the benefits from targeted therapy and immunotherapy. A challenge of targeted
therapies is represented by the development of mechanisms of resistance to the treatment with
EGFR TKsI or ALK TKIs, after 7-11 months of treatment.
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159
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160
Palliative Treatment of Severe Dysphagia in

Advanced Cancer Patients
RAHNEA-NITA Gabriela1,2, CIUHU Anda-Natalia1,

RAHNEA-NITA Roxana-Andreea1, STOIAN Alexandru-Rares2,3
1 Department of Oncology – Palliative Care, “Sf. Luca” Chronic Disease Hospital, Bucharest (ROMANIA)
3 Department of Surgery, “Bagdasar Arseni” Emergency Hospital, Bucharest (ROMANIA)
* All authors had equal scientific contribution and share first authorship.
Emails: gabriela_rahnea@yahoo.com, andadum@yahoo.com, roxana_rahnea@yahoo.com, dr.raresstoian@yahoo.com
Abstract
Introduction
Severe dysphagia is the main symptom in patients with advanced head and neck and
oesophageal cancers. Swallowing disorders can affect nutritional status of the patients and may
delay or contraindicate specific curative or palliative treatment – radiotherapy, surgery and
chemotherapy. In order to increase survival and quality of life it is important to find the most
appropriate medical solution to alleviate this symptom, depending on the causes, performance
status and the patient’s will.

All the patients admitted to Oncology Department of the Chronic Disease “St. Luke”
Hospital, in June-December 2019 period, were assessed for mainly symptomatology caused by
cancer disease. Patients with severe swallowing disorders were evaluated by patient’s medical
history, symptom’s history and characteristics, cancer disease site, stage and complication.
Results and Discussions

Severe dysphagia was mainly diagnosed to patients with head and neck cancers. 42,18%
(28/64) and, respectively, 10.93% (7/64) of these required symptomatic relief by PEG or
nasogastric feeding tube. For lung cancer patients who had severe dysphagia, most commonly
by eso-tracheal fistula, oesophageal stent fitting was the most appropriate medical solution –
75% patients (3/4).
Conclusions
Palliative treatment of severe dysphagia through PEG, oesophageal stent, nasogastric
feeding probe, depending on the indication, allows maintaining the nutritional status of patients
with advanced cancer and the administration of the specific treatment – radiotherapy,
chemotherapy, with minimal and rare complications. If the performance status of the patient is
severely impaired, parenteral nutrition, hydration, corticosteroids, and specific analgesia are the
right short-term solutions, in palliative care.
Keywords: severe dysphagia, advanced cancer, palliative care
1. Introduction
The incidence of dysphagia, defined as any dysfunction in swallowing vary considerably in

cancer patients depending on the site and stage of the disease [1, 2]. Most commonly, this
161
symptom is encountered in head and neck cancers, in oesophageal cancers or gastric cancers
localized pericardial [1]. Severe dysphagia is an important symptom in cancer patients because
it causes an important nutritional deficiency, contributing along with other symptoms such as:
inappetence, nausea, vomiting, hypercomsumtiv syndrome, at the appearance of cachexia. The
causes of dysphagia in cancer patients are multiple: primary/metastatic tumour compression or
invasion, specific antineoplastic treatment (radiotherapy, treatment with Epidermal Growth
Factor Receptor tyrosine kinase inhibitors – EGFR TKIs), paraneoplastic or associated
pathologies [3-6], (Table 1).
Table 1. Causes of severe dysphagia related to advanced cancer disease

Causes by cancer process Caused by specific treatment
Obstruction by mass lesion in mouth, pharynx or Surgery (loss of structure, motor loss, fibrosis, fistula)
oesophagus
External compression (mediastinal tumours) Radiotherapy (fibrosis, mucosal inflammation, fistula)
Perineural tumour spread Chemotherapy (mucosal inflammation)
Upper motor neurone damage (cerebral tumours) Neuroleptics, anticholinergic, metoclopramide drugs
Paraneoplastic neuropathy
Dysphagia is an important symptom that requires prompt medical evaluation. In patients

with head and neck cancers (especially subcricoidian lesions) and with oesophageal tumours
(most commonly located in 1/3 medium of the oesophagus), they usually are presented to the
doctor for swallowing disorders, most frequently, severe.
The management of severe dysphagia in cancer patients requires the correct evaluation of
the symptom (oropharyngeal/oesophageal dysphagia, acute/chronic, causes, existing
complications). The palliative treatment of dysphagia mainly aims at maintaining or
maximizing the nutritional status of the patient, avoiding major complications, increasing the
quality of patients’ lives [7-13]. The therapeutic options are: non-oral feeding, corticosteroids,
proton pump inhibitors, resection or bypass surgery, radiotherapy/palliative chemotherapy,
percutaneous endoscopic gastrostomy (PEG), standard nasogastric tubes (SNT), endoluminal
stenting [14-18].
We reviewed database for all patients admitted in the Oncology – Palliative Care Department
of Chronic Disease “St. Luke” Hospital Bucharest over a 6-months period – June -December
2019 who had palliation of malignant dysphagia. At presentation, dysphagia was reviewed by
the characteristics of the symptomatology (oropharyngeal/oesophageal dysphagia,
chronic/acute, difficulty or avoidance of some food consistencies, nutritional status), medical
history (any history of swallowing difficulties, presence of a feeding tube or oesophageal stent,
any prior surgery to the oral, pharynx, oesophageal or gastric areas, history of pneumonia or
other respiratory diseases, radiotherapy to the thorax or throat or systemic chemotherapy, any
other associated disease),anterior imaging (computer tomography, MRI, chest x-rays, barium
contrast studies) or endoscopic evaluation.
In the period from June to December 2019 there have been hospitalized in the Department
Oncology – Palliative Care of Chronic Disease “St. Luke” Hospital Bucharest a number of 964
patients with cancer. 6,95% patients (67/964) when came to be hospitalized they presented
severe dysphagia. Severe symptoms have been noticed mainly at the patients with head and
neck cancer – 62,5% patients (45/72), as follows: 2,77% (2/72) with supracricoid tumours and
162
59,72% with subcricoid tumours. 84,31% (43/51) of the patients with subcricoid tumours
presented upon hospitalization severe swallowing disorders, compared to 9,52% of the patients
with supracricoid tumours. As well as, severe dysphagia was found at 1,86% (18/964) patients
with digestive cancer, as follows: 0,72% (7/964) with gastric cancers, representing 11,47% of
the evaluated patients with gastric cancer, 0,82% (8/964) patients with oesophageal cancer
representing 40% of the hospitalized patients with oesophageal cancer and 0,31% patients with
pancreatic cancers (3/964) (Figure 1, Figure 2, Figure 3).
Fig. 1. Incidence of severe dysphagia at patients with advanced cancer admitted

in Oncology Department – June-December 2019
Fig. 2. Incidence of severe dysphagia by cancer site
Fig. 3. Incidence of acute/chronic severe swallowing disorders in patients with severe dysphagia
163
From the total number of patients with total dysphagia, 11,94% (8/67) presented acute
dysphagia – 4,47% (3/67) having bronchopulmonary cancers, 4,47% (3/67) with pancreatic
cancers and 2,98% with oesophageal cancers. The evaluation of the patients with acute
dysphagia was done by barium contrast test and revealed that 2 of the 3 patients with lung
cancer presented tracheoesophageal fistula and they were treated by oesophageal stent, the other
6 patients (1 with lung cancer, 2 with oesophageal cancer and 3 with pancreatic cancer)
presented intrinsic and extrinsic obstructive mechanism. Patients with oesophageal cancer were
sent to the Surgery Department to have installed a PEG tube.
Of the 59 patients with chronic swallowing disorder, 22,03% (13/59) refused the installation
of a stent, gastrostomy or jejunostomy, that had been prescribed and they opted for the
parenteral treatment and oral adequate nutrition. At 1,69% (1/59) patients with lung cancer and
severe swallowing disorders it was preferred the hydroelectrolytic and nutritional parenteral
rebalance because of deficitary performance status.
Some of the patients with oesophageal or gastric pericardial cancers (13 patients) with severe
chronical swallowing disorders, 53,84% (7/13) and 23,07% (3/13) had, in their recent medical
history, installed a PEG/oesophageal stent in order to maintain the nutritional status during the
period of chemotherapy and palliative radiotherapy. 42,18% (28/64) and 10.93% (7/64) of the
patients with head and neck cancers with severe dysphagia benefited from pre-therapeutic PEG
installation, respectively, a nasogastric feeding tube for nutrition (Figure 4).
Fig. 4. Specific management of severe dysphagia by cancer site
During the period of hospitalization and periodical evaluation of the patients, 13,51% (5/37)
of the patients with PEG presented the following complications: skin infections, tube blockage
or tube displacement and 28,57% (2/7) of patients with nasogastric tube had tube displacement
[19, 20].
Corticoterapy, treatment with proton pump inhibitors, antalgic treatment and
hydroelectrolyticrebalancing were addressed to all the patients with severe dysphagia who were
hospitalized in our department [21-23].
4. Conclusions
Main interventions against severe dysphagia at patients with advanced cancer, the most
frequent being the mechanic one, are the installation of PEG, oesophageal stent, nasogastric
tube for nutrition. The preference of the medical staff is to install a PEG tube for nutrition to
avoid the risk of tumour invasion of the stent devices. Severe swallowing disorders represent a
164
major problem in the care of the patients with head and neck cancers, as well as of the patients
with digestive cancers (oesophageal, gastric and pancreatic cancer). Parenteral nutrition has
short term benefits – days and weeks, not being indicated for the patients whose symptoms
persist for more than 10 weeks.
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18. Bolocan, A., Paduraru, D.N., Nitipir, C., et al., (2018). Mixed a denoneuroendocrinecarcinoma of the
gastrointestinal tract-features, diagnosis, management and prognostics. Romanian Biotechnological
Letters 23(6), pp. 14193-14202.
19. Ciuhu, A.N., Pantea-Stoian, A.M., Nitipir, C., et al., (2017). Assessment of cachexia in cancer patients
with advanced disease, Conference: 3rd International Conference on Interdisciplinary Management of
Diabetes Mellitus and its Complications (INTERDIAB) Location: Bucharest, ROMANIA, pp. 139-147.
20. Nitipir, C., Diaconu, C.C., Orlov, C., et al., (2018). The Necessity of Nutritional Intervention in the
Oncological Patient. What is the Evidence? Conference: 35th Balkan Medical Week on Healthy Ageing
– An Endless Challenge Location: Athens, pp. 133-137.
21. Grigorean V.T., Rahnea Niță G., Georgescu L., Sandu A.M., Tuinea L.A., Ciuhu A.N., (2014).
Correlations entre l evaluation des symptoms et de la qualite de vie chez les patients oncologues, Archives
of the Balkan Medical Union, Bucharest, vol. 49, pp. 139-148.
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166
The Impact of Chemotherapy on the Quality of Life in Patients

with Gynaecological Cancer
RAHNEA-NITA Roxana-Andreea1, RAHNEA-NITA Gabriela2,3,

ANGHEL RODICA Maricela2,4
1 Department of Oncology, “Constantin Gorgos” Psychiatric Hospital, Bucharest (ROMANIA)
3 “Sf. Luca” Chronic Disease Hospital, Bucharest (ROMANIA)
4 “Prof. Dr Alexandru Trestioreanu” Oncological Institute, Bucharest (ROMANIA)
* All authors had an equal scientific contribution to this research
Email: gabriela_rahnea@yahoo.com
Abstract
Quality of life is a concept with multiple dimensions and is regarded as a subjective report
of patients experience related to the neoplastic disease.
Given the premises that life quality is a prognostic and predictive factor for survival, we
have analysed the relationship between the physical, the psychological symptoms and the
activity as well as their impact on the quality of life among female patients with advanced and
terminal stage gynaecological cancer. Also, the aim of this article was to assess the role of
chemotherapy on the quality of life.
Keywords: gynaecological cancer, quality of life, chemotherapy
1. Introduction
Quality of life is what the patient says it is. It is a concept with multiple dimensions and is
regarded as a subjective report of patients experience related to the neoplastic disease [1].
Impairment of quality of life is not only a public health component commonly found in
patients with oncological diseases but is also common in other patients with chronic conditions
such as diabetes mellitus, obesity, severe cardiovascular disease, chronic kidney disease in
advanced stages and also after surgical interventions [2-4].
A single review of interventions to enhance the quality of life for gynaecological cancer
patients included few articles who met the inclusion criteria regardless of the method, sample
size and instruments employed [1].
We also mention that very few studies regarding the quality of life in elderly patients with
cancer were reported [5-8].
Work hypothesis
Given the premises that anxiously depressive disorders are predictive factors for life quality
and that life quality is a prognostic and predictive factor for survival, we have analysed the
relationship between the symptoms (physical and psychological) and the activity, and also, their
impact on the quality of life among female patients with advanced and terminal stage
gynaecological cancer [9].
The objectives of the paper

• The assessment of the role of physical and psychological symptoms on the quality of
life;
167
• The assessment of the role of activity (mobility and social activity) on the quality of
life;
• The assessment of the quality of life by age category;
• The role of the specific oncological therapies on the quality of life.
Experimental part
We conducted a prospective study over a period of two years (2016-2018), in 124 patients
with advanced and terminal stage of gynaecological cancer (cervical, endometrial and ovarian)
admitted to “Prof. Dr. Alexandru Trestioreanu” Oncological Institute and “Saint Luca” Hospital
of chronic diseases – Chronic Oncological – Palliative Care Unit.
The 124 patients included in this study were divided into two categories, by age (under 55
years and over 56 years), depending on the median. The patients were evaluated at first
admission, after one month, after three months, utilizing one questionnaire:
Rotterdam Symptom Checklist – RSCL [10-13] [Appendix 1].
The patients completed the questionnaires during their admission in the hospital or by
telephone interview.
After six months, the patients received a phone call asking them to assess the quality of their
life at the time of the call. The patient’s record was drawn at first admission, which included
data regarding the age, the background, the studies, as well as the therapies undergone by the
time of admission.
The patients freely agreed to participate in the study by signing the informed consent.
The approval from the Ethical Commission has been obtained.
The data were statistically processed in the SPSS program.
Statistical medical research methods were used [14].
3. Results
The patients in the first group were aged between 29 and 87 years, with a mean age of 57
years (more precisely 56.9 years) and a median of 56 years.
The fact that the median is 56 years justifies, from a statistical point of view, the separation
of patients into two approximately equal groups of age. It is presented in Table 1.
Table 1. Patients distribution according to the age category

Frequency Percentage
Valid The first (under55years) 58 46.8
The second (56years or more) 66 53.2
Total 124 100.0
The distribution of the patients according to the age category and the diagnostic is presented
in Table 2.
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Table 2. Patients distribution according to the age category and the diagnostic
Diagnostic
Cervical Endometrial Ovarian Total
The age category The first (under 55 years) 40 0 18 58
The second (56 years or more) 38 4 24 66
Total 78 4 42 124
Visit 1. The data regarding the quality of life – for the 124 patients initially enrolled in the
study – are presented in the Table 3.
Table 3. Quality of life – Visit 1

Quality of life – eval. 1 excellent 4
good 31
moderately good 15
neither good nor bad 29
rather poor 18
poor 14
extremely poor 13
Total 124
Visit 2. Until the second evaluation, 21 patients were “loss of sight”, a number of 103 patients
remaining in the study.
The following table indicates that for the majority of patients the initial assessment is
retained; only for a number of 11 patients, a positive evolution is noticed (marked by bolded
figures), while for a number of 20 patients a negative evolution is noticed (marked by underline
figures) (Table 4).

Quality of life – eval.2
moderately neither rather extremely
excellent good good good nor poor poor poor Total
bad
Quality of life – excellent 3 1 4
eval.1 good 2 19 6 3 1 31
moderately good 1 10 2 1 14
neither good nor bad 2 2 20 2 2 28
rather poor 2 12 1 1 16
poor 1 8 9
extremely poor 1 0 1
Total 5 24 18 28 14 12 2 103
Visit 3. Another five patients were “loss of sight” by the third evaluation, with only 98
patients remaining in the study.
The following table (Table 5) indicates that, also, in this case, the initial evaluation is retained
for the majority of patients; for a number of 20 patients a positive evolution is noticed (marked
by bolded figures), while for a number of 14 patients a negative evolution is noticed (marked
by underline figures).
169

Quality of life – eval.3
moderately neither rather poor extremely
excellent good good good nor poor poor Total
bad
Quality of life excellent 4 1 5
– eval.2 good 2 21 1 24
moderately good 7 7 1 1 16
neither good nor bad 5 4 13 3 3 28
rather poor 1 8 3 12
poor 1 10 1 12
extremely poor 1 0 1
Total 6 35 13 15 11 17 1 98
Phone call. Another nine patients were “lost” by the time of the telephone call, while only
89 patients remained in the study.
The following table (Table 6) indicates that the initial evaluation is retained for the majority
of patients; only for a number of 7 patients, a positive evolution is noticed (marked by bolded
figures), while for a number of 26 patients a negative evaluation is noticed (marked by underline
figures).
Table 6. Quality of life – Phone call

Quality of life – eval. phone call
moderately neither rather extremely
excellent good good good nor poor poor poor Total
bad
Quality of life – excellent 4 2 6
eval.3 good 1 27 3 1 32
moderately good 1 6 3 1 1 12
neither good nor bad 1 8 5 14
rather poor 3 3 4 10
poor 1 8 6 17
extremely poor 0 0
Total 5 30 10 14 10 13 7 89
The evolution of life quality assessments is shown in the following tables, given the two age
categories (Table 7, Table 8):
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Table 7. The age category – The first (under 55 years)

Quality of life Evaluation Crosstabulation
Evaluation
I II III Phone call
Quality of life excellent 2 3.4% 2 3.4% 3 5.2% 4 6.9%
good 13 22.4% 13 22.4% 21 36.2% 16 27.6%
moderately good 7 12.1% 9 15.5% 6 10.3% 8 13.8%
neither good nor bad 19 32.8% 16 27.6% 8 13.8% 7 12.1%
rather poor 6 10.3% 5 8.6% 3 5.2% 3 5.2%
poor 5 8.6% 7 12.1% 9 15.5% 5 8.6%
extremely poor 6 10.3% 1 1.7% 1 1.7% 3 5.2%
Lost of the study 0 2 3.4% 2 3.4% 5 8.6%
deceased 0 3 5.2% 5 8.6% 7 12.1%
Total 58 100.0% 58 100.0% 58 100.0% 58 100.0%
Table 8. The age category = The second (over 56 years)

Quality of life Evaluation Crosstabulation
Evaluation
I II III Phone call
Quality of life excellent 2 3.0% 3 4.5% 3 4.5% 1 1,5%
good 18 27.3% 11 16.7% 15 22.7% 15 22.7%
moderately good 8 12.1% 9 13.6% 7 10.6% 2 3.0%
neither good nor bad 10 15.2% 12 18.2% 6 9.1% 6 9.1%
rather poor 12 18.2% 9 13.6% 8 12.1% 7 10.6%
poor 9 13.6% 5 7.6% 8 12.1% 8 12.1%
extremely poor 7 10.6% 1 1.5% 0 4 6.1%
Lost of the study 0 6 9.1% 6 9.1% 7 10.6%
deceased 0 10 15.2% 13 19.7% 16 24.2%
Total 66 100.0% 66 100.0% 66 100.0% 66 100.0%
4. Discussion
Rotterdam Symptom Checklist has permission for use for scientific research [10], [11]. Full
inclusion of the questionnaire on a website is only allowed for online responding to the
questionnaire, for scientific research [10-12].
We analyse the results of the Rotterdam Symptom Checklist (RSCL).
In the first category of age, the most common value is the neutral one (“neither good nor
bad”) at visit 1 and visit 2, while at visit 3 and the phone call, the most frequent value is “good”.
Regarding negative values “rather poor”, “poor” and “extremely poor” and “deceased”, at
all evaluation, the percentages are higher at the second category of age, compared to the first
one.
These percentages indicate that patients from the first category of age have a better quality
of life compared to the second category of age. An interesting aspect regarding the quality of
life that emerged is related to the comparison between the two age categories. If the results are
roughly similar for the first two evaluations, respectively on Visit 1 (p-value = 0.375) and on
visit 2 (p-value = 0.059), statistically significant differences are noticed upon final evaluations
(at visit 3, p-value = 0.021 and a phone call, p-value = 0.008), the balance being in favour of
the first category. Practically, it turns out that younger patients have a better quality of life
compared to elderly ones.
171
Another statistically significant difference is the difference between the third evaluation (p-
value = 0.002) and the 6-month telephone call (p-value = 0.001). Unfortunately, they refer to
the worsening of the quality of life for both categories of age.
As for the means of treatment, the two groups of age were compared according to the
therapeutic method from which they benefited.
As for the surgical treatment, the distributions were similar, respectively, approximately
50% of the patients underwent surgery.
As for radiotherapy, the Z-dimensional assay concluded that the ratio of those who
underwent radiotherapy is higher among the first category of age and indicated the statistical
significance of the statement. Several 51.7% of the first category benefited from radiotherapy,
while only 34.8% of the second category did.
Moreover, as for chemotherapy, different percentages were also noticed, the Z proportions’
test attaches a P value of 0.021 to the statement “the proportion of those who underwent
chemotherapy is higher in the first category of age” (while the Fisher test exactly 0.032), which
indicates the statistical significance of the statement.
Regarding the state of patients’ life quality in both categories (whether they have had
chemotherapy or not), at each and every visit: the p values are obtained with the help of the
Mann-Whitney test and they all indicate that the statement according to which the patients who
have undergone chemotherapy have a better quality of life (than those who did not) is
statistically significant.
We notice that many patients in the category of those who did not undergo chemotherapy
resulted “deceased” by the time of the telephone call.
The overall evolution of the quality of life for each category of patients is decreasing.
As a result of all the above mentioned, the treatment has a positive impact on the
improvement of the quality of life.
Regarding the “Rotterdam-physical symptoms” scale: The P values indicate differences
between the two groups, at all visits, in favour of the first category of age. (visit 1 – p-
value=0.012, visit2 – p-value=0.032, visit 3 – p-value=0.013).
For all visits, as a result of Mann-Whitney test, the statement according to which patients in
the second category had a lower quality of life is statistically significant.
Regarding the “Rotterdam – psychic symptoms” obtained from patients at their three visits:
The p-values suggest differences between the two groups at all visits in favour of the first
category of age who had a better quality of life in terms of the psycho-emotional state, compared
to patients in the second category (visit 1 – p-value=0.010, visit 2 – p-value=0.019, visit 3 – p-
value=0.007).
Regarding the activity values obtained from patients at their three visits: The p values do not
suggest any differences between the two groups at any visit. (visit 1 – p-value=0.190, visit 2 –
p-value=0.709, visit 3 – p-value=0.654). As for Activity, the statistical tests indicate the
existence of some statistically significant decrease of activity on visit 2 compared to visit 1 for
the first category, and on visit 3 compared to visit 2 for the second category.
However, a very strong positive correlation between the Quality of Life and the Activity
parameter was noticed, as well as a negative correlation between the Quality of Life and the
Rotterdam – physical symptoms, so it could be concluded that the higher the Activity, the better
the Quality of Life and the more pronounced the Symptoms, the lower the Quality of Life [15-
17].
172
5. Conclusions
As far as the statistical conclusions are concerned, the following were reflected as a result:
• The conclusion related to the quality of life by age category (under 55 years and over
56 years) is that the results were statistically different, the difference being that the
patients under 55 years had a better evolution of the quality of life compared to those
over 56 years;
• Regarding the quality of life in terms of whether a specific treatment was undergone or
not, it was found that the patients who underwent radiotherapy or chemotherapy had a
better quality of life compared to those who did not undergo these treatments;
• In connection with the RSCL, the patients in the first category experienced less
accentuated symptoms than those in the second category from both a physical and
psychological point of view;
• Both categories had decreased Activity, namely, the first category had a negative change
from visit 2 to visit 3, while the second category had a negative change from visit 1 to
visit 2;
• The crucial conclusion to which this study led is the higher the activity and the less
pronounced the symptoms, the better the quality of life.
Gynaecological cancers have a substantial impact on the patient's physical, psychological

and emotional status, and the professionals who treat the patients must monitor their effect on
the quality of life.
Interventions to improve gynaecological cancer patients’ quality of life should help asses
and target the individual physical, psychological, and emotional symptoms of the patient.
Acknowledgement
This article is a result of the research of Roxana-Andreea Rahnea Nita, MD, within the
doctoral studies (2015-2019) at UMF “Carol Davila” Bucharest under the coordination of
Prof.Dr. Rodica Anghel, Md, PhD.
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Appendix 1
Rotterdam Symptom Checklist

date of completion 19
In this questionnaire, you will be asked about your symptoms. Would you please, for all
symptoms mentioned, indicate to what extent you have been bothered by it, by circling the
answer most applicable to you. The questions are related to the past week.
Example: Have you been bothered, during

the past week, by headaches
not at all a little quite a bit very much
Have you, during the past week,
not at all a little quite a bit very much
been bothered by lack of appetite
Irritability not at all a little quite a bit very much
Tiredness not at all a little quite a bit very much
Worrying not at all a little quite a bit very much
Sore muscles not at all a little quite a bit very much
Depressed mood not at all a little quite a bit very much
Lack of energy not at all a little quite a bit very much
Low back pain not at all a little quite a bit very much
Nervousness not at all a little quite a bit very much
Nausea not at all a little quite a bit very much
Despairing about the future not at all a little quite a bit very much
Difficulty sleeping not at all a little quite a bit very much
Headaches not at all a little quite a bit very much
Vomiting not at all a little quite a bit very much
Dizziness not at all a little quite a bit very much
Decreased sexual interest not at all a little quite a bit very much
Tension not at all a little quite a bit very much
Abdominal (stomach) aches not at all a little quite a bit very much
Anxiety not at all a little quite a bit very much
Constipation not at all a little quite a bit very much
Diarrhoea not at all a little quite a bit very much
Acid indigestion not at all a little quite a bit very much
Shivering not at all a little quite a bit very much
Tingling hands or feet not at all a little quite a bit very much
Difficulty concentrating not at all a little quite a bit very much
Sore mouth/pain when swallowing not at all a little quite a bit very much
Loss of hair not at all a little quite a bit very much
Burning/sore eyes not at all a little quite a bit very much
Shortness of breath not at all a little quite a bit very much
Dry mouth not at all a little quite a bit very much
175
A number of activities is listed below. We do not want to know whether you actually do
these, but only whether you are able to perform them presently. Would you please mark the
answer that applies most to your condition of the past week?
without help, with

unable only with help without help
difficulty
care for myself (wash etc.) 0 0 0 0
walk about the house 0 0 0 0
light housework/household jobs 0 0 0 0
climb stairs 0 0 0 0
heavy housework/household jobs 0 0 0 0
walk out of doors 0 0 0 0
go shopping 0 0 0 0
go to work 0 0 0 0
All things considered; how would you describe your quality of life during the past week?
excellent 0
good 0
moderately good 0
neither good nor bad 0
rather poor 0
poor 0
extremely poor 0
Would you please check whether you answered all questions?

Thank you for your help.
Patient number.......
176
Hypertension and Diabetes Mellitus throughout History
GAIȚĂ Laura-Adriana1, TIMAR Romulus Zorin1,2, TIMAR Bogdan1,2

1“Victor Babeș” University of Medicine and Pharmacy, Timisoara (ROMANIA)
2“Pius Brînzeu” Emergency Hospital, Timisoara (ROMANIA)
Emails: gaita.laura@umft.ro, timar.romulus@umft.ro, timar.bogdan@umft.ro
Abstract
The management of arterial hypertension has undergone multiple changes throughout recent
history. From 1957 when it was shown that this condition increases the risk of developing the
so-called arteriosclerotic heart disease, to the near future where its approach involves the digital
sector, artificial intelligence, biotech and population sciences, hypertension has been diagnosed
and treated in different approaches. Its increasing prevalence alongside another epidemic of the
modern world, diabetes mellitus, with their amplified risk of cardiovascular events, led to more
and more effective ways of assessing their consequences, to updated recommendations and to
complex algorithms of pharmacologic agents and lifestyle changes. The new antidiabetic drugs,
the GLP-1 receptor agonists and the SGLT-2 inhibitors, not only lower the blood glucose levels,
but also have protective cardiovascular effects and contribute to safely lowering the blood
pressure. This review represents an overview on the beginnings of hypertension as a
cardiovascular risk factors, of the perspectives of approach and of the current European
guidelines on hypertension in patients with diabetes mellitus. In other words, it is about
hypertension and diabetes mellitus yesterday, today and tomorrow.
Keywords: arterial hypertension, diabetes mellitus, atherosclerotic cardiovascular disease
1. Introduction
Hypertension is considered to be one of the most important cardiovascular risk factors.

However, this was not always the case. Not long ago, in the recent history, in 1947, the U.S.
Public Health service began to plan a large epidemiological study regarding cardiovascular
disease, the Framingham Study. This research shed light upon the gaps in knowledge about one
of the main causes of mortality of the mid-20th century, disease with a growing epidemic trend
[1]. In 1957, the first results from the 4 years follow-up were published with the main focus on
the so-called arteriosclerotic heart disease (ASHD). It was shown that high values of blood
pressure increase the risk of developing ASHD, with a gradient of risk starting with
normotensive individuals and followed by patients with borderline hypertension, possible
hypertensive heart disease, definite hypertension and definite hypertensive heart disease. The
conclusion that was drawn is that high blood pressure is a cardiovascular risk factor, followed
by adding high levels of serum cholesterol on the list [2].
2. Material and Methods
The findings from the Framingham Study have shaped the world of cardiology and soon
after the first published work, the concept of “risk factors” was defined by Dr. William B.
Kannel [3]. Fast forwarding to the present, arterial hypertension is the main preventable cause
of death, with a prevalence of 1 in 3 adults with this condition [4] and with an even higher
prevalence in Romania – 45,1% of the Romanians aged between 18 and 80 [5]. Another
177
epidemic of the modern world is diabetes mellitus (DM). In 2019, the International Diabetes
Federation estimated that worldwide there are 463 million patients with this disease – or 1 in
11 adults – and that 1 in 2 adults are undiagnosed [6]. DM itself doubles or even triples the risk
of developing atherosclerotic cardiovascular disease (ASCVD) [7]. Moreover, hypertension
and DM not only increase the risk of a cardiovascular event, they are also frequently associated,
sometimes included with dyslipidaemia and abdominal obesity in the metabolic syndrome. The
two conditions are interlinked through endothelial dysfunction, vascular inflammation, arterial
remodelling, atherosclerosis, dyslipidaemia and obesity.
Hypertension is twice as prevalent in diabetic patients that in those without and the presence
of both in a patient further increases the cardiovascular risk, the risk of developing an acute
myocardial infarction, a stroke or heart failure [8, 9].
This amplified risk of cardiovascular events leads to a necessity of an effective screening
and an early diagnose of hypertension and DM, with an interdisciplinary approach of these
conditions and with treating to target not only the blood pressure and the glycemia, but also all
the existing cardiovascular risk factors in order to achieve cardio-, reno-, neuro- and metabolic
protection. The importance of a collaboration between diabetologists and cardiologists was
suggested in the guidelines already in 2013 when the European Society of Cardiology
Guidelines on diabetes, pre-diabetes and cardiovascular diseases developed in collaboration
with the European Association for the Study of Diabetes suggested an algorithm in investigating
patients that have a primary diagnosis of DM or CVD. If the main diagnosis is DM, several
tests should be performed in order to assess either the presence or the status of CVD-ECG,
echocardiography, exercise test and Holter monitoring. After the evaluation, a cardiology
consultation and subsequently the possibility of invasive or non-invasive ischemia treatment
should be considered, if any of the tests have a positive result. Similarly, if the main diagnosis
is CVD, patients should be advised to check their fasting plasma glucose levels, HbA1c, blood
lipids and, if needed, to perform an oral glucose tolerance test and a screening for
microangiopathy, followed by a diabetology consultation [10].
In 2018, the European Society of Cardiology in collaboration with the European Society of
Hypertension released a new guideline regarding the management of arterial hypertension.
One of its recommendations regarding patients with DM suggests not only frequently
measuring the blood pressure in office, but also to consider using a 24-hour ambulatory blood
pressure monitoring in apparently normotensive individuals with DM, due to the frequent
association of these two risk factors. And while this guideline maintained the same
classification of office blood pressure and the definitions of the hypertension grade, it included
a new classification of hypertension, in stages, according to blood pressure levels, the presence
of cardiovascular risk factors, hypertension-mediated organ damage (HMOD) and
comorbidities. The rationale behind this classification lies partly in the fact that not all features
of HMOD are included in the SCORE system, such as pulse pressure, carotid-femoral pulse
wave velocity, left ventricular hypertrophy, microalbuminuria or advanced retinopathy, and that
they lead to an increased cardiovascular risk even in the absence of the classical risk factors
[11]. The role of hypertension in the development of microalbuminuria and retinopathy proves
the contribution of this condition not only to the macrovascular complications of DM, but also
to the microvascular ones. The predominant role is, however, the one on the progression of
diabetic nephropathy through glomerular hyperfiltration [12].
Consequently, to prevent the development of complications or to slow the progression of the
existing ones, it is of extreme importance to have a correct approach of both hypertension and
DM. One of the first tasks is to choose the adequate goals for our patients. Similar to the
178
glycaemic targets, when planning the antihypertensive treatment, the desired goals should be
individualized according to the patient’ characteristics. Although we should consider the age,
the presence of ASCVD, the presence of other comorbidities, the risk of progression of chronic
kidney disease, the presence of retinopathy or the tolerance of the patient for different values
of blood pressure, the 2018 guideline recommends, for patients with DM aged between 18-65
to target the systolic blood pressure between 120-130 mmHg and for the patients over 65 years
to target the systolic blood pressure between 130-139 mmHg, if tolerated. For both age groups,
the goal for the diastolic blood pressure is between 70-79 mmHg. Pharmacologic treatment of
hypertension should be initiated when the systolic blood pressure is higher than 140 mmHg in
patients under 80 years old and higher that 160 mmHg in older patients, or in both age groups
when the diastolic blood pressure is higher than 90 mmHg. Similar targets, of a systolic blood
pressure of 130-139 mmHg, and not lower due to reduced renal perfusion pressure, are
recommended for patients with diabetic chronic kidney disease and hypertension. Moreover,
when choosing the blood pressure goal, besides the tolerance of the treatment, we should always
assess the presence of postural hypotension that can be caused by diabetic autonomic
neuropathy [11].
Regarding the individualization of treatment, the same principle applies to choosing the
pharmacological antihypertensive agent. Although all blood pressure lowering drugs are
beneficial in patients with DM and hypertension, multiple studies prove the importance of
renin-angiotensin system blockers that are proven to be the only class of drugs that are more
effective in DM than in individuals without DM when reducing the cardiovascular risk [13].
Important positive effects of angiotensin-converting enzyme inhibitors (ACE inhibitors) and
angiotensin II receptor blockers (ARBs) is the reduction of albuminuria and the progression of
diabetic nephropathy. The benefits apply not only to patients with albuminuria, but also to
patients with type 2 DM and normo-albuminuria in whom the risk of developing
microalbuminuria is reduced [14]. Therefore, the recommended algorithm in the management
of hypertension in patients with DM starts with a combination of a renin-angiotensin system
blocker with a calcium channel blocker or a thiazide/thiazide-like diuretic, while the
simultaneous administration of an ACE inhibitor and an ARB is not indicated because it
increases the risk of hyperkalaemia and of acute kidney injury. Monotherapy should be
considered in patients older than 80 years or in frail individuals. If the targets are not met, then
the second step comprises in the combination of an ACE inhibitor or and ARB, a calcium
channel blocker and a thiazide/thiazide-like diuretic. The next step is represented by the
previous triple combination with spironolactone or some other drugs. However, when
combining spironolactone and an ACE inhibitor or an ARB, potassium levels should be checked
regularly. Beta-blockers should be considered at any step if there is a specific indication for
their use [11, 15].
Nevertheless, treating hypertension in patients with DM also implies complex lifestyle
changes including diet, physical activity and smoking cessation. Regarding diet, the 2018
guidelines recommend a salt restriction to less than 5g per day, an alcohol consumption
restriction to less than 14 units per week for men and less than 8 units per week for women,
while also avoiding binge drinking and to increase the consumption of vegetables, fresh fruit,
low-fat dairy products, wholegrains, fish, nuts and olive oil, with a low consumption of red
meat [11]. These suggestions are based on the results of the Dietary Approaches to Stop
Hypertension (DASH) trial which has shown that consuming a diet rich in fruit, vegetables and
low-fat dairy product and simultaneously reduced in saturated fat and cholesterol lowered
systolic blood pressure and diastolic blood pressure. Another study, the DASH-Sodium trial
has shown that a sodium reduction has a significant impact on the blood pressure independently
on the effects of the DASH diet [16].
179
Physical activity is recommended on a regular basis – at least 30 minutes of moderate

dynamic aerobic exercise on 5-7 days per week. Although it induces an acute rise in blood
pressure, it is followed by a short decline below baseline [11]. An extensive meta-analysis
demonstrated that endurance training, dynamic resistance training, combined training and
isometric resistance training significantly reduce diastolic blood pressure and that all except
combined training reduce systolic blood pressure. Also, some of the analyses suggest that in
patients with hypertension endurance training might be superior to dynamic resistance training
or combined training, with the most important reductions in endurance training after shorter
exercise program durations at moderate to high intensity and under 21 minutes of weekly
exercise [17].
Lifestyle changes also include smoking cessation, with a recommendation of ensuring
supportive care and referral to smoking cessation programs [11]. This suggestion is based not
only on the fact that smoking is an important cardiovascular risk factor in itself, but also on
results that prove that heavy smoking is associated with a persistent rise in blood pressure and
also with an increase in blood pressure variability [18].
The 2018 guidelines also recommend body-weight control, aiming at a healthy body mass
index and waist circumference to reduce blood pressure and cardiovascular risk. The reduction
of weight towards an ideal body weight decreases blood pressure, while also reducing insulin
resistance and improving glycaemic control. [11] In consequence, bariatric surgery should have
favourable results on the cardiovascular risk factors, hypothesis proposed by a meta-analysis
on 6587 patients. Its data showed an overall cardiovascular risk reduction after bariatric surgery,
while a BMI reduction of 5 kg/m2 corresponded to a hypertension reduction of 27%, a type 2
DM reduction of 33% and a hyperlipidaemia reduction of 20% [19].
Furthermore, some of the antidiabetic agents, alongside the blood glucose lowering effects,
also have cardioprotective and blood pressure lowering effects. Metformin usually represents
the first-line drug for type 2 DM and although it is neutral on the body weight and on the blood
pressure, it has documented cardiovascular benefits. Other classes of antidiabetic drugs that are
neutral on the blood pressure are sulfonylureas and insulin, while thiazolidinediones are
associated with fluid retention and heart failure. The DPP-4 inhibitors are more or less neutral
on blood pressure and also on the cardiovascular risk, but saxagliptin was associated with an
increased risk of hospitalization for heart failure [20]. However, the GLP-1 receptor agonists
decrease the blood pressure – liraglutide and exenatide have been shown to reduce significantly
the systolic blood pressure with 1-5 mmHg. Even if the mechanisms of this effect remain
uncertain, a nervous system pathway, vasodilatation, diuresis and natriuresis are suggested [21,
22]. The blood pressure lowering effects are even more clear in SGLT-2 inhibitors. They are
linked to clinically relevant reductions both of the systolic blood pressure of 4-6 mmHg and
also of the diastolic blood pressure of 2-3 mmHg. The mechanisms include osmotic diuresis,
mild natriuresis, weight loss and possible indirect effects on nitric oxide release secondary to
reduced oxidant stress through an improved glycaemic control [23, 24].
And the therapeutic agents that can improve the cardiovascular risk in patients with both
hypertension and DM are being developed continuously and paradigms are frequently
changing. In order to improve the control of blood pressure, a transformative system-wide
approach is mandatory, a strategy that should include the digital sector, artificial intelligence,
data sciences, biotech and biomedical sectors, healthcare delivery an also population sciences.
The digital sector offers the opportunity to shift the care away from the hospitals and clinics
and into the daily setting, with a greater focus on prevention, from using technology-based
tools. These include mainly apps and devices for monitoring and control and for hypertension
promising technologies could enable home blood pressure measurements that can be
transmitted directly to health providers and the electronic medical record. This comprehensive
assessment would have better outcomes than the discontinuous office measurement and would
180
diagnose cases of masked hypertension [25].There are already studies made on new methods in
predicting systolic blood pressure through a machine learning technique, specifically an
artificial neural network [26].
4. Conclusions
Data science and artificial intelligence together with advances in genomics and
metabolomics could lead to the development of precision medicine and to major benefits from
big data, real-time evidence and real-life results. Consequently, biotechnology will focus on
precision medicine and will have the potential to develop new pharmacological agents that
could have a targeted effect on RNA or DNA or that could regenerate tissues affected by
hypertension – therefore could create a regenerative medicine [25]. Some of the drugs that are
currently under development focus on the complex cellular signalling pathways that modulate
vascular smooth muscle cell contraction and growth involved in the regulation of vascular
tonicity and some on the HMOD [27]. Furthermore, the healthcare delivery should implement
coordinated and integrated measures, with an interdisciplinary approach of the non-
communicable diseases, while involving also educational, political or economic decision
factors [25]. All these institutions could improve blood pressure values or the evolution of
hypertension through education programs, public awareness campaigns, legislation to limit salt
intake and to promote a healthy diet, encouraging physical activity through opening new public
spaces and facilities and supporting smoking cessation [28]. And all of the above are only the
beginning of the new era of the management of hypertension, a condition that less than 7
decades ago was not yet considered, with evidence, to have such a paramount importance in
enhancing the development of cardiovascular disease.
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Pharmacoepidemiologic Implications at Patients with Metabolic

Syndrome
GĂNCEANU-RUSU Ana-Roxana1,2, LUPUŞORU Elena-Cătălina1,
DIMA Nicoleta1,2, MĂRĂNDUCĂ Minela-Aida1,2, CLIM Andreea2,
POP Ana-Maria2, BĂDESCU Minerva-Codruţa1,2, TĂNASE Daniela-Maria1,2,
OUATU Anca1,2, REZUŞ Ciprian1,2
2Department of Internal Medicine III, “Sf. Spiridon” Clinical Emergency Hospital, Iasi (ROMANIA)
Emails: roxanarusu12@yahoo.com, celupusoru@yahoo.com, nicoleta2006r@yahoo.com, sminelaaida@yahoo.com,
andreea.clim13@yahoo.com, ana.strainu.16@gmail.com, codruta.badescu@gmail.com, tanasedm@gmail.com,
ank_mihailescu@yahoo.com, ciprianrezus@yahoo.com
Abstract
Metabolic syndrome is a major public health problem and a global challenge following
urbanization and sedentary life, having a complex etiopathogenesis and pathophysiology. It is
characterized by several interconnected biochemical, clinical and metabolic factors that directly
increase the risk of cardiovascular disease and type 2 diabetes causing increased mortality. The
current definition of metabolic syndrome is present if three or more of the following criteria:
high blood pressure, dyslipidaemia (increased triglycerides and/or low-intensity cholesterol
lipoproteins), hyperglycaemia, and central obesity.
Keywords: metabolic syndrome, cardiovascular diseases, mortality, pharmacology
1. Introduction
Clinical management of the metabolic syndrome is difficult because there is no GOLD

STANDARD method to prevent or ameliorate the entire syndrome, using specific
pharmacological agents or other treatment methods for each of the components involved
(hyperglycaemia, high blood pressure, atherogenic dyslipidaemia), resulting thus
polypharmacy. Within this model, targeting excess visceral fat/ectopic fat through healthy
eating, physical activity and, if necessary, pharmacotherapy, may be useful in further managing
the risk of cardiovascular diseases associated with the treatment of traditional risk factors [1].
2. Material and Methods
Patients with type 2 diabetes have been shown to have an increased prevalence of lipid
abnormalities, thus contributing to the occurrence of cardiovascular disease. Multiple clinical
studies have demonstrated the beneficial effects of drug therapy (with statins) on subjects with
or without cardiovascular diseases [2]. Meta-analyses, including data from more than 18,000
diabetes patients from 14 randomized statin therapy studies (mean follow-up of 4.3 years), show
a 9% reduction in overall mortality and a 13 reduction % of vascular mortality for each mmol/l
(39 mg/dl) when reducing low-intensity cholesterol lipoproteins [3]. A study was performed on
250 patients monitored in the 3rd Medical Clinic of the County Emergency Hospital “Sf.
Spiridon” Iasi, which analysed the pharmacoepidemiologic investigations carried out in patients
diagnosed with metabolic syndrome. The objectives of the study consisted of carrying out
clinical, paraclinical investigations of patients with metabolic syndrome, highlighting the
183
biochemical changes produced during the metabolic syndrome therapy and evaluating the
correlations between the biochemical markers and the specific medication in the same category
of patients. In order to carry out this study, the clinical observation sheet of the patient was used
as a working tool, with the elaboration of special records, regarding the clinical, biochemical
and treatment aspects.
The following inclusion criteria were used: age over 18 years, presence of metabolic
syndrome, absence of end-stage chronic kidney disease, absence of severe hepatic impairment.
A total of 143 patients returned to the clinic for reassessment after 12 months of treatment.
All data were collected and centralized, keeping the confidentiality of the patient’s personal
information. The data were centralized and statistically processed using SPSS software version
22.0 and ANOVA method. In the statistical analysis, both descriptive and analytical methods
were used at the 95% significance threshold (95% CI). The values of coefficient p less than
0.05, respectively lower than 0.01 were considered to be statistically significant.
3. Results
Obesity, especially in the central regions of adipose tissue, is an important risk element for
the onset of type 2 diabetes, which is frequently associated with hypertension through the path
of inter-pathophysiological mechanisms. By measuring the abdominal circumference, both the
visceral distribution of adipose tissue and the cardiovascular risk induced by it are estimated.
Table 1. Descriptive indicators of abdominal circumference (cm)

Standard Standard 95% Median Confidence Range
Sex N Average Min. Max.
deviation error Inferior limit Superior limit
Male 102 118,34 17,75 1,76 114,86 121,83 75 168
Female 148 118,89 22,52 1,85 115,23 122,55 79 172
The analysis of the data regarding this parameter revealed that a small number of patients in
all the studied cases were within the normal limits, thus 84.3% of men and 91.9% of women
had abdominal obesity (AC ≥102 cm M; AC ≥88 cm F), statistically significant percentage
differences (p=0.049) (Table 1) (Fig. 1). It can be seen that the average values of the abdominal
circumference were approximately equal for men and women, having values of 118.34 cm and
118.89 cm respectively.
High blood pressure is considered the most important risk factor for cardiovascular disease,
the relationship between blood pressure and the occurrence of cardiovascular events being
continuous, constant and independent of other predisposing factors [4]. In the study group, it
was found that hypertension was present at the most of the patients, with 82.4% of men and
79.7% of women, differences, which, however, proved to be statistically insignificant (p=0.364)
(Fig. 2). These results are in agreement with those in the literature, which shows a predisposition
of the female sex to increases in tensile values.
184
Fig. 1. Group structure according to the presence / absence of abdominal obesity
Fig. 2. Group structure according to blood pressure level
Hyperglycaemia is commonly associated with atherosclerotic vascular disease [5]. Even in

the absence of glycaemic values that define diabetes, there is a close relationship between blood
glucose levels and cardiovascular risk [6]. Patients with diabetes have increased mortality and
increased vascular morbidity, which reduces their lifespan by approximately 5-15 years
(depending on the age at which the disease was diagnosed) [7] In the present study, an
insignificant difference of hyperglycaemia could be observed by sex, 70.6% of men and 72.3%
of women had blood glucose levels ≥100 mg/dl (p=0.438) (Fig. 3).
Epidemiological aspects. Of the 250 patients initially included in the study, 143 (57.2%) of
them were re-evaluated during the study. The mean age of the entire group of re-evaluated
patients was 67.10±9.90 years (age limits 37-91 years). In terms of gender distribution of the
143 re-evaluated patients, 86 were women (60.1%) and 57 men (39.9%). It was found that in
the studied group, the female sex is more frequently affected, with a ratio of 1.5 to 1
women/men (Fig. 4). This distribution of cases is consistent with data from the literature, which
report a higher prevalence of metabolic syndrome in women.
Cardiometabolic risk increases proportionally with the presence of each constituent within
the metabolic syndrome [8]. Within the studied group, the cardiometabolic risk was
significantly reduced upon re-evaluation (35.13 vs. 24.25; p=0.001) (Fig. 5). This decrease in
cardiometabolic risk is due in large part to both a significant decrease in systolic and diastolic
blood pressure and blood glucose levels.
185
Fig. 3. Group structure according to blood glucose level
Fig. 4. Structure of the lot at re-evaluation
Fig. 5. Initial cardiometabolic risk and re-evaluation
Because there is at least partially an inflammatory process in cardiovascular disease, serum

C-reactive protein modifications have been investigated in the context of atherosclerosis and
subsequent vascular disorders [9]. Thus, in Table 2 we can observe an improvement of the
inflammatory process by the statistically significant decrease of the inflammatory markers
(p=0.001), represented by C-reactive protein, erythrocyte sedimentation rate, and fibrinogen.
Finding the presence of elevated fibrinogen in obesity, diabetes, and cardiovascular diseases
has led to the idea that this is an independent risk factor for increased cardiovascular morbidity
and mortality, and as such should be added to cardiovascular risk factors.
186
Table 2. Evolution of biological markers in patients with metabolic syndrome

Marker Initial Re-evaluation P value for paired T-test
C-reactive protein 2,97±3,59 2,61±3,25 0,001

Erythrocyte
9,81±7,07 8,54±7,06 0,001
sedimentation rate
Fibrinogen 384,78±114,10 358,22±98,60 0,001
In our study, patients with inflammatory syndrome present and treated with Rosuvastatin
had the smallest differences in mean values from baseline and a reassessment, and the decrease
in fibrinogen values was approximately 30 mg/dl regardless of statin type. A similar situation
was also observed with regard to C-reactive protein, where a decrease of approximately 0.3
mg/L was recorded in all evaluated patients, regardless of the type of statin administered (Fig.
6).
Fig. 6. Distribution of the initial and re-evaluation of mean fibrinogen and C-reactive protein values according to
the type of statin administered
4. Discussions
The obesity epidemic has altered the landscape of cardiovascular risk factors. There are no
convincing evidence that overweight/obese patients with excess visceral/ectopic fat
accumulation represent the subgroup of people most likely to be insulin resistant.
Cardiovascular risk is directly associated with glycaemic control, regardless of the presence
or absence of diabetes diagnosis. Obesity affects the quality of life, having social, professional
and personal repercussions, being shown that the weight loss of at least 5% of the initial weight
improves the perception of individuals on their image, influencing the quality of life favourably
[10, 11].
Although the metabolic syndrome increases the relative cardiovascular risk, its diagnosis is
not sufficient for the correct assessment of the overall cardiometabolic risk, which must first be
evaluated using current algorithms based on known risk factors (age, sex, blood pressure, low-
intensity cholesterol lipoproteins, high-intensity cholesterol lipoproteins, diabetes, smoking,
family history of cardiovascular disease) [12]. However, there is evidence that metabolic
syndrome can further increase cardiometabolic risk globally beyond what could be estimated
by traditional risk factors [13]. In the current study, we found that female sex was more
frequently affected, with a 1.5: 1 ratio of women/men, a possible explanation being that women
had greater abdominal circumference and obesity was more common than in men. At re-
evaluation, the cardiometabolic risk was significantly lower, this fact is largely due to the
significant decrease of the blood pressure, as well as the glucose level.
Statins suppress vascular and myocardial inflammation, favourably modulate the vascular
and myocardial structure and improve the bioavailability of nitric oxide. Early initiation of
187
statin treatment in normolipidemic subjects without cardiovascular disease or increased levels

of C-reactive protein has a significant positive impact on cardiovascular risk [14, 15].
5. Conclusions
The metabolic syndrome should be recognized as an entity that needs the attention of the
medical world and public health authorities, having the gift of encouraging the doctor to
evaluate all its components so that the therapeutic approach of patients at risk for cardiovascular
disease and diabetes is as high as possible, judicious and complete. The identification of patients
with cardiometabolic risk as quickly as possible allows the evaluation and therapeutic approach
of the environmental risk factors, offering the chance of diminishing the prevalence of
cardiovascular diseases and type 2 diabetes.
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15. Cannon CP, et al., (2015). Ezetimibe Added to Statin Therapy after Acute Coronary Syndromes. N Engl
J Med 372(25), pp. 2387-2397.
188
Natural Killer Cells in Type 1 Diabetes – Phenotypic

Characteristics
ISVORANU Gheorghiţa1, MANOLE Emilia1, SURCEL Mihaela1,2,

MUNTEANU Adriana Narcisa1,2, NEAGU Monica Teodora1,2
1“Victor Babeș” National Institute of Pathology, Bucharest (ROMANIA)
2Faculty of Biology, University of Bucharest (ROMANIA)
Emails: gina_isvoranu@yahoo.com, emilia_manole@yahoo.com, msurcel2002@yahoo.com, iadinuta@yahoo.com,
neagu.monica@gmail.com
Abstract
Animal models have been used to elucidate the pathophysiology of large variety of diseases
and to provide insight into potential targets for therapy. Treatments for type 1 diabetes are aimed
at resetting the adaptive arm of immune system, the innate immune system being often ignored
in the design of novel immune-based therapies. The objective of this study was to characterize
Natural Killer (NK) cells phenotype in diabetes. A single high-dose treatment with
streptozotocin was used to generate a type-1 diabetic animal model. We found significantly
differences regarding activation and maturation markers on splenic NK cells.
They displayed an activated surface phenotype in the diseased mice.
Keywords: Type 1 diabetes, NK cells, animal model
1. Introduction
Diabetes mellitus (DM) is a chronic metabolic disorder that is characterized by a relative or

absolute lack of insulin, resulting in hyperglycaemia. According World Health Organization
diabetes was the seventh leading cause of death in 2016, and its incidence is constantly
increasing worldwide [1]. Type 1 diabetes (T1D) is an autoimmune disease leading to the
destruction of the insulin-secreting pancreatic β-cell in the islets of Langerhans [2]. T1D have
a multi-factorial pathogenesis, and it is generally believed to be a result of the interaction of the
immune system with both genetic and environmental factors [3]. T1D development and
progression are primarily caused by T effector cells, but recently evidence has linked
abnormalities within innate immunity as potentially having a role in T1D pathogenesis. There
are data that suggesting a participating role of NK cells in the disease development. An altered
NK cell number and function was found both in the peripheral blood and affected tissues of
patients with T1D and in experimental mouse models of T1D [4].
NK cells provide crucial defences against viral pathogens and tumour cells. There are studies
that reported a significant alteration of NK cell functions in cancer [5], [6]. They are innate
lymphocytes, but they also show typical characteristics of the adaptive immune system, such
as expansion and generation of memory cells [7]. Previous studies have demonstrated that NK
cells show a dual behaviour, they can either protect against or promote autoimmunity [8], [9],
[10]. Similarly, in animal models of autoimmune diabetes, NK cells can either be protective or
harmful.
The role for NK cells in autoimmune diseases, especially in diabetes is less studied. For a
better understanding of divergent results on NK cells in diabetes, a more detailed
189
characterization of the phenotype and function of NK cells will be necessary. The aim of this
study was to explore the phenotype of splenic NK cells in a mouse model for type 1 diabetes.
As the main characteristic of T1D is an autoimmune destruction of the pancreatic β-cell,
leading to lack of insulin production, in animal models this deficiency can be achieved by
chemical ablation of the pancreatic β-cell. Streptozotocin (2-deoxy-2-(3-methyl-3-nitrosourea)
1-D-glucopyranose) is widely used to induce experimental type 1 diabetic animal models [11],
[12]. In chemically induced models of type 1 diabetes, a high percentage of the endogenous
beta cells are destroyed, a high single dosage administration induces complete death of
pancreatic β-cells within 2 days, followed by hyperglycaemia and weight loss.
Animals
All animal procedures were approved by the Ethics Committee from Victor Babeș Institute
and by the National Sanitary Veterinary and Food Safety Authority. Procedures were conducted
in accordance with recognized principles of laboratory animal care in the framework of EU
Directive 2010/63/EU for animal experiments. C57BL/6J mice males, 8-10 weeks old and
weighing 20-26 g, were used in this study. Mice were provided by the Animal Husbandry of
Victor Babeș National Institute. Mice were maintained in a specific pathogen-free animal
facility with a temperature-regulated fashion (22±2 °C), humidity 55±10%, and on a 12h
light/dark cycle. Mice had free access to pure water and fed with chow diet.
Type 1 diabetic mouse model

Mice received streptozotocin (STZ, Sigma-Aldrich, St. Louis, MO, USA) by intraperitoneal
injection at a dose of 150mg/kg in freshly prepared Hank’s Balanced Salt Solution (HBSS,
ThermoFisher Scientific, USA). Group control received an injection of HBSS alone. After
streptozotocin injection, mice received 10% sucrose in drinking water for 3 days. On day five
after injection the fast blood glucose level of mice was measured using the blood from the tail
vein and glucose monitor (ACCU-CHEK Performa Nano, Roche Diagnostics, Switzerland).
One week after the streptozotocin injection the animals were sacrificed and the spleens were
harvested and spleen cell suspensions were prepared according ref [13].
NK cells phenotype
Spleen cell suspensions were labelled with the following monoclonal antibodies from
BioLegend, San Diego, CA, USA: FITC anti-mouse CD3ɛ (clone 145-2C11), PE/Cy7 anti-
mouse CD335 (NKp46) (clone 29A1.4), PE/Cy7 anti-mouse CD69 (clone H1.2F3), APC anti-
mouse CD11b (clone M1/70), APC/Cy7 anti-mouse CD43 (clone 1B11), PerCP/Cy5.5 anti-
mouse/rat/human CD27 (clone LG.3A10), PE anti-mouse KLRG1 (clone 2F1), APC/Cy7 anti-
mouse CD25 (IL-2Rα) (clone PC61); and eFluor 450 anti-mouse CD49b (DX5) (clone DX5)
from eBioscience Inc, San Diego, CA, USA. Data acquisition and analysis were performed on
a BD FACSCanto II cytometer with BD FACSDiva v.6.1 software (BD Biosciences, San Jose,
CA, USA).
Statistical analysis
Results were analysed using Student t-test (two-tailed, assuming equal variance) and 𝑃<0.05
was considered significant. Data are presented as mean ±SD (standard deviation).
190
3. Results
On day 5, after a high single dosage of STZ, the mice in the diabetes group presented
hyperglycaemia (˃590 mg/dL) and weight loss. No significant differences were found in the
glycaemia or body weight of the control group, while the body weight of the diabetes group
was lower than that on starting point, day 0. The changes in body weight during the experiment
are displayed in Figure 1.
Fig. 1. Changes in body weight during the experiment. Data were presented as mean ± SD (n=5 per group)
30
25
20
Body weight (g)
15
10
0
day 0 day 5
Control Diabetes
Analysis of NK cells population in spleen cells suspension showed an increasing of the

percentage of CD49b+ (DX5) cells in diabetic mice (10.79±6.12) compared to healthy animals
(5.36±0.86) (Fig. 2).
Fig. 2. Percentage of CD49b cells in diabetes group, and the control group.
The results are presented as percentage from CD3ɛ- lymphocytes (mean ± SD, n=5 per group)
20,00
18,00
16,00
% from CD3- lymphocytes
14,00
12,00
10,00
8,00
6,00
4,00
2,00
0,00
Control Diabetes
In diabetes group splenic NK cells that express CD49b showed a distinct phenotype
compared with NK cells in healthy mice. In diabetes NK cells had a higher expression of the
191
activation markers CD69 (39.96±16.2), CD25 (35.36±10.96) than control, and expressed more
KLRG1 (12.9±1.3), a receptor that is upregulated on NK cells after activation and proliferation
(Fig. 3).
Fig. 3. Expression of CD69, CD25 and KLRG1 for CD49b cells.

The results are presented as percentage from CD49b+lymphocytes (mean ± SD, n=5 per group)
100,00
90,00
80,00
70,00
% from CD49b+ cells
60,00 *
50,00 *
40,00
30,00
20,00 *
10,00
0,00
CD49b+CD69+ CD49b+CD25+ CD49b+ KLRG1+
Control Diabetes
The activating receptor NKp46 (CD335) was downregulated on NK cells in diabetes

(34.6±9.3) (Fig. 4). Also, splenic NK cells had low expression of the CD27 (23.9±5.3), CD11b
(26.8±7.9) and CD43 (61.0±6.2), markers associated with NK cells maturation [4, 14].
Fig. 4. Expression of CD27, CD11b, CD43 and CD335 for CD49b cells.
100,00
90,00
80,00
70,00 *
% from CD49b+ cells
60,00
50,00 *
40,00 *
30,00
20,00
10,00
0,00
CD49b+CD27+ CD49b+CD11b+ CD49b+CD43+ CD49b+CD335+
Control Diabetes
The expression pattern of CD27 and CD11b define four stages of NK cell maturation:
immature NK cells (CD27-CD11b-), early mature NK cells (CD27+CD11b-), mature NK cells
(CD27+CD11b+) and late mature cells (CD27-CD11b+). Analysis of NK cell subsets, defined
by the differential expression of a combination of CD27 and CD11b, indicated a significant
difference in the distribution of NK cell subsets with the immature subset being dominant in
the diabetic mice (Fig. 5).
192
Fig. 5. NK cell subsets defined by the expression level of CD27 and CD11b.
100,00
90,00
80,00
70,00
*
% from CD49b+ cells
60,00
50,00
40,00
30,00 * *
20,00
*
10,00
0,00
CD27-CD11- CD27+CD11b- CD27+CD11b+ CD27-CD11b+
Control Diabetes
Although many details of the complex pathophysiology of type 1 diabetes have been
clarified, the role of NK cells and is still unknown. In this study we present a phenotypic
characterization of the splenic NK cells in a T1D mouse model.
In diabetes group NK cells showed signs of activation, expressing high levels of CD69,
CD25, and KLRG1. The finding reveals that diabetes can induce high activation in secondary
lymph. It is possible that this increase to be a consequence of inflammation. KLRG1 is a
maturation receptor expressed on NK cells that have undergone homeostatic or infection-
induced proliferation, but it is also a molecular signature of exhausted CD8+T cells. High
expression of KLRG1on NK cells may be a consequence of proliferation in response to
inflammation. Cytokines such as IFN-γ, IL-15, IL-12, and IL-18 known as activators of NK
cells have been associated with inflammation in the pancreatic islets [15], [16].
The NKp46 receptor, which recognizes ligands expressed on stressed cells was
downregulated on diabetic NK cells. NKp46 (similar NCR1 or CD335) is part of natural
cytotoxicity receptor group together with NKp44 and NKp30 and is found on all mature NK
cells [17]. Analysis of maturation markers CD27, CD11b and CD43 revealed a tendency to
decrease their expression on NK cells. Distinct differences have been described between
diabetic and healthy mice in the NK cell expression of the maturation markers CD27 and
CD11b. In diabetes group, our data revealed significant increased values for the immature
stages, while the percentages for mature subsets were significant decreased. We have found the
same phenotypic characteristics of splenic NK cells in a cutaneous melanoma mouse model
[13].
4. Conclusions
Streptozotocin-induced type-1 diabetic model was analysed in order to evaluate the

involvement of NK cells in the pathogenesis of this autoimmune disease. Evaluating a large
panel of NK surface markers from the activation and maturation sets we obtained important
differences in experimentally induced mouse NK cell phenotypes as compared to the control
group. Our study has provided new insights into NK cell phenotype in diabetes as future new
approaches to diabetes immunotherapy. Studies that examine the role of NK cells in type 1
diabetes progression, which NK subsets impact the other immune cells, and the characterization
193
of the immune environment following immunomodulation are needed in order to develop novel
immune-based therapies.
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sheets/detail/diabetes
2. Kerner, W., Bruckel, J., German Diabetes Association, (2014). Definition, classification and diagnosis of
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9. Fogel, L.A., Yokoyama, W.M., French, A.R. (2013). Natural killer cells in human autoimmune diseases.
Arthritis Research & Therapy 15, 216, pp. 1-9.
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Zaharescu, I., Sima, L., Costache M., Neagu, M. (2019). Reinforcing involvement of NK cells in
psoriasiform dermatitis animal model. Experimental and Therapeutic Medicine, pp. 4956-4966.
11. Dekel, Y., Glucksam, Y., Elron-Gross, I., Margalit, R. (2009). Insights into modelling streptozotocin-
induced diabetes in ICR mice. Lab Animal 38(2), pp. 55-60.
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resulting from streptozotocin challenge in experimental animals, its practical use and potential risk to
humans. Journal of Diabetes and Metabolic Disorders 12(1), pp. 1-7.
13. Isvoranu, G., Surcel, M., Huica, R.I., Munteanu A.N., Pirvu, I.R., Ciotaru, D., Constantin, C., Bratu, O.,
Neagu, M., Ursaciuc, C. (2019). Natural killer cell monitoring in cutaneous melanoma – new dynamic
biomarker. Oncology Letters 17(5), pp. 4197-4206.
14. Abel, A.M., Yang, C., Thakar, M.S., Malarkannan, S., (2018). Natural Killer Cells: Development,
Maturation, and Clinical Utilization. Frontiers in Immunology 9,1869, pp. 1-23.
15. Brauner, H. Elemans, M., Lemos, S., Broberger, C., Holmberg, D., Flodström-Tullberg, M., Kärre, K.,
Höglund, P. (2010). Distinct phenotype and function of NK cells in the pancreas of nonobese diabetic
mice. The Journal of Immunology 184(5), pp. 2272-2280.
16. Cardozo, A.K., Proost, P., Gysemans, C., Chen, M.C., Mathieu, C., Eizirik, D.L., (2003). IL-1β and IFN-
γ induce the expression of diverse chemokines and IL-15 in human and rat pancreatic islet cells, and in
islets from pre-diabetic NOD mice. Diabetologia 46(2), pp. 255-66.
17. Hadad, U., Thauland, T.J., Martinez, O.M., Butte, M.J., Porgador, A., Krams, S.M. (2015). NKp46
clusters at the immune synapse and regulates NK Cell polarization. Frontiers in Immunology 6:495, pp.
1-11.
194
Risk of Hypoglycemia in Patients with Stage Three of Chronic

Kidney Disease and Type 2 Diabetes Mellitus Treated with
Different Classes of Oral Antidiabetic Drugs
GHERBON Adriana1, FRANDES Mirela2

1 Second Department of Internal Medicine, Clinic of Diabetes, Nutrition and Metabolic Diseases, Emergency Clinical County
Hospital, “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
2 Department of Functional Sciences, “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
Emails: gherbon.adriana@umft.ro, mirela.frandes@umft.ro
Abstract
Chronic kidney disease is a medical condition which can evolve into terminal renal failure,
requiring dialysis. Initially, patients are asymptomatic, so they are underdiagnosed, especially
elderly patients. However, several oral antidiabetic drugs are contraindicated in case of elderly
diabetic patients with chronic kidney disease, the main risk remaining hypoglycaemia.
We aimed to assess the risk of hypoglycaemia in elderly patients with both stage three of
chronic kidney disease and type 2 diabetes mellitus treated with different classes of oral
antidiabetic drugs. The study group included 165 elderly Romanian patients with type 2
diabetes and stages three of chronic kidney disease. Measured data included the patients’
clinical data, anthropometric parameters, and biochemical investigations. The patients treated
only with Biguanide had a shorter duration of the disease, higher body mass index, and better
values of the glycemia and glycated haemoglobin. The patients treated with Sulphonylureas or
with both Biguanide and Sulphonylureas had a rather good glycaemic control with a trend to
hypoglycaemia. Some of the patients from our study group received Glibenclamide, which is
inappropriate for older adults because of the risk of severe and prolonged hypoglycaemia. In
the group treated with both Biguanide and Dipeptidyl Peptidase-4 Inhibitors, the values of
fasting glycemia and glycated haemoglobin were higher than in patients treated with Biguanide
and Sulphonylureas, so the risk of hypoglycaemia was lower. Inappropriate administration or
intensification of therapy (especially for sulfonylureas) represents the main cause of
hypoglycaemia in elderly patients. It is necessary to evaluate the renal function of elderly
patients and to change the oral antidiabetic treatment according to the current guidelines and
stages of chronic kidney disease to slow the progression of renal impairment.
Keywords: oral antidiabetic drugs, elderly, chronic kidney disease, type 2 diabetes mellitus, glomerular filtration rate
1. Introduction
Chronic kidney disease (CKD) is a severe condition that can evolve into terminal renal
failure requiring dialysis. CKD global prevalence varies between 11% and 13%, with a
predominance of stage 3 [1]. The Predator study from Romania showed a CKD prevalence of
7% (1.3 million adults), most of whom were over 60 years old [2]. The leading causes of CKD
are: glomerulonephritis, type 1 and 2 diabetes mellitus (DM), uncontrolled hypertension, renal
polycystic disease, some drugs, such as: acetaminophen and ibuprofen, atherosclerosis, urinary
obstruction by calculus bladder, prostate adenoma, strictures or tumours, vascular pathologies:
arteritis, vasculitis or fibro-muscular dysplasia, HIV infection, amyloidosis, systemic lupus
erythematosus, renal calculi, renal diseases and some tumours [3].
195
Elderly patients have cardiovascular risk factors such as hypertension, DM, or metabolic
syndrome, which together increase the risk of CKD progression [4]. On the other hand, CKD
is considered an independent risk factor for cardiovascular disease [5]. However, several studies
showed that there is an inverse relationship between cardiovascular risk and estimated
glomerular filtration rate (eGFR), regardless of age, sex, or other factors. Most patients die due
to cardiovascular complications than due to the progression to terminal-stage of renal disease
[6]. The cause of CKD in DM patients can be diabetes itself or other conditions (e.g.,
hypertension, glomerulonephritis, prostate adenoma, etc.). DM treatment consists of oral
antidiabetic drugs (OADs) or insulin. Until the last years, insulin was the only one indicated in
the advanced stages of CKD. More recently, with the discovery of new classes of OADs, insulin
is reported in stage five of CKD; in stages three and four being administered some OADs.
Because of the presence of CKD, some OADs may accumulate in the blood, with an
increased risk of hypoglycaemia. Therefore, some ODAs are contraindicated or require dose
reduction.
Generally, elderly patients with diabetic kidney diseases (DKD) are at a higher risk of
hypoglycaemia than patients without CKD, even if the glycaemic control of both groups of
patients is comparable [7]. The decreased GFR affects the clearance of insulin and many
diabetes medications, and therefore it raises the risk of hypoglycaemia. It was shown that
hypoglycaemia is the primary barrier to achieving normal glycemia, which is the goal for
delaying the progression of DKD [8]. Moreover, hypoglycaemia is also related to the
progression of cognitive impairment, more exactly, cognitive impairment increases the
subsequent risk of hypoglycaemia, and at the same time, the presence of severe hypoglycaemia
raises the risk of dementia [9]. The targets of glycaemic control should be tailored to each
patient, taking into account the functional status, comorbidities, especially the presence of
CVD, CKD, and the risk of hypoglycaemia. A patient without DM complications or having
only mild microvascular DM complications, but without other major concurrent illnesses,
should have an HbA1c level of ~7%. A patient with longer-duration of DM (more than ten
years) or with DM complications, needing treatment with insulin, should have an HbA1c level
of ~8%. In the case of a patient with major concurrent illness and advanced microvascular
complications, the target level of HbA1c should be of 8-9% [10]. Our study aimed to evaluate
the risk of hypoglycaemia in the elderly with stage three of chronic kidney disease and type 2
diabetes mellitus treated with different classes of oral antidiabetic drugs.
Study design and patients

This study was a cross-sectional, noninterventional study, including 165 patients with type
2 DM and CKD stages three, aged ≥65 years old. They were consecutive outpatients of the
Emergency Hospital Timisoara from 1 January 2018 to 31 December 2018. All included
patients signed the informed written consent. Our study was approved by the Institutional Ethics
Committee (The Ethics Committee of the Emergency Hospital, Timisoara). The inclusion
criteria consisted of: age ≥65 years, presence of type 2 DM, presence of CKD stages three, and
existence of an OAD treatment; while the exclusion criteria consisted of: presence of type 1
DM or secondary diabetes (pancreatic disease, drugs, endocrine diseases, etc.), CKD at other
stages, and treatment based only diet or insulin.
Clinical, anthropometric, and laboratory data

The methods of investigation included the clinical patients data, e.g., case history, current
status; anthropometric parameters, e.g., height, weight, body mass index (BMI), abdominal
circumference (AC), blood pressure; biochemical parameters, e.g., for glycaemic balance:
196
fasting blood glucose, postprandial glucose, glycated haemoglobin, lipid profile (cholesterol,
triglyceride, HDLc, LDLc), for kidney function: eGFR, albuminuria, serum creatinine,
creatinine/albumin ratio.
Body mass index (BMI) was computed with the following formula: actual weight/height2.
Abdominal circumference was measured with a centimetre and was considered normal
values <80 cm for females and <94 cm for males. Determination of plasma glucose was
performed by enzyme technique with glucose oxidase. We considered normal values for fasting
glucose, values between 70-110 mg%, and for postprandial glycemia, values below 140 mg%.
For the diagnosis of diabetes mellitus, we considered the criteria consisting of values equal
to or greater than 126 mg% for fasting glucose and 200 mg for postprandial glycemia.
Determination of glycated haemoglobin (HbA1c) was done through the DiaStat for
measuring HbA1c reported to the total value of HbA. A normal value was <5.7%. For the
diagnosis of DM, HbA1c was ≥6.5%, and for prediabetes, it was between 5.7%-6.4%.
According to KDIGO classification, based on eGFR, stages of CKD are: stage one (I): if
eGFR ≥90 ml/min/1.73 m2, stage two (II): if eGFR is between 60-89 ml/min/1.73 m2, stage
three a (III-a): if eGFR is between 45-59 ml/min/1.73 m2, stage three b (III-b): if eGFR is
between 30-44 ml/min/1.73 m2, stage four (IV): if eGFR is between 15-29 ml/ min/1.73 m2,
and stage five (V): if the eGFR is <15 ml/min/1.73 m2.11 A series of formulas based on age,
sex, race, and serum creatinine (CKD-EPI, MDRD, etc.) are used to estimate GFR.12 In our
study, the GFR estimation was made applying the CKD-EPI formula. Serum creatinine was
determined by Jaffe kinetics (colorimetric enzymatic method) as normal values at F <1 mg%
and B <1.2 mg%. Urinary creatinine was determined by spectrophotometric method
(colorimetric enzyme), assuming normal values of 600-2000 mg/day. Albuminuria was
determined by the immunoturbidimetric method, considering the following interpretation: A1
<30 mg/day: normal or slightly increased albuminuria, A2: 30-300 mg/day: moderate
albuminuria, A3 >300 mg/day: severe albuminuria. We performed the ratio of urinary albumin
to urinary creatinine, with the following interpretation: A1 <30 mg/g: normal, A2: 30-300 mg/g:
moderate, A3 >300 mg/g: severe.
Statistical Analysis
Continuous variables with a Gaussian distribution were described as mean (± standard
deviation), while continuous variables without Gaussian distribution and nominal variables
were expressed by their median (interquartile range) and absolute frequency (percentage),
respectively. The normality of continuous variable distributions was tested using the
Kolmogorov-Smirnov’s test, while the equality of variances was tested using Levene’s test.
The significance of the difference between groups was assessed by applying the Student’s t-
test (means, Gaussian populations), Mann-Whitney U test (medians, non-Gaussian
populations), and Pearson Chi-square test or Fisher’s exact test (proportions). Data were
analysed by using the SPSS v.20 software (SPSS Inc., Chicago, IL, USA). We considered a P-
value <0.05 as the threshold for statistical significance, and a confidence level of 0.95 for
estimating intervals.
3. Results
Our study group included 165 patients with an actual age of 72.96±6.4 years, 62.05% were
females and 37.95% males, who had a DM duration of 10.00 (7.00-12.50) years. The glycaemic
balance expressed by median glycemia level and HbA1c were 133.00 (109.00-164.50) mg%
and 6.70 (6.20-7.45) %, respectively. The characteristics of the studied group are presented in
Table 1.
197
Table 1. Characteristics of the study group

Parameters Values
Age (years)a 72.96±6.4
DM duration (years)b 10.00 (7.00-12.50)
Sex (Male)c 63 (38.19%)
Cholesterol (mg%)b 187.50 (165.00-216.00)
Triglyceride (mg%)b 150.50 (109.00-199.00)
BMI (kg/m2)b 30.50 (28.00-35.00)
Glycemia (mg%)b 133.00 (109.00-164.50)
HbA1c (%)b 6.70 (6.20-7.45)
Albuminuriac 70 (42.42%)
Uric acid (mg/dL)b 6.00 (5.00-7.00)
GFR (mg/min/1.73 m2)b 56.00 (52.00-58.00)
Hypertension 102 (61.81%)
Angina pectorisc 51 (30.9%)
Notes: acontinuous variables (with gaussian distribution) are presented by their mean (SD). bcontinuous variables
(with non-gaussian distribution) are presented by median (IQR). ccategorical variables are presented by absolute
frequency (percentage) in the sample.
Abbreviations: DM, diabetes mellitus; BMI, body mass index; GFR, glomerular filtration rate.
The ethiology of CKD in patients with DM was either diabetes or other diseases. In our study
group, diabetes represented 42.42% of cases, followed by hypertension (29.7%),
arteriosclerosis (5.45%), renal diseases (12.73%) and prostate adenoma (9.7%). The patients
were treated with different classes of OADs as follows, 24.24% of patients received Biguanide,
20.60% Sulphonylureas, 39.39% Biguanide and Sulphonylureas, and 15.77% Biguanide and
Dipeptidyl peptidase-4 inhibitors. A percentage of 79.39% received Biguanide (Metformin),
60% received Sulphonylureas (37.37% Glimepiride, 43.34% Gliclazide, 5.05% Gliquidone and
14.24% Glibenclamide), and 15.77% received Dipeptidyl peptidase-4 inhibitors (96.15%
Sitagliptin and 3.84% Saxagliptin). When considering of the CKD stages, we observed that
92.12% of the patients were at stage III-a of CKD, from which 24.34% received Biguanide,
20.39% Sulphonylureas, 38.82% Biguanide and Sulphonylureas, and 16.45% Biguanide and
Dipeptidyl peptidase-4 inhibitors. Also, we observed that 7.88% of the patients were at stage
III-b of CKD, from which 23.08% received Biguanide, 23.08% Sulphonylureas, 46.15%
Biguanide and Sulphonylureas, and 7.7% Biguanide and Dipeptidyl peptidase-4 inhibitors.
Considering the oral antidiabetic treatment, we presented the characteristics of the study
group in Table 2.
Table 2. Characteristics of the study group stratified by used OADs

Biguanide and
Biguanide and Dipeptidyl p-
Parameters Biguanide Sulphonylureas
Sulphonylureas peptidase-4 valued
inhibitors
Agea 72.3±5.95 75.14±7.08 72.62±6.4 72.03±5.87 0.214
DM durationb 9.00 (7.00-10.00) 9.00 (7.00-10.00) 10.00 (6.00-14.00) 13.00 (10.50-13.50) 0.010
Sex (Male)c 14 (35%) 9 (26.47%) 24 (36.92%) 16 (61.54%) 0.042
Cholesterola 187.02±38.14 192.83±38.74 194.72±53.58 169.65±59.49 0.160
142.00 136.00 156.50 164.00
Triglycerideb 0.140
(101.00-166.00) (109.00-221.00) (118.00-222.50) (108.00-196.00)
BMIa 33.15±5.91 29.46±5.52 30.52±5.02 31.08±4.78 0.062
122.00 132.00 128.50 182.00
Glycemia b
0.010
(103.00-150.00) (116.00-164.00) (106.50-162.00) (136.00-261.00)
HbA1ca 6.34±1.14 6.53±0.65 7.01±1.34 7.4±1.19 0.001
Albuminuriac 10 (14.3%) 12 (17.14%) 31 (44.28%) 17 (24.28%) <0.001
54.00 56.00 55.00 56.00
GFRb 0.779
(51.00-57.00) (53.50-57.00) (52.00-58.00) (53.50-58.00)
198
Hypertensionc 22 (13.33%) 22 (13.33%) 40 (24.24%) 18 (10.9%) <0.001

Angina
13 (7.88%) 10 (6.06%) 20 (12.12%) 8 (4.84%) <0.001
pectorisc
Notes: acontinuous variables (with Gaussian distribution) are presented by their mean (SD). bcontinuous variables
(with non-gaussian distribution) are presented by their median (IQR). ccategorical variables are presented by
absolute frequency (percentage) in the sample. dcontinuous variables (with non-gaussian distribution) are
compared using the Kruskal-Wallis H test; categorical variables are compared using the Chi-square test or the
Fisher’s exact test.
Abbreviations: DM, diabetes mellitus; BMI, body mass index; HbA1c, haemoglobin A1c; GFR, glomerular
filtration rate.
We observed that the DM duration was the longest in case of patients treated with Biguanide
and Dipeptidyl peptidase-4 inhibitors, 13.00 (10.50-13.50) years; on the second place was the
class of patients treated with Biguanide and Sulphonylureas, 10.00 (6.00-14.00), and afterward,
there were the groups treated separately with Sulphonylureas and Biguanide.
When comparing all the four classes, we observed significant differences between the DM
duration (Kruskal-Wallis H test X2(3)=4.479, p=0.010) (Table 2). In addition, when comparing
every two classes of OADs, we observed significant differences only between the group of
patients treated with Biguanide and the group treated with Biguanide and Sulphonylureas
(Mann-Whitney U test, U=1,665.00, p=0.016), and between the group of patients treated with
Biguanide and the group treated with Biguanide and Dipeptidyl peptidase-4 inhibitors (Mann-
Whitney U test, U=757.50, p=0.002) (Table 3).
Table 3. Comparison between the studied sub-groups divided by used OADs

OADs-1 OADs-1 OADs-1 OADs-2 OADs-2 OADs-3
Parameters vs. vs. vs. vs. vs. vs.
OADs-2 OADs-3 OADs-4 OADs-3 OADs-4 OADs-4
Agea 0.068 0.794 0.861 0.086 0.068 0.678
DM duration b
0.121 0.016 0.002 0.052 0.056 0.922
Sex (Male)c 0.263 0.840 0.002 0.158 < 0.001 0.002
Cholesterola 0.519 0.393 0.208 0.841 0.102 0.078
Triglycerideb 0.807 0.102 0.468 0.130 0.42 0.559
BMIa 0.017 0.021 0.175 0.399 0.276 0.655
Glycemiab 0.073 0.117 0.001 0.692 0.042 0.022
HbA1ca 0.427 0.009 0.001 0.024 0.002 0.189
Albuminuriac 0.511 <0.001 0.001 <0.001 0.140 <0.001
RFGb 0.490 0.580 0.525 0.819 0.964 0.861
Hypertensionc 0.897 <0.001 0.318 <0.001 0.318 <0.001
Angina pectorisc 0.314 0.036 0.083 0.002 0.462 <0.001
Notes: acontinuous variables (with gaussian distribution) are compared using t-Student test.
b
continuous variables (with non-gaussian distribution) are compared using Mann-Whitney U test. ccategorical
variables are compared using Chi-square test or Fisher’s exact test.
Abbreviations: OADs-1, oral antidiabetic drugs including Biguanide; OADs-2, oral antidiabetic drugs including
Sulphonylureas; OADs-3, oral antidiabetic drugs including Biguanide and Sulphonylureas; OADs-4, oral
antidiabetic drugs including Biguanide and Dipeptidyl peptidase-4 inhibitors; DM, diabetes mellitus; BMI, body
mass index; HbA1c, haemoglobin A1c; GFR, glomerular filtration rate.
We obtained significance differences regarding BMI – higher in patients treated with

Biguanides vs. patients treated with Sulphonylureas, and with Biguanides and Sulphonylureas.
More precisely, we observed significant differences between the group treated with
Biguanide and the group treated Sulphonylureas (Mann-Whitney U test, U=260.50, p=0.017),
and between the group of patients treated with Biguanide and the group treated with Biguanide
and Sulphonylureas (Mann-Whitney U test, U=542.50, p=0.021) (Table 3). Glycemia was
higher in patients treated with Biguanides + Dipeptidyl Peptidase-4 Inhibitors vs. patients from
the other three groups, e.g., the group of patients treated with OADs-1 (Mann-Whitney U test,
199
U=763.50, p=0.001). HbA1c was lower in patients treated with OADs-1 vs. patients treated
with OADs-2, and also significantly lower in patients treated OADs-1 vs. patients treated with
OADs-3 (Mann-Whitney U test, U=1,328.50, p=0.009).
Moreover, HbA1c was significantly lower in patients treated OADs-1 vs. patients treated
with OADs-4 (Mann-Whitney U test, U=651.50, p=0.001).
4. Discussion
According to the current guidelines, Metformin is the first therapeutic line for type 2 DM
patients [10]. In our study, most of the patients received Metformin (79.39%) according to the
actual guidelines [11] and some studies [12]. In case of renal insufficiency at stage III-b,
Metformin is not initiated and can be continued at a maximum dose of only 1 g/day, is
contraindicated in stage four or five of CKD [13]. In elderly patients, there are still discussions
about the use of Metformin [14]. Some studies showed the benefit of other newer drugs, such
as DPP-4 inhibitors [15]. The main risk of Metformin administration in elderly patients with
CKD is lactic acidosis, being necessary to monitor the renal function.
In our study, the patients treated only with Metformin had a shorter duration of the disease,
higher BMI, and better values of both glycemia and HbA1c. The Metformin dose was
unchanged in stage III-a of CKD, being reduced at 1 g/day in CKD stage III-b. The beneficial
effects of Metformin therapy included low oscillations of body weight and decreased the risk
of hypoglycaemia [16]. Metformin also has some other benefits, e.g., improving cardiovascular
prognosis in overweight or obese patients with newly diagnosed type 2 DM, favourable effects
in hepatic steatosis (commonly present in patients with DM), possible protection against some
forms of cancer, positive influence on intestinal microbial flora [17].
Sulphonylureas have been for a long time the only non-insulin drugs used to treat type 2
DM. The most critical side effects are hypoglycaemia, with all degrees of severity, especially
in the elderly or in case of renal failure, and weight gain. Sulfonylurea-induced hypoglycaemia
has the particular feature that it is prolonged and causes frequent relapses after apparent
correction under treatment [18]. In renal impairment, Glibenclamide is contraindicated, and
Gliclazide, Glipizide, and Gliquidone may, however, be administrated with caution and with
dose adjustment because their elimination is done in a lower quantity by the kidneys. Over 65
years, it is also a contraindication for this medication (especially for Glibenclamide) [19].
In our study, 60% of patients received Sulphonylureas (37.37% Glimepiride, 43.34%
Gliclazide, 5.05% Gliquidone, and 14.24% Glibenclamide). The Sulphonylureas were
administrated such as second therapeutic line when the Biguanide is contraindicated or non-
tolerated. Some of the patients from our study group received Glibenclamide, which is
inappropriate for older adults because of the risk of severe and prolonged hypoglycaemia.
Also, in CKD, these drugs are contraindicated, and they were partially eliminated [20]. The
consequence is accumulation into the blood with increased risk of hypoglycaemia in older
patients with CKD. The patients treated with Sulphonylureas or with Biguanide and
Sulphonylureas had a better glycaemic control expressed through HbA1c values, with a trend
to hypoglycaemia.
In our study, 15.77% receive DPP-4 inhibitors (96.15% Sitagliptin and 3.84% Saxagliptin).
The values of fasting glycemia and HbA1c were higher than in patients treated with
Biguanide and Sulphonylureas, so the risk of hypoglycaemia was lower. SGLT-2 inhibitors
(Gliflozins) are the newest class of non-insulin antidiabetics. Its representatives are
Dapagliflozin, Empagliflozin (registered in Romania so far), Canagliflozin, etc. The
mechanism of action is the inhibition of glucose reabsorption in the proximal convoluted tube
[21]. Contraindications are chronic kidney disease at 3-5 stages, age <18 years, pregnancy and
lactation [22]. The use of Dapagliflozin is not recommended in patients with moderate to severe
200
renal impairment. Renal function monitoring is recommended before initiating Dapagliflozin

treatment and then at least once a year.
5. Conclusion
Inappropriate administration or intensification of therapy (especially for sulfonylureas)

represents the main cause of hypoglycaemia in elderly patients. It is necessary to evaluate the
renal function of elderly patients and to change the oral antidiabetic treatment according to the
current guidelines and chronic kidney disease stages to slow the progression of renal
impairment.
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202
The Impact of Metabolic Syndrome on Pregnancy
POPESCU (BANACU) Ina1,2, PACU Irina1,2, IONESCU Cringu Antoniu1,2,

MĂDAN Victor2,3, SOCEA Bogdan2,4, DIMITRIU Mihai1,2,
BACALBASA Nicolae2, POPESCU Mihail6, ZYGOUROPOULOS Nikolaos1,
DAN Adelina1, BANACU Mihail1,2
3 Department of Urology, “Dr. Carol Davila” Central Military Emergency University Hospital, Bucharest (ROMANIA)
4 Department of General Surgery, “Sf. Pantelimon” Clinical Emergency Hospital, Bucharest (ROMANIA)
5 Municipal Hospital, Ramnicul Sarat (ROMANIA)
6 Elias Emergency University Hospital, Orthopaedics and Traumatology Department, Mărăști Blvd, no 17, Bucharest
(ROMANIA)
Emails: ina.popescu@yahoo.com, irinapacu@hotmail.com, antoniuginec@yahoo.com, victmad@gmail.com,
bogdansocea@gmail.com, drmihaidimitriu@yahoo.com, nicolae_bacalbasa@yahoo.com, nz@zitaquality.ro,
danadelina90@gmail.com, mbanacu@gmail.com
Abstract
Introduction
The metabolic syndrome (MetS) is a constellation of conditions such as insulin resistance,
hyperinsulinemia, abdominal obesity, impaired glucose tolerance, dyslipidaemia, and
hypertension. Each of these conditions, and, more so, their sum, has a tremendous impact on
the pregnancy development and its outcomes.
Methods
This integrative review was written after reviewing the databases for studies on this matter
published during the last two decades.
Discussion
During the last decade, the researchers who were studying pregnant women found a
correlation between the development of preeclampsia and the subsequent risk of developing the
MetS after delivery. They concluded that the number of MetSs features increases the risk of
developing preeclampsia, placental dysfunction and gestational diabetes.
Conclusion
There are still unanswered questions as to the way these risk factors work. Whether they
work in an additive or multiplicative manner remains an open question.
Keywords: metabolic syndrome, pregnancy, preeclampsia
Introduction
The metabolic syndrome (MetS) is a constellation of conditions such as insulin resistance,

hyperinsulinemia, abdominal obesity, impaired glucose tolerance, dyslipidaemia, and
hypertension. Each of these conditions, and, more so, their sum has a tremendous impact on the
pregnancy development and its outcomes. Taking into consideration the fact that the metabolic
syndrome becomes a global epidemic, we think it is becoming more and more important to
diagnose it either at the prenatal consult or, at least, during the first trimester of the pregnancy.
203
What to expect during pregnancy in case of patients with metabolic syndrome, which are the
mechanism leading to these outcomes and how to treat a patient with metabolic syndrome who
is also pregnant are the main subjects of this article.
Methods
This integrative review was written after reviewing the databases for studies on this matter
published during the last two decades.
Discussions
To have this discussion, we need to postulate the definition of metabolic syndrome (MetS).
Even though there are several definitions of the syndrome, the main traits are similar.
Taking into consideration the fact that the most used definition is the one issued by the
National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III) in 2005
we will proceed by listing the criteria. To diagnose the metabolic syndrome, one should meet 3
out of 5 following criteria [1]:
1. Abdominal obesity, defined as a waist circumference ≥102 cm in men and ≥88 cm in
women
2. Serum triglycerides ≥150 mg/dL (1.7 mmol/L) or drug treatment for elevated
triglycerides
3. Serum high-density lipoprotein (HDL) cholesterol <40 mg/dL (1 mmol/L) in men and
<50 mg/dL (1.3 mmol/L) in women or drug treatment for low HDL cholesterol
4. Blood pressure ≥130/85 mmHg or drug treatment for elevated blood pressure
5. Fasting plasma glucose (FPG) ≥100 mg/dL (5.6 mmol/L) or drug treatment for elevated
blood glucose [2].
Since the pregnancy can sometimes mimic the changes seen in MetS by showing an increase
in blood glucose level, in triglyceride level and possibly in blood pressure [3], we find it
important to establish the diagnosis before the pregnancy as there haven’t been established
modified criteria applicable for this estate.
Regarding the patients outside the pregnancy, the risks associated with MetS are relatively
well described. The literature describes the risk of developing type 2 diabetes, cardiovascular
disease (CVD) [4], fatty liver disease [5], chronic kidney disease [6], polycystic ovary
syndrome [7], sleep apnea and other sleep-disordered breathing [8-11].
During the last decade, the researchers who were studying pregnant women found a
correlation between the development of preeclampsia and the subsequent risk of developing the
MetS after delivery [12]. The downside of the study is the fact that they did not evaluate the
presence of MetS before the pregnancy; thus, even though the correlation was present, the
causality direction was unclear. Since then, a significant number of studies researched the
association between the pre-pregnancy presence of various MetSs components (obesity,
hypertension, dyslipidaemia, insulin resistance or hyperglycaemia) and the development of
preeclampsia [13]. They also concluded that the number of MetSs features increases the risk of
developing preeclampsia. One study discovered a strong correlation between the number of
MetSs features present before the pregnancy and the growing threat of developing placental
dysfunction [14]. The same study concluded that the greater the number of features, the higher
the risk of developing fetal growth discrepancies or intrauterine fetal death in association with
placental dysfunction.
A recent study researched the MetS as a syndrome rather than its components and its impact
on pregnancy. It concluded that women with MetS have an increased risk of developing
preeclampsia, with features like raised blood pressure, elevated triglycerides and high waist
204
circumference as key contributors to the poor outcome [15]. It also states that patients who
suffer from MetS are at risk of developing gestational diabetes with components as raised
triglycerides, raised blood pressure and raised glucose contributing as individual factors [15].
The researchers excluded MetS as a particular risk factor, outside of its distinctive traits, for
small for gestational age (SGA), large for gestational age (LGA) or spontaneous premature
birth. We note the importance of this study while acknowledging one of its limitations – the
fact that the diagnosis of MetS was established at 15 weeks of pregnancy rather than before the
pregnancy using the criteria for adult patients thus emphasizing the lack of criteria tailored for
pregnancy.
As to the pathophysiological processes that lead to the poor pregnancy outcomes of patients
with MetS it appears that the metabolic stress, the excessive inflammatory response and the
endothelial dysfunctions are ultimately involved [13]. Both endothelial dysfunction and
excessive inflammatory response are traits of MetS, causing the common cardiovascular disease
[16, 17] while being known causes of placental dysfunction leading to preeclampsia.
In conclusion, regarding the potential pregnancy outcomes, the chronic “dysmetabolism”
present before the pregnancy and during the beginning of it, seems to be at fault for the
abnormal remodelling of the spiral arteries and its’ results [14].
The management strategies for metabolic syndromes, such as dietary and lifestyle
interventions, lower the risk of long-term CVD but are also known to reduce the risk of
preeclampsia [18]. Targeting modifiable factors such as excessive weight gain through physical
activity and dietary advice, focusing on the reduction of calories, dietary glycaemic load and
saturated fats [19] lower the risk for gestational diabetes [20]. Lifestyle interventions also bring
long-term benefits since women who have diabetes, arterial hypertension or obesity are known
to be at risk for multiple cancer types [21-23].
Conclusion
In the end, we find ourselves asking the same questions as other practitioners – Do the MetS
imply different pathophysiology, or does it confer any additional risk as a syndrome besides the
risks conferred by its traits? Is the cluster of features stronger than the sum of its components
regarding the dangers it poses? Are the risk factors working in an additive manner or preferably
in a multiplicative manner? These questions remain unanswered until further studies.
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206
Serum Adipokines in Obese Bariatric Patients – Correlations with

Metabolic Parameters
HRISTOV Ioana1, TIMOFTE Daniel Vasile2,

CREȚU-SILIVESTRU Iustina Silvia1, MIHAI Bogdan3, ANISIE Ecaterina4,
MOCANU Veronica1
1 Department of Morpho-Functional Sciences II, “Grigore T. Popa” University of Medicine and Pharmacy, Iasi (ROMANIA)
2 Department of Surgical Sciences, “Grigore T. Popa” University of Medicine and Pharmacy, Iasi (ROMANIA)
3 Department of Diabetes and Nutrition, “Grigore T. Popa” University of Medicine and Pharmacy, Iasi (ROMANIA)
4 Laboratory of Immunology, “Sf. Spiridon” Hospital, Iasi (ROMANIA)
Email: ioanahristov@yahoo.com
Abstract
Introduction
There has been growing evidence of significantly reduced serum concentrations of
adiponectin in obese individuals compared to those with normal body mass index (BMI).
Hypoadiponectinemia is associated with the development of insulin resistance and type 2
diabetes mellitus, independently of BMI and metabolic syndrome. Leptin/Adiponectin ratio is
considered a more precise instrument for cardio-metabolic risk evaluation in obese patients,
compared to adipokines value separately.
Methods
Our clinical study evaluated serum adipokines in a group of morbidly obese patients (N=106)
with a mean BMI= 45.0±6.3 kg/m2 (36.4-58.9) before bariatric surgery procedure.
We assessed the relationship between serum leptin, adiponectin and leptin/adiponectin ratio
values, morning blood glucose and insulin levels, lipid profile, morning cortisol and thyroid
hormones profile in a group of obese bariatric patients with metabolic syndrome (N=64) versus
“metabolic healthy” obese patients (N=42) according to IDF criteria.
Results
The plasma adiponectin values varied between 8,68-33,84 µg/dl, with a mean value of
29.0±3.2 ug/dl, with similar results in “metabolic healthy obese” patients vs subgroup with
associated metabolic syndrome (29.76 vs 28.45 µg/dl; p=0.120). Mean leptin serum level was
0.93±0.3 ng/dl and varied between 1.54-44.15 ng/dl in the studied group of obese patients and
there was no statistical difference between the two obese patients’ sub-groups. We found a
positive correlation between Leptin levels and CT/HDL values (r=+0.492; p=0.046).
Leptin/Adiponectin ratio (LAR) had higher values in obese patients with associated
metabolic syndrome 0.877 vs 1.045; p=0.05, compared to the “metabolic healthy obese sub-
group”. In our study, the correlation between adiponectin and HOMA-IR was indirect and
reduced in intensity, and by drawing the ROC curve, it is noted that the leptin/adiponectin ratio,
with a probability of 65.8%, can be a good predictor in the determinism of the metabolic
syndrome (AUC=0.658; 95% CI: 0.437-0.699).
Conclusion
Low adiponectin levels in obese patients are associated with the development of insulin
resistance and type 2 diabetes, and the Leptin/Adiponectin Ratio was shown to have higher
207
values in patients with metabolic syndrome. LAR was demonstrated in our study to be a
valuable marker for metabolic syndrome in obese patients and can be useful for the evaluation
of the associated cardiovascular risk in these patients.
Funding
Research relating to this abstract was funded by “Grigore T. Popa” University of Medicine
and Pharmacy Iasi, Romania, through the grants Ideas-Teams contract number
29032/28.12.2016 and number 30340/28.12.2017.
Keywords: obesity, adipokines, metabolic syndrome
Background
The obese populations are reportedly at high risk for cardio-metabolic diseases. As metabolic
syndrome is a significant predictor, not only for cardiovascular mortality but also for all-cause
mortality. The evaluation of the metabolic status of obese patients needs to be performed
precociously and accurately [1]. The present definition of the International Diabetes Federation
(IDF) for the metabolic syndrome [2] includes the assets of the cardiovascular risk factors that
are considered essential predictors for cardiovascular mortality for obese and non-obese
patients and also contains parameters that can be easily accessible for screening [3]. Insulin
resistance evaluation by HOMA-IR is considered as an excellent cardiovascular risk predictor
[4], is also described as a valuable criterion for recognition of the obese individuals with higher
mortality risk [1].
Insulin resistance is well known associated with disturbances of fat metabolism [5]. The
rising storage capacity of the subcutaneous adipose tissues results in lipotoxicity. This condition
is characterized by fatty acid infiltration of insulin target tissues, i.e., skeletal muscle and liver,
that could lead to insulin resistance [5].
High leptin levels and leptin- adiponectin ratio are predictors of obesity-related
complications as type 2 diabetes mellitus (T2DM) and hypertension independent of BMI or the
metabolic syndrome (MetS) criteria [6-8].
Adipose tissue is an essential source of circulating leptin and adiponectin, especially the
subcutaneous tissue, is responsible for the secretion of large amounts of leptin, with its
expansion increasing mRNA expression for leptin [9-10]. Leptin also has an impact on food
appetite, energy consumption, adipogenesis and insulin sensitivity. The broad spectrum of
biological action of this hormone is, however, closely related to the functions of adipose tissue
and maintenance of body weight [11]. The neurohormonal mechanism, which acts at the
hypothalamic level, regulating the balance between food intake and energy consumption, is
initiated by coupling at this level of leptin with the specific receptor, causing a decrease in the
secretion of neuropeptide Y and α-MSH (melanocytes stimulating hormone), which will lead
to a reduction of appetite and an increase in the energy consumption rate through the generation
of caloric energy and lipolysis. This process is mediated by the sympathetic nerve fibers,
stimulated by leptin, which acts on the adipocytes via epinephrine receptors on their surface,
stimulating lipolysis.
Characteristics of our study group: The study included 106 obese patients (OB group),
referred for Laparoscopic Sleeve Gastrectomy (LSG) that were characterized using IDF criteria
for metabolic syndrome in 2 subgroups: obese patients with associated metabolic syndrome-
Metabolic Unhealthy Obese (MUHO) (N=64) and metabolic healthy obese (MHO) (N=42)
208
The mean body mass index (BMI) was 45.02±6.31 kg/m2. The age ranges of the two groups
matched, with a mean of 42.05±9.91 years for the MUHO group and 42.00±10.67 years for the
MHO group. Biochemical measurements: Blood samples were collected after 12h of fast,
before the bariatric surgery procedure. The lipid profile was established based on the following
serum determinations: Fasting plasma glucose (FPG), Total Cholesterol (TC), Low-Density
Lipoproteins (LDL) Cholesterol, High-Density Lipoproteins (HDL) Cholesterol, Triglycerides
(TG) and TC/HDL ratio. Plasma concentrations were measured using
electrochemiluminescence immunoassay method. HOMA-IR was calculated using the formula:
FPI x FPG /405, where FPI = fasting plasma insulin (μUI/ml) and FPG = fasting plasma glucose
(mg/dl). Leptin and adiponectin were measured in plasma using ELISA method (Sigma Aldrich
kit RAB0005 was used for adiponectin and Fine Test EH0216 for leptin serum concentration
quantification.
Statistical analysis: Data was analysed using IBM SPSS Statistics 21 Software and expressed
as mean ± standard deviation. Comparisons between parameters in obese patients and normo-
ponderal controls were performed using the Mann-Whitney test. Pearson’s correlation analysis
was used.
To evaluate correlations between lipid accumulation evaluation and biochemical and
hormonal parameters. Significance was defined as p<0.05.
Results
The plasma adiponectin values varied between 8.68-33.84 µg/dl, with a mean value of
29.0±3.2 ug/dl, with similar results in MHO patient’s vs MUHO subgroup (29.76±2.55 vs
28.45±4.86 µg/dl; p=0.120). Mean Leptin serum concentration was 0.93±0.3 ng/dl and varied
between 1.54-44.15 ng/dl in the studied group of obese patients and there was no statistical
difference between the two obese patients’ sub-groups. (Table 1). Leptin/Adiponectin ratio had
higher rates in MUHO compared to MHO bariatric patients (1.045±0.353 vs. 0.8777±0.308,
p=0.05). For the lipid profile evaluation, results show lower LDL cholesterol levels in the MHO
subgroup and higher HDL cholesterol, as this is associated with a reduced atherogenic risk for
this phenotype of obese patients. For the thyroid hormones and morning cortisol levels, there
were no statistically significant differences between the two obese phenotypes subgroups.
Table 1. Biochemical and endocrine profile evaluation in the two sub-groups of obese patients
Parameters MUHO MHO (n=42) p-value
(n=64)
Age (Years) 43.21±9.95 40.31±11.23 0.143
Female n(%) 43 (67.6%) 38 (89.8%) 0.004
BMI, kg/m2 43.02±8.37 38.68±9.85 0.012
Blood glucose, mg/dl 123.57±47.24 91.72±11.40 0.001
Insulinemia, µUI/ml 26.04±18.36 16.47±9.74 0.001
HOMA-IR 9.40±7.00 4.43±3.72 0.001
Total Cholesterol, mg/dl 223.23±41.26 202.31±32.12 0.006
LDLc, mg/dl 148.62±32.73 136.36±29.65 0.050
HDLc, mg/dl 43.16±8.58 50.41±9.56 0.001
Triglycerides, mg/dl 208.20±99.33 106.06±38.74 0.001
TGO, UI/l 26.52±15.58 21.88±7.38 0.069
TGP, UI/l 40.09±26.83 28.87±15.07 0.014
CT/HDL 5.30±1.20 4.14±0.78 0.001
TSH, µUI/ml 2.60±2.44 3.09±2.53 0.324
FT4, pmol/l 1.04±0.15 1.09±0.15 0.147
FT3, pmol/l 4.34±1.03 4.70±1.10 0.134
Cortisol, µg/dl 12.64±3.94 11.20±3.92 0.061
Adiponectin, µg/dl 29.76±2.55 28.45±4.86 0.120
Leptin, mg/dl 26.54±9.30 27.08±10.84 0.811
LAR 0.8777±0.308 1.045±0.353 0.05*
209
Correlation with insulin-resistance and metabolic syndrome:

We found a positive correlation between Leptin levels and CT/HDL values (r=+0,492;
p=0,046). Leptin/Adiponectin ratio (LAR) had higher values in obese patients with associated
metabolic syndrome 0.877 vs 1.045; p=0.05, compared with the “metabolic healthy obese sub-
group”. In our study, the correlation between adiponectin and HOMA-IR was indirect and
reduced in intensity (Fig. 1).
40
35
30
Adiponectina
25
20
15
10
0
0 50 100
HOMA-IR
Fig. 1. The sensitivity/specificity balance of the leptin/adiponectin ratio in the determinism
of the metabolic syndrome
By drawing the ROC curve, it is noted that the leptin/adiponectin ratio, with a probability of
65.8%, can be a good predictor in the determinism of the metabolic syndrome (AUC=0.658;
95% CI: 0.437-0.699) (Fig. 2).
ROC Curve
1,0
0,8
Sensitivity
0,6
0,4
0,2
0,0
0,0 0,2 0,4 0,6 0,8 1,0
1 - Specificity
Fig. 2. ROC curve for LAR as a predictor for the metabolic syndrome in the study group of obese patients
Discussions
There has been growing evidence of critical reduced levels of adiponectin in obese patients
compared to subjects with normal body mass index (BMI) [10]. An inverse relationship with
BMI was observed in both men and women, as well as a negative correlation of adiponectin
with visceral fat accumulation. It was demonstrated that hypoadiponectinemia (levels less than
4 mg/l) is associated with the increasing level of insulin resistance and development type 2
210
diabetes mellitus, independently of BMI and metabolic syndrome. The lower level of
adiponectinemia is considered an independent risk factor for developing hypertension. Kern et
al., measured adiponectin plasma concentrations and mRNA levels in adipose tissue in
nondiabetic individuals with varying degree of obesity and insulin resistance [12]. They found
a strong correlation between these two parameters. Obese individuals had significantly lower
plasma level of adiponectin. When BMI was less than 30 kg/m2, women had twice more the
body fat than men, but adiponectin levels were higher on average of 65% than in men (14.2
mg/l vs.8.6 mg/l). Peoples with the highest levels of mRNA secreted the lowest levels of TNF
alpha in adipose tissue [13].
Since the early 2000s, adiponectin has begun to be considered a valuable biomarker for
metabolic pathology, with an inverse correlation with both visceral adiposity and basal blood
glucose, triglycerides, insulinemia or blood pressure [14]. Thus, results similar to those obtained
in our study were published by Kotani et al., [15] and showed higher levels of LAR in patients
with metabolic syndrome or Ayina et al., [16], which demonstrated statistically significant
correlations between LAR and metabolic syndrome or HOMA-IR. In our study, the correlation
between adiponectin and HOMA-IR was indirect and reduced in intensity.
Still, given the small group of patients included in the study, it can be estimated that for a
larger group of obese patients, this relationship should be considered. Although the validation
of LAR as a marker of insulin resistance still requires extensive studies, it could be a useful
predictor, which does not require fasting for the risk of type 2 diabetes, early warning of obesity-
associated adipocyte dysfunction, which precedes metabolic disorders. Thus, results similar to
those obtained in our study were published by Kotani et al., [15], which showed higher levels
of LAR in patients with metabolic syndrome or Ayina et al., [16], which demonstrated
statistically significant correlations between LAR and metabolic syndrome or HOMA-IR. In
our study, the correlation between adiponectin and HOMA-IR was indirect and reduced in
intensity. Still, considering the small group of patients included in the study, it can be estimated
that for a larger group of obese patients, this relationship should be maintained and maintained.
It obtains statistical significance.
For the Leptin/Adiponectin Serum (LAR) ratio, although slightly lower values were detected
in patients with metabolic syndrome without obtaining statistical significance, however, by
tracing the ROC curve, it is noted that LAR can be a good predictor of the syndrome metabolic.
Regarding the association of metabolic syndrome in obese patients with adipokine secretion,
there are data that associate increased levels of leptin with abdominal type obesity and increased
risk for metabolic pathology and peripheral insulin resistance, respectively [17, 18]. Serum
adiponectin concentration has constant individual values but varies significantly inter-
individually depending on genetic factors, body weight, sex and general health status.
Leptin and adiponectin are adipokines secreted exclusively by adipocytes. Although certain
aspects related to the regulation of their secretion and synthesis remain incompletely explored,
the value of leptin is directly correlated with fat mass. In contrast, the secretion of adiponectin
decreases with weight gain.
Hypoadiponectinemia represents an independent risk factor of BMI and metabolic syndrome
for the development of insulin resistance, type 2 diabetes, but also for high blood pressure,
being associated with visceral adiposity. Low serum adiponectin levels are considered an
independent risk factor for high blood pressure, as shown by the study by Kim et al., who
analysed both serum adiponectin levels and mRNA adiponectin expression in adipocytes and
demonstrated that obese patients had significantly lower levels of adiponectin and for the
female subgroup, 65% higher levels of adiponectin were found, although body composition
showed a double amount of adipose tissue compared to men [19].
Data analysis in our study did not detect statistically significant differences for leptin and
adiponectin between obese patients with metabolic syndrome and those without metabolic
211
syndrome but confirmed for the serum leptin level a strong positive correlation with BMI, the
dependence of the serum fat adipokine concentration being described in the literature [20].
In our study, we found significantly lower mean values of leptin/adiponectin ratio in patients
with metabolic syndrome, and by tracing the ROC curve, it is noted that adiponectin may be a
good predictor for metabolic syndrome. The results obtained represent an additional argument
regarding the close relationship between the serum level of leptin and adiponectin and in
particular the relationship between these two adipokines with the metabolic syndrome elements
and thereby with the cardiovascular risk of obese patients.
Conclusion
The leptin/adiponectin ratio for our group of patients was statistically significantly higher in
obese patients with metabolic syndrome compared to those without associated metabolic
pathology criteria. At the same time, this leptin/adiponectin ratio is a good predictor for the
metabolic syndrome in our study. For the groups of obese patients, the data in the literature are
not yet very numerous in terms of this correlation, which suggests that further studies are needed
to establish the particularities of the mechanisms of metabolic dysfunction for obese patients.
Progress in deciphering the pathophysiological mechanisms of the metabolic syndrome
associated with obesity brings to the fore the adipose tissue and the particularities related to the
expansion, the storage capacity of the fatty acids resulting from the positive energy balance but
also disturbances in the endocrine activity of the adipose tissue.
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213
Insulin Resistance and Male Infertility
PACU Irina1,2, GHEORGHIU Diana, MĂDAN Victor3, CIOBANU Ana Maria1,

DAN Adelina-Loredana1,2, ZYGOUROPOULOS Nikolaos1, MATEI Alexandra1,2,
ROȘU George-Alexandru1,2, PACU Ovidiu3, NAVOLAN Dan4,
CĂLIN Florin Daniel1,2
3 Department of Urology, “Dr Carol Davila” Central Military Emergency Hospital, Bucharest (ROMANIA)
Emails: irinapacu@hotmail.com, cdgheorghiu@hotmail.com, victmad@gmail.com, danadelina90@gmail.com,

nz@zitaquality.ro, ale.matei@gmail.com, george.rosu@ymail.com, navolan@yahoo.com, dkcalin@gmail.com
Abstract
Metabolic syndrome is particularly prevalent and multifaceted, causing significant

disturbance of numerous physiological processes. Connections between obesity, dyslipidaemia,
hypertension and insulin resistance are each examined concerning their associated detrimental
effects on male fertility. Insulin resistance may induce a prejudicial impact on male infertility
due to hyperinsulinemia and being part of metabolic syndrome. In our study, we aimed to look
at the intercourse among insulin resistance and men infertility.
Keywords: insulin resistance, male infertility, metabolic syndrome, diabetes mellitus
Introduction
Insulin resistance, dyslipidaemia, hypertension and obesity are components that signal the
presence of metabolic syndrome, initially described by a committee of experts from the World
Health Organization (WHO) in 1998 [1]. All these conditions determine the perturbation of
physiologic processes [2]. The increase in the prevalence of this syndrome has coincided with
a decrease in birth rates and fertility potential [3]. Male fertility can be affected by the presence
of metabolic syndrome through several mechanisms. Obesity, dyslipidaemia, hypertension, and
insulin resistance may separately influence male fertility in different ways, but when these
conditions coexist, there is a cumulative effect on male fertility. This review will investigate
the relationship between insulin resistance and men infertility.
Epidemiology
It is known that, after one year of regular unprotected intercourse, 15% of couples who want
to conceive are affected by infertility [2]. Men determine up to 40% of infertility issues, 40%
by women only and up to 20% are estimated to be male and female infertility problems [4]. 5-
8% of obese premenopausal women have polycystic ovarian syndrome characterized by chronic
anovulation, amenorrhea or oligomenorrhea, hyperandrogenism, polycystic ovary morphology
in pelvic ultrasound, hyperinsulinemia, and insulin resistance [5, 6].
The causes of male infertility are distributed into four principal categories:
1. Endocrine and systemic abnormalities (frequently with hypogonadotropic
hypogonadism) – 2-5%.
214
2. Primary testicular defects in spermatogenesis – 65-80%, of which the majority have

idiopathic dysspermatogenesis (obscure molecular mechanisms were supporting the
weaknesses [7, 8]).
3. Sperm transport abnormalities – 5%.
4. Idiopathic male infertility – 10-20%. Idiopathic male infertility should be
differentiated from idiopathic dysspermatogenesis (whereas, idiopathic male
infertility is characterized by average seminal fluid analysis and no aetiology for
infertility, in the idiopathic dysspermatogenesis the analysis of seminal fluid is
abnormal) [8].
One theory is that insulin resistance can determine hypogonadism and other metabolic
disorders and cause male infertility [9].
Pathophysiology
Insulin resistance represents the low sensitivity of cells to stimulation by insulin in normal
or elevated levels of glucose. Therefore, the pancreas will produce more insulin, causing
hyperinsulinemia, leading to type 2 diabetes mellitus [10].
Oxidative stress, vascular endothelial dysfunction, an abnormal lipid profile, hypertension,
and vascular inflammation are consequences of insulin resistance, the associated
hyperinsulinemia and hyperglycaemia and adipocyte cytokines (adipokines-leptin, adiponectin,
tumour necrosis factor-alpha and resisting [11-15]), all of these leading to atherosclerotic
cardiovascular disease [16-18]. For preventing cardiovascular events, healthy dietary patterns
must be adopted [19]. Insulin resistance is the primary deficiency in the metabolic syndrome
[20, 21] involved in the pathogenesis of type 2 diabetes mellitus.
Hyperinsulinemia caused by resistin-induced insulin resistance affects normal
spermatogenesis through its inhibitory effect [22, 23]. A study shows nuclear and mitochondrial
DNA damage in the sperm of the patients with diabetes mellitus [24] deteriorating male fertility.
Outcomes of Male Reproductive Health
Erectile and ejaculatory dysfunction and low libido, endothelial dysfunction of the corpus
cavernosum caused by neurovascular modifications leading to retrograde ejaculation are often
encountered in patients with insulin resistance [3].
Increased insulin resistance was correlated to decreased testosterone release by the Leydig
cells without alterations in hypothalamic or pituitary function [24, 25].
Lower concentration, total count, motility and ejaculate volumes of the sperm were
confirmed in men with diabetes mellitus [26-28]. Also, there is no association with pregnancy
morbidity in adolescent or with postpartum pain in adolescent [29-31].
Preliminary results of a study highlighted that there is no opposite correlation between
insulin resistance and the male reproductive system; additional studies were necessary [32, 33].
Conclusion
Long-term outcomes of insulin resistance include the appearance of type 2 diabetes mellitus
and cardiovascular disease; thus, a healthy lifestyle represents the most important interference
to prevent unwished effects. In men with infertility, insulin resistance may inflict
hypogonadism, idiopathic oligozoospermia and other metabolic disorders, but further studies
are required to improve the understanding of the underlying pathophysiology and to discover
new therapeutic interventions.
215
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02-04, 2017 Sponsor(s): Assoc Renal Metab & Nutrit Studies; AstraZeneca Diabetes; MSD Diabetes;
novo nordisk; SANOFI INTERDIAB 2017: DIABETES MELLITUS IN INTERNAL MEDICINE
Book Series: International Conference on Interdisciplinary Management of Diabetes Mellitus and its
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217
Oral Mucosa Vascularization in Early Diabetic Diagnosis
VYSOCHANSKAYA Julia1, YEROSHENKO Galina2, RYBALOV Oleg2

1Senior Scientist, PhD Odontologia Somatica LTD (RUSSIAN FEDERATION)
2Ukrainian Medical Stomatological Academy (UKRAINE)
Email: zlider@inbox.ru
Abstract
Long term diabetes may cause microangiopathy. The mucous membrane of the oral cavity
is characterized by a developed network of microvessels. The smallest vessels that are the first
to respond to the pathology of glucose uptake are located in the macula, fingertips and in the
oral mucosa. The oral mucosa is interesting for the study and early detection of
microangiopathy, as it undergoes daily damage by the food lump and has various anatomical
and physiological areas for a comparative analysis of the ability to self-repair in a small study
area. The aim of our work was to study the mucous membranes micro circulatory of the hard
palate and alveolar processes in patients with and without diabetes mellitus, in age groups from
9 to 80 years. To determine the zonal characteristics of the microvessels of the hard palate
mucous membrane, a morphometric study of histological sections of the mucosa from10 people
of 40 and 80 years old was carried out, the cause of death of which was not associated with
pathology of the oral cavity and cardiovascular system. Using the rheography method, the
following was studied: 1) a group of 30 children aged 9 to 18 years without pathology of the
oral cavity and cardiovascular system for the zone study of microcirculation of the hard palate
– control; 2) a group of 30 people aged 15-25 who do not have pathology from the
cardiovascular system and oral cavity for a comparative assessment of the state of
microcirculation of the mucous membrane of the vestibular, palatine and lingual sides of the
upper and lower jaws; 3) a group of 100 patients aged 15 to 20 years with diabetes mellitus.
Our study showed, that the rheographic index from diabetic patients sharply decreases, the
elasticity index rises (which indicates vasodilation), the peripheral tone of arterioles rises and
the peripheral tone of venules decreases. The filtration rate increases sharply, which indicates
a decrease in the quality of trophic processes and a possible increase in pressure.
Keywords: vascularization, diabetic, oral, mucosa, diagnosis
Introduction
Long term diabetes may cause microangiopathy. In this case, high blood glucose levels cause
the endothelial cells lining the blood vessels to take in more glucose than normal. The presence
of insulin receptors on endothelial cells is well documented [1]. The insulin‐stimulated
production of nitric oxide is a feature of the action of insulin on endothelial cells.
They then form more glycoproteins on their surface than normal, and also cause the
basement membrane in the vessel wall to grow abnormally thicker and weaker. Therefore they
bleed, leak protein, and slow the flow of blood through the body [2].
The role of the dentist in the diagnosis of somatic pathology is growing every year. The oral
cavity is well visualized, participates in many physiological processes, carries an active immune
and digestive function. Somatic changes occur in the oral cavity in the form of metabolic and
neurovascular pathologies [3]. The mucous membrane of the oral cavity is characterized by a
developed network of microvessels. The smallest vessels that are the first to respond to the
218
pathology of glucose uptake are located in the macula, fingertips and in the oral mucosa. The
oral mucosa is interesting for the study and early detection of microangiopathy, as it undergoes
daily damage by the food lump and has various anatomical and physiological areas for a
comparative analysis of the ability to self-repair in a small study area. By the functional state
of this link of the cardiovascular system, one can determine the state of the organ and predict
the progression of somatic pathology [4]. The mucous membrane of the oral cavity is well
studied, but meanwhile there is not enough work on zonal microcirculation of the hard palate
in the norm and with pathology in the age aspect [5].
The aim of our work was to study the mucous membranes micro circulatory of the hard
palate and alveolar processes in patients with and without diabetes mellitus, in age groups from
9 to 80 years.
Morphology:
To determine the zonal characteristics of the microvessels of the mucous membrane of the
hard palate, a morphometric study of histological sections of the mucosa of 10 people of 40 and
80 years old was carried out. The cause of death of which was not associated with pathology of
the oral cavity and cardiovascular system. The material of the mucous membrane of the oral
cavity was poured into Epon-812 according to the generally accepted method [7], after which
half-thin sections were stained with hematoxylin-eosin and examined under a light microscope.
Using the MOV-16 eyepiece micrometer, the diameter of microvessels was studied. Based
on the made two-dimensional reconstructions corresponding to the zones of the hard palate, the
topography of the microvessels and the type of their branching were determined [9, 10].
Rheography of the microvascular function:

To study the physical parameters of the vascular link, the method of zonal registration of the
potentials of the vessels of the hard palate was used [6] attaching a two-channel rheograph RPG-
2-02 and specially adapted lead sensors with a diameter of 0.5 cm, readings were taken in the
second standard lead. The essence of the method consists in applying two electrodes to the
mucous membrane by a gauze pad folded in half and moistened with saline. The electrodes are
fixed using iron clothespins, coated with rubber and located far from 0.5 cm from each other.
The characteristic of the rheogram differs from the generally accepted one in that “insisura”
in our case characterizes the metabolic processes in the capillaries, therefore we called it
“capillary”. The peripheral resistance index and dicrotic index are not determined, but a new
indicator is introduced – the capillary filtration rate. A rheogram can be evaluated qualitatively
(Table 1) and quantitatively. When a rheovasogram is quantified, the amplitude-time indices
are estimated by measuring the distance between certain points on the reovasogram and their
formula processing, it is possible to calculate the exact digital indices for the microvasculature:
i. Elasticity Index (IE) – displays the degree of elasticity of the vascular wall, it
increases with vasodilation. IE=(a/s) x100%, where c is the amplitude of slow blood
supply.
ii. The Rheographic Index (RI) is an indicator of the intensity of blood supply and the
degree of expansion of the vessel wall RI=(w/h) x0.1 Ohm where in is the length of
the anacrotic wave, h is the height of the dicrotic wave, 0.1 Ohm is the calibration
index.
iii. The Rheographic Index (RI) is an indicator of the intensity of blood supply and the
degree of expansion of the vessel wall RI=(w/h) x0.1 Ohm where in is the length of
the anacrotic wave, h is the height of the dicrotic wave, 0.1 Ohm is the calibration
index.
219
iv. Venulus tone index (PTSv) — indicates the state of the arteriole wall tone
PTSv=B/(a+B) x100%.
v. Filtration rate (Vf) – displays the degree of metabolic processes in a given area,
indirectly shows the trophic state of the test object Vf=fx0.032, where f is the height
of incisura corresponding to the degree of filtration, 0.032 (s) is the division price
of the calibration scale, depending on the settings of the rheograph [8].
Table 1. Qualitative assessment of the tone and functional state of regional vessels
Pulse Curve assessment Vascular tone The functional state of
the vessels
Anacrota steep Normal Not changed
(ascending part) Sharp steep Reduced Dilatation
sloping Elevated Constriction
Sharp Normal Normal
peak Pointy Reduced Dilatation
Flattened Elevated Constriction
The location of dicrota In the middle third Normal Normal
In the lower third Reduced Dilatation
In the upper third Elevated Constriction
The severity of dicrota Distinctly Normal Normal
Very sharp Reduced Dilatation
Weak Elevated Constriction
Using this method, the following was studied: 1) a group of 30 children aged 9 to 18 years
without pathology of the oral cavity and cardiovascular system for the zone study of
microcirculation of the hard palate – control; 2) a group of 30 people aged 15-25 who do not
have pathology from the cardiovascular system and oral cavity for a comparative assessment
of the state of microcirculation of the mucous membrane of the vestibular, palatine and lingual
sides of the upper and lower jaws; 3) a group of 100 patients aged 15 to 20 years with diabetes
mellitus. All patients and parents signed the informed consent form.
The statistical analyses were realized using SPSS ® version 20.0 software. A p-value of 0.05
was counted statistically significant. We used mean ± standard deviation to describe continuous
variables with a normal distribution and median for skewed variables. The variables were tested
for normal distribution and the group comparison; we used paired Student’s t-test, chi-square
or Mann-Whitney U test depending on their Gaussian distribution.
Microcirculatory:
The type of branching of the bringing vessels of the suture zone looks like a crest, the
branches depart almost at a right angle. The delivery vessel is placed superficially and gives off
twigs that run parallel to the epithelial layer. From the rest, the capillary leaves in each
connective tissue papilla at an angle of 90 degrees (Table 2).
The branching of the microvessels of the marginal zone corresponds to the tangential type.
The angle of departure of arterioles is 120-130 degrees.
In the fatty and glandular zones, the type of branching of arterial vessels can be described as
loose.
220
Table 2. The microvasculature reconstruction of the hard palate mucosa
The suture zone The marginal zone
The fatty zone The glandular zone
The vessels are most densely located in the marginal zone. The venous link according to the
type of branching corresponds to zonal features. The highest density of this link corresponds to
the fat zone. The following patterns were revealed by the depth of location:
- in the suture area, the main vessels are placed superficially, this applies to both venules
and arterioles. Given the smallest thickness of the epithelial layer in this region, even
superficial damage will be accompanied by damage to large vessels. A corresponding
defect in other areas will correspond to capillary bleeding. Therefore, the massiveness
will be less, and the duration will be longer because in other areas of the hard palate the
depth of the vessels is almost twice as great as in the seam zone.
- there is a clear distribution of arterial and venous cells in the marginal and suture zones,
while in the fat and glandular zones they are scattered.
- estimating the diameters of microvessels, we can say that the largest arterioles (101.25
microns) and capillaries (20.75 microns) are in the glandular zone, and venules (51
microns) are in the fatty zone.
- The smallest indicators characteristic of the marginal zone: 16.9 microns – arterioles,
26.7 microns – venules and 10.7 microns – capillaries.
It is interesting that with age, the smallest diameters are characteristic of the fat zone in all
respects, the largest are for the suture zone, while the narrowing of the microvessels in the
glandular zone is determined and the microvessels of the edge zone remain almost unchanged.
This indicates high regenerative abilities of the marginal zone [3].
The Rheography data can be presented in (Table 3).

1. The maximum anacrota is observed in the marginal and glandular zones, but the
glandular zone has the steepest rise and a pointed apex, which indicates the highest
arteriole resistance in this area. The top of the anacrota in the fat zone is presented in
the form of a plateau (that is, the smallest tone of the venous network).
2. The depth of the capillary notch is directly proportional to the exchange processes of
the capillaries: the descending part – filtration into the pre-vascular space, the ascending
part – characterizes re-absorption. The weld zone has the greatest filtration ability and
the least readsorption (normally, the readsorption amplitude is inversely proportional to
filtration). The fat readsorption zone is characterized by the greatest readsorption ability,
which is explained by functional features.
221
It can be assumed that with pathology will be observed:

- an increase in the filtration amplitude, which indicates an increase in its intensity and
exudative inflammation
- if the base of the capillary notch has a sharp base, but forms a plateau, there is an
infectious process, as this is a manifestation of a decrease in the tone and dilatation of
capillaries
- if there is a hump instead of a tenderloin, this indicates ischemia
3. The greatest steepness of dicrota is observed in the marginal and glandular zones, this
should be understood as the largest volume of blood flow. In the fat zone, the venous
curve is uniform, has no elevations, which indicates the same vascular tone along the
entire length, so the outflow in this zone is stable, without drops. The fastest evacuation
of blood occurs in the marginal zone.
4. The blood supply index is ascending from the seam zone to the fat zone, which is due
to the function of these zones.
5. The greatest elasticity of blood vessels is observed in the seam zone, and resistance is
the smallest.
6. The greatest tone of arterioles can be determined in the glandular zone, and venules in
the fatty zone, which also corresponds to the functional features of these zones.
Table 3. The hard palate microcirculation – healthy people (n=60)

The suture zone The marginal zone The fatty zone The glandular zone
14.28-15.13 17.1-18.0 18.57-19.98 20.0-21.2 RI (1)
74-76% 69-71% 61-63% 58-60% IE (2)
40-44% 39-42% 37-40% 43-46% PTSa (3)
56-59% 57-61% 74-79% 54-57% PTSb (4)
0.14-0.16 0.08-0.10 0.04-0.06 0.10-0.12 Vf (mm/s) (5)
With diabetes, the following picture is observed.

The rheographic index sharply decreases, the elasticity index rises (which indicates
vasodilation), the peripheral tone of arterioles rises and the peripheral tone of venules decreases.
The filtration rate increases sharply, which indicates a decrease in the quality of trophic
processes and a possible increase in pressure (Table 4).
Table 4. The state of the microvascular bed of the mucosa of the hard palate in patients with diabetes mellitus
The suture zone The marginal zone The fatty zone The glandular zone
RVG arm
RVG hard
palate
14.28-15.16 17.1-18.2 18.57-19.98 20.0-21.2

1
74-76% 69-71% 61-63% 58-60%

2
40-44% 39-42% 37-40% 43-46%

(p≤0,05)
Normal
5 4 3
56-59% 57-61% 74-79% 54-57%

0.24-0.36 mm/s 0.20-0.29 mm/s 0.12-0.18 mm/s 0.16-0.22 mm/s
222
RVG arm
RVG hard
palate
6,78-8,93 ↓↓ 3,16-5,12 ↓↓↓ 3,5-7,9 ↓ 6,2-8,7↓↓↓
5 4 3 2 1
57-66% ↓ 93-113%↑↑ 78-84% ↑ 61-75% ↑
46-52% ↑ 52-57% ↑ 41-46% ↑ 39-43%↓
Diabetes
(p≤0,05)
60-67% ↓ 43-46% ↓ 53-57%↓ 57-59%↓
0,60-0,64 mm/s ↑ 0,45-0,48mm/s ↑ 0,34-0,37mm/s ↑ 0,39-0,43mm/s↑
Conclusion
Our study showed persistent changes in the circulatory bed of microvasculature of the
mucous membrane of the hard palate in patients with diabetes mellitus.
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223
Clinical and Pathological Characteristics of Patients with Type 2

Diabetes and Early Stage Breast Cancer – a Single Centre
Experience
MAZILU Laura1, SUCEVEANU Adrian-Paul1, PAREPA Irinel-Raluca1,

GHEORGHE Andreea Daniela1, PANTEA STOIAN Anca2,
HAINAROSIE Razvan2, STEFANESCU Dragos Cristian2, NITIPIR Cornelia2,
ARDELEANU Valeriu3,4,5, SUCEVEANU Andra Iulia1
1 Faculty of Medicine, “Ovidius” University, Constanta (ROMANIA)
3 Arestetic Clinic, Galati (ROMANIA)
4 Faculty of Medicine, “Ovidius” University, Constanța (ROMANIA)
5 Department of Surgery, General Hospital CFR, Galati (ROMANIA)
Email: dr.ancastoian@gmail.com
Abstract
Breast cancer represents the leading type of cancer in women and the second cause of death
related to cancer worldwide. Diabetes and obesity had a rapid increase in terms of incidence
over the last 30 years. Obesity and type 2 diabetes represent independent risk factors for
development of breast cancer, diabetic women having a 15-20% increased risk for breast
cancer. This study it is an observational study, in a single centre, and was conducted in
Oncology Department of Clinical Emergency Hospital of Constanta, on a group of 226 patients
with early stage breast cancer with or without diabetes, between 2017 and 2019. A signed
informed consent form was obtaining from all participants before collection of data for study,
even if it’s not an interventional study. Statistical analysis was performed using the Statistical
Package for the Social Sciences version 21 software (SPSS). Total number of studied patients
accounted for 226 female patients with early-stage breast cancer, and of these 38 patients,
representing 16.8%, had associated type 2 diabetes. Mean age was 51.77 years for non-diabetic
patients, comparing with 62.82 years for diabetic patients. No significant difference was found
between groups for tumour size, number of involved lymph nodes, histology of breast cancer,
hormonal receptor status, and HER2 status. Regarding tumour size, our observation is that
diabetic patients tend to have larger tumours, 71% of all diabetic patients presented with breast
tumours larger than 21 mm. Regarding the histological grade, clinically significant difference
was observed for grade 2 and 3 tumours. In our analysis, in diabetic group, a significant number
of patients were classified as luminal A (57.9%), followed by luminal B subtype (23.7%), triple
negative breast cancer accounting for 13.2% of the patients, and HER2 enriched subtype for
5.3% of patients. At the time of this analysis all patients were alive, some had disease
progression in both groups, but without any clinically significant difference (p=0.771).
Keywords: breast cancer, type 2diabetes, molecular subtypes
Introduction
Breast cancer represents the leading type of cancer in women and the second cause of death
related to cancer worldwide [1]. Diabetes and obesity had a rapid increase in terms of incidence
over the last 30 years. Obesity and type 2 diabetes represent independent risk factors for
224
development of breast cancer, diabetic women having a 15-20% increased risk for breast cancer
[2, 3]. Studies evaluating body weight and risk of breast cancer, showed that increased body
weight is associated with and increased risk for premenopausal triple negative breast cancer
(TNBC) and luminal B breast cancer [4, 5]. Diabetic women have an increased risk for triple
negative breast cancer, comparing with non-diabetic women [6]. Type 2 diabetes associates
increased risk for development of breast cancer in postmenopausal population [7]. Usually,
breast cancer in diabetic women is diagnosed in advanced stages and mortality is higher in
patients with diabetes and breast cancer, diabetes having a direct influence on breast cancer
prognosis, due to insulin-resistance status, and indirectly influencing breast cancer prognosis
due to its complication [6, 8, 9].
Another possible reason for poor prognosis in patients with breast cancer and diabetes may
be related to the fact that this patient tends to present tumour subtypes that are more aggressive,
and non-responsive to treatment [10, 11].
There are few studies evaluating breast cancer subtypes and characteristic in diabetic
population, despite that breast cancer subtype have been studied in numerous trials.
The objective of this analysis is to evaluate clinical and pathological features of patients with
early-stage breast cancer and diabetes.
Methodology
This study is an observational study, in a single centre, and was conducted in Oncology
Department of Clinical Emergency Hospital of Constanta, on a group of 226 patients with early
stage breast cancer with or without diabetes, between 2017 and 2019. A signed informed
consent form was obtaining from all participants before collection of data for study, even if it's
not an interventional study. All information about the patients were obtain using medical
records of the Department and medical charts of patients – age, menopausal state,
histopathological and immunohistochemistry results, treatment, diabetes status.
Statistical analyses were performed using the Statistical Package for the Social Sciences
version 21 software (SPSS). All statistical test performed were 2-sided and p value <0.05 was
considered statistically significant.
Results
We studied a total number of 226 female patients with early-stage breast cancer diagnosis.
Of these, 38 patients had associated type 2 diabetes, representing 16.8% of patients included
in study. Patients were divided into two groups and comparison between groups was performed
according to the presence or absence of type 2 diabetes. Characteristics and comparisons of the
two groups are described in Table 1.
Table 1. Characteristics of patients with breast cancer
Characteristic Diabetes – Diabetes + P-value Analysis
(n=188) (n=38) between groups
Age 51.77 62.82 p=0.003
(31-75) (48-79)
Menopausal state
Pre-Menopausal 92 (48.9%) 4 (10.5%)
Post-Menopausal 96 (51.1%) 34 (89.5%) p=0.001
BMI
Pre-Menopausal p<0.05
<25kg/m2 85 (45.21%) 12 (31.57%)
25-30kg/m2 68 (36.17%) 15 (39.47%
>30kg/m2 35 (18.61%) 19 (50.05%)
BMI
225
Post-Menopausal
<25kg/m2 103 (57.78%) 9 (23.68%)
25-30kg/m2 55 (25.25%) 14 (36.84%) p<0.05
>30kg/m2 30 (15.95%) 15 (39.47%)
Breast Tumor Size
≤20 mm 62 (33.0%) 11 (28.9%)
21-50 mm 94 (50.0% 23 (60.5%) p=0.914
>50 mm 32 (17.0%) 4 (10.5%)
Number of positive lymph nodes
1-3
≥4 85 (45.2%) 23 (60.5%) p=0.085
103 (54.8%) 21 (39.5%)
Histology
Ductal 163 (86.7%) 35 (92.1%) p=0.319
Lobular 14 (7.4%) 3 (7.9%)
Mixed 11 (5.9%) 0
Grade
Grade 1 21 (11.2%) 6 (15.8%) p=0.002
Grade 2 62 (33.0%) 22 (57.9%)
Grade 3 103 (55.9%) 10 (26.3%)
Oestrogen
Positive 123 (67.2%) 31 (81.6%) p=0.080
Negative 60 (32.8%) 7 (18.4%)
Progesterone
Positive 121 (66.5%) 27 (71.1%) p=0.587
Negative 61 (33.5%) 11 (28.9%)
HER2
Positive 48 (25.5%) 6 (16.2%) p=0.226
Negative 140 (74.5%) 31 (83.8%)
Ki67 (median) 28.30 23.07 p=0.044
Classification – subtype
Luminal A 72 (38.3) 22 (57.9%) p=0.042
Luminal B 67 (5.6) 9 (23.7%)
TNBC 30 (16.0) 5 (13.2%)
HER2 enriched 19 (10.1%) 2 (5.3%)
Surgery
Conservatory 68 (36.2%) 14 (36.8%) p=0.938
Radical 120 (63.8%) 24 (63.2%)
Radiotherapy
Yes 151 (80.3%) 22 (57.9%) p=0.013
No 30 (16.6%) 14 (36.8%)
Unknown 7 (3.7%) 2 (5.3%)
Hormonotherapy
Yes 135 (71.8%) 28 (73.7%) p=0.812
No 44 (23.4%) 9 (23.7%)
Unknown 9 (4.8%) 1 (2.6%)
Disease Progression
Yes 64 (34.0%) 12 (31.6%) p=0.771
No 124 (66.0%) 26 (68.4%)
Median age of the entire group of patients was 52.93 years, and was significantly higher in
diabetic patients (62.82 years) group, comparing with non-diabetics (51.77 years), with a
statistically significant difference (p=0.003). According to menopausal status of the patients,
the majority of patients with breast cancer and diabetes were postmenopausal, representing
89.5%. In non-diabetic group, the proportion of patients in pre- and postmenopausal status was
almost equivalent (48.9% versus 51.1%). Body mass index was calculated for all included
patients, according to data available at the time of first presentation in our clinic, and was
significantly higher in patients with diabetes, 39.47% of diabetic patients were obese comparing
226
with 15.95% in the non-diabetic group. Size of the tumour was similar in the two groups. Most
of the patients in both groups presented tumours between 21-50 mm. In terms of the number of
involved lymph nodes, most of the patients in diabetic group had 1 to 3 lymph nodes affected,
and in the non-diabetic group, a larger number of patients had 4 or more lymph nodes affected.
Majority of patients in both groups had ductal carcinoma, 92.1% in diabetic group, and
86.7% in non-diabetic group. According to subtype classification of breast cancer, we found
that luminal A subtype was the most frequent type in both groups, 57.9% in diabetic group and
38.3% in non-diabetic patients, followed by luminal B subtype 23.7% versus 35.6%, triple-
negative breast cancer 13.2% versus 16.0%, and HER2 enriched accounting for 5.3% in diabetic
group versus 10.1% in non-diabetic group. All patients were evaluated for progression, disease
free survival and overall survival. At the time of this analysis, all patients were alive, and
progressive disease was found in 31.6% of diabetic patients and 34.0% of non-diabetic patients.
Discussions
According to epidemiological data, breast cancer and diabetes represent important health
problems, breast cancer being the second cause of cancer related death in women and obesity
and diabetes incidence are increasing worldwide, in part due to adoption of “Western” way of
life by developing countries [12-15].
Age it is another risk factor for cancer in general, patient over 65 years old are more
susceptible to develop cancer [16-19]. In our study mean age was 51.77 years for non-diabetic
patients, comparing with 62.82 years for diabetic patients. Results are comparable with data
from literature, showing that maximum incidence reaches highest level in menopause, after that
incidence decreases slowly or remains stable [20]. Association between obesity, diabetes and
breast cancer is supported by a multitude of epidemiological studies [21, 22]. Body mass index
was calculated for all patients included in this analysis, according to data available at the time
of first presentation in our clinic, and was significantly higher in patients with diabetes,
regardless of menopausal status. BMI play an important role in survival of patients with breast
cancer as an independent predictor factor [23, 24, 25]. No significant difference was found
between groups for tumour size, number of involved lymph nodes and histology of breast
cancer. Regarding tumour size, even if no clinical significant value of p was obtain in this
analysis, our observation is that diabetic patients tend to have larger tumours, 71% of all
diabetic patients presented with breast tumours larger than 21 mm, and this result is important
due to the fact that size of the tumour influence survival of the patients[26]. Regarding the
histological grade of the tumour there was no difference for grade 1 tumours, but clinically
significant difference was observed for grade 2 and 3 tumours. In diabetic group majority of
patients had moderate differentiated tumours, while non-diabetic patients in our analysis
presented with poorly differentiated tumours. No difference was observed in terms of hormonal
receptor status and HER2 status. In terms of subtype classification, a number of studies showed
that patients with diabetes tend to develop more frequent triple negative breast cancer [6, 27].
In our analysis, in diabetic group a significant number of patients were classified as luminal
A (57.9%), followed by luminal B subtype (23.7%), triple negative breast cancer accounted for
13.2% of the patients, and HER2 enriched subtype for 5.3% of patients. Comparing our data to
literature data, with the exception of luminal A percent, all data was comparable with data
reported in other analysis.
All patients were evaluated for disease progression, disease free survival, and overall
survival.[28] At the time of this analysis all patients were alive, some had disease progression
in both groups, but without any clinically significant difference (p=0.771).
227
Conclusion
Obesity and type 2 diabetes are very well-known risk factors for a multitude of cancers,
including breast cancer. Pre-existing diabetes can impact quality of life, prognosis, treatment
options and mortality of patients with breast cancer.
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(2015). Overweight, obesity, and postmenopausal invasive breast cancer risk: a secondary analysis of
the Women’s Health Initiative randomized clinical trials. JAMA Oncol 1(5), pp. 611-621.
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and breast cancer outcomes: a systematic review and meta-analysis. J Clin Oncol 29, pp. 40-46.
10. Hou, G., Zhang, S., Zhang, X., Wang, P., Hao, X., Zhang, J. (2013). Clinical pathological characteristics
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807-816.
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Suceveanu, A.P., Bondari, S., Bondari, D., Voinea, F.(2015) Clinical presentation of a patient with Cutis
Laxa with systemic involvement: A case report Romanian Journal of Morphology and Embryology
56(3), pp. 1205-1210.
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women suffering from breast cancer in support groups. Procedia Social and Behavioural Sciences 128,
pp. 10-15.
13. Aschie, M, Mitroi, A., Cozaru, G., Poinareanu, I. (2015). Molecular profiles of invasive breast
carcinomas 467, S63-S63, Supplement: 1 Meeting Abstract: PS-01-063.
14. Aschie, M., Mitroi, A.F., Poinareanu, I., Cozaru, G.C. (2014). Comparison of HER2 status in breast
cancer versus gastric carcinoma. Virchows Archiv 465(S1), pp. S324-S325.
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pp. 394-424.
17. Aschie, M., Poinareanu, I., Cozaru, G. C., Mitroi, A. F., Branzan, C., Aschie, D et al., (2017).
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Spring St, New York, NY 10013 USA: Springer.
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NY 10013 USA: Springer.
228
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229
Clinical and Pathological Features of Patients with Colorectal

Cancer and Diabetes – a Single Centre Experience
MAZILU Laura1, SUCEVEANU Adrian-Paul1, PAREPA Irinel Raluca1,

GHEORGHE Andreea Daniela1, PANTEA STOIAN Anca2,
ARDELEANU Valeriu1, SUCEVEANU Andra Iulia1
2“Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
Emails: lauragrigorov@yahoo.com, asuceveanu@yahoo.com, irinel_parepa@yahoo.com,
andreea_gheorghe78@yahoo.com, ancastoian@yahoo.com, valeriu.ardeleanu@gmail.com, andrasuceveanu@yahoo.com
Abstract
In our days, according to GLOBOCAN, colorectal cancer represents the third leading cause
of death and fourth cancer in terms of world incidence. The incidence of colorectal cancer is
globally high, now representing 11% of all cancers, especially in developing countries that
adhere to the “western way of life”. Diabetes is one of the most frequent metabolic diseases,
the number of diabetic patients increased two times over the last 30 years.
Diabetes is known to be related to a different type of cancer, and it is now a recognized risk
factor for cancer development, including colorectal cancer. This study is a single-centre,
observational study, and was conducted in the Oncology Department of Clinical Emergency
Hospital of Constanta, on a group of 324 patients with colorectal cancer. Of these, 99 patients,
representing 30.6% of cases, had associated type 2 diabetes. Male were the majority of patients
in the diabetic group, but in the non-diabetic group, the majority of patients were women
(p=0.005). The median age was significantly higher in the diabetic patients’ group.
Regarding tumour location, most of the patients in both groups have left-sided tumours, with
no significant difference between groups. According to the stage, most of the patients have the
advanced disease in both groups. RAS mutation was found in 22.9% of non-diabetic patients,
and in 21.6% of diabetic patients with colorectal cancer. Four % of diabetic patients with
colorectal cancer and 7.1% of non-diabetic patients had microsatellite instability high (MSI-H).
Body mass index was significantly higher in patients with diabetes. Type 2 diabetes
represent a risk factor for a multitude of cancer types, including colorectal cancer. Colorectal
cancer prevention is based on the improvement of lifestyle, factors that are modifiable, and can
reduce the risk for both colorectal cancer and diabetes.
Keywords: colorectal cancer, diabetes
Introduction
In our days, according to GLOBOCAN, colorectal cancer represents the third leading cause
of death and the fourth cancer in terms of world incidence. The incidence of colorectal cancer
is globally, now representing 11% of all cancers, especially in developing countries that adhere
to the “Western way of life”. Risk factors associated with colorectal cancer development are
obesity, alcohol, smoking, red meat consumption, and reduced physical activity [1]. Trends in
colorectal cancer incidence and mortality are classified in three categories according to Human
Development Index (HDI): medium HDI countries with increasing incidence and mortality,
high-HDI countries with increased incidence but decreasing mortality due to improved
230
treatment modalities, and countries with highest HDI with a decrease in both incidence and
mortality [2-4]. Diabetes is one of the most frequent metabolic disease; the number of diabetic
patients increased two times over the last 30 years.
Diabetes is known to be related to a different type of cancer, and it is now a recognized risk
factor for the cancer occurrence. Diabetes influence the quality of life in a serious matter,
causing neurological and vascular complications, have a negative impact on adherence to
oncological treatment, and also on the prognosis of patients with colorectal cancer [5-7].
Methodology
This study is a single centre, observational study, and was conducted in Oncology
Department of Clinical Emergency Hospital of Constanta, on a group of 326 patients with
colorectal cancer, during four years, between 2016 and 2019. Informed consent was signed by
all participants before the collection of data. All data about the patients were obtained using
medical records of the Department. Statistical Package for the Social Sciences version 21
software (SPSS) was used to perform statistical analyses. All statistical tests performed were 2-
sided, and p-value <0.05 was considered statistically significant.
Results
The total number of studied patients accounted for 324 patients with a colorectal cancer
diagnosis. Of these 99 patients, representing 30.6% of patients with colorectal cancer included
in study had associated type 2 diabetes. Patients were split into two groups in reference to the
presence or absence of type 2 diabetes. Characteristics and comparisons of the two groups are
described in Table 1.
Table 1. Characteristics of patients with colorectal cancer

Characteristic Diabetes - Diabetes + P-value Analysis
(n=225) (n=99) between groups
Age 64.88 68.10 p=0.01
(34-88) (42-88)
Sex
Male 110 (48.9%) 65 (65.7%) p=0.005
Female 115 (51.1%) 34 (34.3%)
Age group
<50 years 19 (8.4%) 6 (6.1%) p=0.014
>50 years 206 (91.6%) 93 (93.9%)
Tumor location
Right 68 (30.2%) 28 (28.3%)
Left 157 (69.8%) 71 (71.7%) p=0.726
Colorectal cancer stage
I 22 (9.8%) 7 (7.1%)
II 77 (34.2% 35 (35.4%) p=0.722
III 72 (32.0%) 33 (33.3%)
IV 54 (24.0%) 24 (24.2%)
RAS mutation
WT 172 (77.1%) 76 (78.4%) p=0.810
MT 51 (22.9%) 21 (21.6%)
MSI status
MSI-H 16 (7.1%) 4 (4.0%) p=0.292
MSS 209 (92.9%) 95 (96.0%)
BMI
<25kg/m2 55 (70.5%) 59 (59.6%) p=0.045
25-30kg/m2 22 (28.2%) 33 (33.3%)
>30kg/m2 1 (1.3%) 7 (7.1%)
231
Male were the majority of patients in the diabetic group, but in the non-diabetic group, the
majority of patients were women, a difference that was statistically significant (p=0.005). The
median age was significantly higher in diabetic patients’ group, compared with non-diabetics.
Regarding tumour location, most of the patients in both groups have left-sided tumours, with
no significant difference between groups. According to the stage, most of the patients had the
advanced disease in both groups, with no significant difference between diabetic and non-
diabetic patients. RAS mutation was found in 22.9% of non-diabetic patients, and 21.6% of
diabetic patients with colorectal cancer. 4% of diabetic patients with colorectal cancer and 7.1%
of non-diabetic patients had microsatellite instability-high (MSI-H) with no significant
difference.
Body mass index was evaluated in all patients included in the analysis and was significantly
higher in patients with diabetes.
Discussions
Many environmental and host factors concur to colorectal cancer occurrence. According to
available epidemiological data, colorectal cancer is more frequent in males’ patients, patients
that have a 1.5-fold risk for the occurrence of colorectal cancer [1, 8]. This was also confirmed
in our study, and colorectal cancer was more frequent in men compared with women [9]. Still,
in the group of non-diabetic patients with colorectal cancer, the number of female patients was
higher. The specific climate and local risk factors can be involved in epidemiological
particularities in our geographic area [10-12].
Age is another risk factor for cancer in general, and patients over 65 years old are more
susceptible to develop cancer [13, 14]. In our study, the mean age was 64.88 years for non-
diabetic patients, compared with 68.10 years for diabetic patients. This data does not correspond
with literature data showing that cancer develops in diabetic patients at a younger age.
According to tumour location, no significant difference was found between the two groups
(p=0.726). Tumour location is a prognostic factor in colorectal cancer, as it is demonstrated by
a meta-analysis that included 1.427.846 patients, and which showed that left-sided tumour
location is associated with reducing risk of death independent of stage, race, and use of adjuvant
chemotherapy [15, 16]. No correlation was found between diabetes and stage of the disease
(p=0.722).
Accumulation of genetic and epigenetic mutations is involved in the pathogenesis of
colorectal cancer. RAS and BRAF mutations have prognostic value in colorectal cancer [12].
KRAS mutation can be found in 12-75% of patients with colorectal cancer, and it is
associated with poor prognosis [17-21]. In our analysis, RAS-WT (wild type) was found in
77.1% of non-diabetic patients and in 78.4% of diabetic patients with no clinical significance.
In our study, MSI-H tumours were found in 4.0% of diabetic patients, and in 7.1% of non-
diabetic patients. MSS tumours were found in 92.9% of non-diabetic patients, and 96.0% of
diabetic patients. There was no significant difference was found between groups (p=0.292).
The total number of MSI-H patients in our study was 20 (6.2%), higher than 3.5% reported
in literature [22]. Other studies conducted in Japan reported an MSI-H frequency between 3.1%
and 5.9% in patients with colorectal cancer, and also reported MSI-H more frequent in left
colon cancer, comparing to right colon cancer [23, 24].
Conclusion
Type 2 diabetes is an already known risk factor for a multitude of cancer types, including
colorectal cancer. Both type 2 diabetes and colorectal cancer represent a significant cause of
morbidity and mortality all over the world. “Western lifestyle” that includes diet, physical
232
inactivity and obesity are linked to an enhanced risk for colorectal cancer, and this is supported
by an impressive number of epidemiological analysis. Colorectal cancer prevention is now
based on the improvement of lifestyle, factors that are modifiable, and can reduce the risk for
both colorectal cancer and diabetes.
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Springer.
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survival, and risk factors. Prz Gastroenterol 14(2), pp. 89-103.
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risk. World J Gastroenterol 21, pp. 5167-75
9. Tulin, A., Slavu, I., Tulin, R. et al., (2018). Does sex of the patient play a role in survival for MSI
colorectal cancer? Journal Of Mind And Medical Sciences 5(1), pp. 101-108.
10. Lupu A.A., Ionescu E.V., Iliescu M.G., Almasan R.E., Oprea C, Ion I., Iliescu D.M. Effects of Techirghiol
specific climate factors on the patient’s quality of life with degenerative lumbar pain – Journal of
environmental protection and ecology 2018, 19 (4), pp. 1857-1863.
11. Iliescu, M.G., Profir, D., Surdu, O., Marin V., Demirgean, S., Almasan, R.E., Stanciu, L.E., Oprea, C.,
Iliescu, D.M. (2018). Statistical view through balneal activity in Techirghiol medical area. Journal of
environmental protection and ecology 19(1), pp. 382-391.
12. Stefanescu, H., Muntean, D., Pilut, C., et al., (2018). Using Blood and Plasma MicroRNAs as a Non-
Invasive Biomarker in Patients with Colorectal Cancer. Clinical Laboratory 64(3), pp. 257-262.
13. Rawla, P., Sunkara, T., Muralidharan, P., Pradeep Raj, J. (2018). Update in global trends and aetiology
of hepatocellular carcinoma. Contemp Oncol 22, pp. 141-50.
14. Tofolean, D.E., Mazilu, L., Staniceanu, F., Mocanu, L., Suceveanu, A.I., Baz, R., Parepa, R.I., Suceveanu,
A.P., Bondari, S., Bondari, D., Voinea, F.(2015) Clinical presentation of a patient with Cutis Laxa with
systemic involvement: A case report Romanian Journal of Morphology and Embryology 56(3), pp. 1205-
1210.
15. Petrelli, F., Tomasello, G., Borgonovo, K., et al., (2016). Prognostic Survival Associated with Left-Sided
vs Right-Sided Colon Cancer: A Systematic Review and Meta-analysis. JAMA Oncol; p. 211.
16. Nitipir, C., Baetu, A., Voinea, A., et al., (2018). Peripheral Neurotoxicity Induced by Taxanes, Cisplatin,
Oxaliplatin, Fluoropyrimidines and Vinorelbine A clinical perspective. REV. CHIM. (Bucharest) 69(12),
pp. 3427-3432.
17. Modest, D.P., Ricard, I., Heinemann, V., et al., (2016). Outcome according to KRAS-, NRAS – and
BRAF-mutation as well as KRAS mutation variants: pooled analysis of five randomized trials in
metastatic colorectal cancer by the AIO colorectal cancer study group. Ann Oncol 27, p. 1746.
18. Taieb, J., Le Malicot, K., Shi, Q., et al., (2017). Prognostic Value of BRAF and KRAS Mutations in MSI
and MSS Stage III Colon Cancer. J Natl Cancer Inst, p. 109.
19. Aschie, M., Poinareanu, I., Cozaru, G. C., Mitroi, A. F., Branzan, C., Aschie, D et al., (2017).
Genotyping-RAS mutations analysis in colorectal cancer. Virchows archiv, 471, pp. S198-s198. 233
Spring St, New York, Ny 10013 USA: Springer.
20. Maiorean, A., Aschie, M., Poinareanu, I., Mitroi, A., Cozaru, G., Enciu, M., Chisoi, A. (2017).
Morphological and immunohistochemical aspects of premalignant lesions in colonic polyposis. Case
reports with short literature review. Virchows Archiv, 471, pp. S317-S317. 233 Spring St, New York, Ny
10013 USA: Springer.
233
21. Brinzan, C., Aschie, M., Grasa, C. N., Mitroi, A. F., Matei, E., & Cozaru, G. C. (2019). The Mutation
Profiles of KRAS and BRAF Genes in a Romanian Colorectal Cancer Cohort. REV.CHIM(Bucharest),
70(4), pp. 1346-1350.
22. Smith, CG., Fisher, D., Claes, B., et al., (2013). Somatic profiling of the epidermal growth factor receptor
pathway in tumours from patients with advanced colorectal cancer treated with chemotherapy ±
cetuximab. Clin Cancer Res 19, p. 4104.
23. Nakayama, Y., Iijima, T., Wakaume, R., et al., (2019). Microsatellite instability is inversely associated
with type 2 diabetes mellitus in colorectal cancer. PLoS One 14(4), e0215513
24. Asaka, S., Arai, Y., Nishimura, Y., Yamaguchi, K., Ishikubo, T., Yatsuoka, T., et al., (2009).
Microsatellite instability-low colorectal cancer acquires a KRAS mutation during the progression from
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234
Statins and Risk Biomarkers for Coronary Heart Disease (CHD)

in Diabetes Mellitus Patients with Normal LDL Cholesterol
Levels. 1 Years Prospective Study in Constanta County, Romania
PAREPA Irinel Raluca1, SUCEVEANU Andra Iulia1, PANTEA-STOIAN Anca2,

MAZILU Laura1, CATRINOIU Doina1, IONESCU Paris1, IONESCU Mihaela1
2Faculty of Medicine, “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
Emails: irinel_parepa@yahoo.com, ciucea_mihaela@yahoo.com
Abstract
Introduction
Statin therapy is currently used for primary prophylaxis in patients with abnormal lipid
profile and for secondary prophylaxis in patients with established coronary heart disease (CHD)
and/or diabetes mellitus.
The objective of the study

Our study aimed to evaluate if statin therapy has a protective role against CHD risk
biomarkers other than dyslipidaemia in a high-risk category of patients.
Materials and methods

32 patients with type 2 diabetes mellitus were followed-up for 1 year. Inclusion criteria were:
normal LDL-cholesterol levels, no antidiabetic therapy (diet only), no previous evidence of
CHD (normal resting/effort electrocardiography, normal coronary arteries at angiography),
normal blood pressure, normal creatinine clearance and no previous statin therapy. Patients
received rosuvastatin 10mg/day. C reactive protein (CRP), fibrinogen, microalbuminuria and
fasting glucose levels were evaluated at baseline and after 1 year of prophylactic statin
treatment; patients undergone also another angiographic investigation at the end of the study
period. Transaminases serum levels were evaluated at baseline and every 2 months during study
period.
Results
The average levels of the evaluated biomarkers have shown a significant descendant trend:
fibrinogen from 410±15 mg/dl at baseline to 312±12mg/dl after 1 year (p<0.0001); CRP from
0.82±0.05mg/dl to 0.48±0.04 mg/dl (p<0.0001); microalbuminuria from 74±12μg/min to
36±10μg/min (p<0.0001) and fasting glucose from 156±7mg/dl to 133±4mg/dl (p<0.02).
After 1 year 3 patients (9.3%) developed non-significant coronary arteries stenosis (less than
40% obstruction) and no patient developed clinical signs of CHD. Only 1 patient (3.1%) had to
be withdrawn from statin therapy due to pathologic increase in transaminases levels.
Conclusion
In our study, statin therapy significantly decreased the levels of the evaluated CHD risk
biomarkers (others than abnormal lipid profile) in a population with high cardiovascular risk.
Keywords: diabetes mellitus, coronary heart disease, statin, biomarkers
235
Introduction
“It is more important to know what sort of person has a disease than to know what sort of
disease a person has.” Hippocrates.
Although on an individual level, it is important to diagnose each person with his own
diseases, for the benefit of the public’s health, it is necessary to find ways to identify groups
of people who may be at risk and to aspire to relieve those risks either before an event happens
or prior to its reoccurrence [1].
Statin therapy is currently used for primary prophylaxis in patients with abnormal lipid
profile and for secondary prophylaxis in patients with established coronary heart disease
(CHD) and/or diabetes mellitus and is considered as the cornerstone of clinician’s efforts
toward primary and secondary CHD prevention in patients with type 2 diabetes mellitus
(T2DM) [2]. Patients with diabetes have a considerably higher risk of atherosclerotic vascular
disease. Data from randomized controlled trials and observational studies made on T2DM
patients with normal LDL-cholesterol levels who take statins for primary prevention of CHD
are limited.
It is known that statins have an active effect in preventing cardiovascular (CV) events and
reducing CV mortality and their use is associated with a small number of adverse events.
Because of the high-risk profile of patients with T2DM, intensive statin treatment should
be used on an individualized basis [3, 4, 5]. Although statins are significantly proven to reduce
the CV-related events and all-cause mortality [6], underutilization of statins in patients at high
risk such as those over 40 years with T2DM is reported in the form of discontinuation,
inappropriate dosing and adherence issues [7].
Similar to the 2013 ACC/AHA guidelines for CV disease prevention [8], the new American
Diabetes Association (ADA) statement has not specified any LDL-cholesterol goals for statin
therapy. Statin is dosed based on patient’s risk factors for atherosclerotic cardiovascular
disease (CVD) rather than on single LDL-cholesterol levels. High-dose statin therapy is
recommended for all patients with T2DM with a history of clinical CHD or with at least one
additional CVD risk factor. Moderate-dose statin therapy is advised for patients younger than
40 or older than 70 who have CVD risk factors [9].
Methods
Study population, definition of risk factors, and clinical follow-up

Our prospective study was conducted in the Diabetology and Cardiology Departments of
the County Clinical Emergency Hospital of Constanta, Romania – between January 2018 and
January 2019. The study aimed to evaluate if statin therapy has a protective role against CHD
risk biomarkers other than dyslipidaemia in a high-risk category of patients. The study
population consists of 32 patients with type 2 diabetes mellitus who started a prophylactic,
moderate-to-high statin therapy (rosuvastatin 10 mg /day) over a period of one-year follow-
up.
Study entrants had no documented previous history of CHD (normal resting/effort
electrocardiography, normal coronary arteries at angiography), no antidiabetic therapy (diet
only), normal LDL-cholesterol levels (less than 100 mg/dl), normal blood pressure, normal
creatinine clearance and no previous statin therapy. Normal blood pressure was defined as a
systolic blood pressure of <130 mmHg and/or a diastolic blood pressure of <80 mmHg and
diabetes mellitus was defined as a fasting blood glucose level ≥126 mg/dl in at least two
different measurements, and a glycated haemoglobin A1c (HbA1c) level ≥6.5%.
Biomarkers play an important role in the evaluation of disease as well as in the
development of drug treatments for disease conditions, thus there are many CHD biomarkers
236
currently under study. The biomarkers we survey can be grouped according to the pathologic
process they represent, such as inflammatory (e.g., C-reactive protein), prothrombotic (e.g.,
fibrinogen), metabolic-related (e.g., fasting glucose, microalbuminuria) [10]. C reactive
protein (CRP), fibrinogen, microalbuminuria and fasting glucose levels were evaluated at
baseline and after 1 year of prophylactic statin treatment; patients undergone also another
coronary artery angiography at the end of the study period. Transaminases serum levels were
checked at baseline and every 2 months along study period to prevent this incident [11].
Results
Demographic, clinical and biological features of enrolled subjects at baseline are resumed
in Table 1.
Table 1. Baseline features of studied subjects

Demographic/clinical/laboratory feature Study group
Age (mean, years) ±SD 43.4±11.6
Gender (M/F) 17/15
Urban/Rural 21/11
Smoking status Non-smokers all
BMI (mean, kg/m2) 28.3±6.4
eGFR (mean, ml/min/m2) 76.4±10.6
LDL-cholesterol (mean, mg/dl) 92.2±7.3
Fasting glucose (mean, mg/dl) 146±7
Systolic blood pressure (mean, mmHg) 123.2±6.3
Diastolic blood pressure (mean, mmHg) 72.8±6.4
The average levels of the evaluated biomarkers have shown a significant descendant trend:
fibrinogen from 410±15 mg/dl at baseline to 312±12mg/dl after 1 year (p<0.0001); CRP from
0.82±0.05mg/dl to 0.48±0.04 mg/dl (p<0.0001); microalbuminuria from 74±12μg/min to
36±10μg/min (p<0.0001) and fasting glucose from 146±7mg/dl to 133±4mg/dl (p<0.02).
Table 2. Biomarkers trend along studied period

Biomarker Baseline End-of-study p
Fibrinogen (mean, mg/dl) 410±15 312±12 <0.0001
CRP (mean, mg/dl) 0.82±0.05 0.48±0.04 <0.0001
Microalbuminuria (mean, μg/min) 74±12 36±10 <0.0001
Fasting glucose (mean, mg/dl) 146±7 146±7 <0.02
After 1 year, only 3 patients (9.3%) developed non-significant coronary arteries stenosis
(less than 40% obstruction) and no patient developed clinical signs of CHD. Only 1 patient
(3.1%) had to be withdrawn from statin therapy due to pathologic increase in transaminases
levels.
Discussion
In this prospective evaluation of prophylactic prevention in a population with high CV risk,

we found significant reductions in all evaluated biomarkers: fibrinogen (23.9% lower), CRP
(41.46% lower), microalbuminuria (51.35% lower and fasting glucose (14.74% lower).
The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) have been
assumed to have direct anti-inflammatory effects, an important issue since inflammation plays
a major role in determining atherosclerotic plaque vulnerability [12]. In accordance with other
237
studies and trials, these data provide evidence that statins may have anti-inflammatory effects
in addition to the lipid-lowering effects, as well as they might slow down the development of
diabetes, a disease triggered in part through inflammatory mechanisms [13, 14].
Both in nondiabetic and diabetic subjects, microalbuminuria is associated with an increased
CV risk, independent of other risk determinants. The association between microalbuminuria
and CV disease is probably explained by a common pathophysiologic process, such as
endothelial dysfunction or chronic, low-grade inflammation [15].
Therapy with statins used among patients with T2DM has been associated with absolute
benefits in terms of CHD prevention, as our data demonstrate that the effect of rosuvastatin
on CRP, initially observed in the CARE trial of secondary prevention, is also present among
patients with no prior history of CVD. AFCAPS trial also suggests that the use of statins may
be effective in the primary prevention of acute coronary events among those with elevated
levels of CRP, even in the absence of overt dyslipidaemia, concluding that the effect of statins
on CRP levels may have clinical importance [16, 17].
Only 1 patient (3.1%) had to be withdrawn from statin therapy due to pathologic increase
in transaminases levels. Safety concerns, mainly regarding hepatotoxicity, have driven
numerous studies and trials, which have evidenced the low incidence of statin-related hepatic
adverse effects; the most commonly reported has been the occurrence of “transaminitis”, in
which liver enzyme levels are elevated in the absence of proven hepatotoxicity. This class
effect is usually asymptomatic, reversible, and dose-related [11].
Further trials are needed to test whether diabetic patients with low LDL-cholesterol levels,
but with a high CV risk profile also achieve a substantial benefit from prophylactic statin
therapy.
Disclosure
All authors had equal scientific contribution to this study.
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1. Montgomery JE, Brown JR. Metabolic biomarkers for predicting cardiovascular disease. Vasc Health
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eligibility, coronary artery calcification, and cardiovascular events. JAMA. 2015; 314 pp. :134-41.
3. Valensi P, Picard S. Lipids, lipid-lowering therapy and diabetes complications. Diabetes Metab 2011;
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5. Stone NJ, et al., 2013 ACC/AHA guideline on the treatment of blood cholesterol to reduce
atherosclerotic cardiovascular risk in adults: A report of the American College of Cardiology/American
Heart Association Task Force on Practice Guidelines. J Am Coll Cardiol. 2014; 63: pp. 2889-934.
6. LaRosa JC, He J, Vupputuri S. Effect of statins on risk of coronary disease: A meta-analysis of
randomized controlled trials. JAMA. 1999; 282: pp. 2340-6.
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world population of patients at high cardiovascular risk. J Manag Care Spec Pharm. 2016; 22: pp. 685-
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cardiovascular risk: a report of the American College of Cardiology/American Heart Association Task
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risk: a scientific statement from the American Heart Association. Circulation. 2009; 119(17): pp. 2408-
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13. Freeman DJ, et al., Pravastatin and the development of diabetes mellitus: evidence for a protective
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14. Ridker PM. High-sensitivity C-reactive protein: potential adjunct for global risk assessment in the
primary prevention of cardiovascular disease. Circulation. 2001; 103: pp. 1813-1818.
15. Coen D.A. Stehouwer and Yvo M. Smulders. Microalbuminuria and Risk for Cardiovascular Disease:
Analysis of Potential Mechanisms. JASN August 2006, 17(8) pp. 2106-2111.
16. Ridker PM, Stampfer MJ, Rifai N. Novel risk factors for systemic atherosclerosis: a comparison of C-
reactive protein, fibrinogen, homocysteine, lipoprotein-a, and standard cholesterol screening as
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2016; 388: pp. 2532-61.
239
The Relationship Between Arterial Stiffness and 25-OH Vitamin

D3 in Type 2 Diabetes Mellitus Patients
IURCIUC Stela1, TUDOR Anca2, RADA Maria1,

BADALICA-PETRESCU Marius1, PAH Ana1,
STOICHESCU-HOGEA Gheorghe1, CRACIUN Laura1, IURCIUC Mircea1,
ORAVITAN Mihaela3
1 Department of Cardiology, “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
2 Department of Functional Sciences, “Victor Babes” University of Medicine and Pharmacy, Timisoara (ROMANIA)
3 Department of Physical Therapy and Special Motricity, West University, Timisoara (ROMANIA)
Emails: stela_iurciuc@yahoo.com, atudor@umft.ro, radamariam@gmail.com, marius_badalica@yahoo.com,

ana11p@yahoo.com, goguhogea@yahoo.com, drlauracraciun@gmail.com, mirceaiurciuc@gmail.com,
mihaela.oravitan@gmail.com
Abstract
Objective/Purpose
We evaluate the vascular stiffness parameters in a group of 50 type 2 diabetes mellitus
(T2DM) patients.
Design and Method

The study included 50 T2DM patients compared with a control group of 50 healthy age-
matched subjects (CON). Serum levels of 25-OH vitamin D3 were determined by ELISA.
Carotid-femoral pulse wave velocity (c-f PWV) was determined using the Arteriograph
device. The measurement of the intima-media thickness (IMT) was made using ultrasound at
the carotid artery level. Renal resistivity index (RRI) was measured by duplex Doppler.
Results
We observed a significant differences between the two groups regarding HbA1c (8.44±2.26
in T2DM vs. 6.50±1.40 in CON, p<0.001), c-f PWV (9.86±3.09 m/s in T2DM vs. 7.62±1.47 in
CON, p<0.001), IMT (1.9±0.8 mm in T2DM vs. 0.8±0.5 mm in CON, p<0.001) and 25 OH-
D3 (20.49±7.31 mg/dL in T2DM vs. 37.09±4.59 in CON, p<0.001). A statistically significant
correlation was found between 25-OH-D3 and HbA1c values (p<0.001), and c-f PWV (p<0.001)
and IMT (r=-0.473, p<0.001).
Conclusion
This study demonstrated that measurement of c-f PWV and IMT are convenient and reliable
methods in T2DM patients and together with determination of 25-OH-D3 can be used to
monitoring arterial stiffness in these patients.
Keywords: vascular stiffness, type 2 diabetes mellitus, 25-OH vitamin D3, carotid femoral pulse wave velocity,
intima media-thickness
Introduction
Cardiovascular disease and diabetes mellitus are the most common diseases of our times and
their rate is still rising. Mortality rates from cardiovascular diseases in the last decades were
240
reported to be approximately 1.7 times higher in adults diagnosed with diabetes mellitus than
in adults without diabetes [1].
The association between diabetes and increased arterial stiffness has been widely reported
in recent years. Muhammad et al., in a study conducted on more than 2400 subjects explore the
relationship between incidence of diabetes and arterial stiffness during a follow-up of 5 years.
They observed that the incidence of diabetes has been associated with increased carotid-
femoral pulse wave velocity (c-f PWV), independent of other risk factors. These findings
support the idea that increased arterial stiffness is an early marker of risk for development of
diabetes [2]. It has also been shown by numerous studies that arterial rigidity is closely related
to the presence of diabetes complications (diabetic retinopathy, nephropathy and neuropathy)
[3]. In a study conducted by Gencay et al., it was observed a decrease in arterial stiffness after
performing high‐intensity anaerobic cycling exercise [4]. Furthermore, dynamic stability
training was showed to improve life style in healthy people [5, 6] and also in type 2 diabetic
mellitus patients [7].
Assessing and studying circulating biomarkers involved in various pathways in arterial
stiffness is a potential method to elucidate key factors associated with this multifactorial
disorder [8, 9]. Inflammatory biomarkers have a major role in arterial stiffness process, related
to atherosclerosis [10]. The role of vitamin D in the pathogenesis of atherosclerosis is not only
linked by the presence of high blood pressure and arterial stiffness, (8) but also by vascular
calcification, a universal feature of atherosclerosis, which can be considered synonymous in
this perspective.
As a result, the aim of this study was to correlate arterial stiffness measured by the vascular
calcification parameters, with 25-OH vitamin D3 levels in T2DM patients compared with
healthy age-matched subjects.
Patients and Methods
This study included 50 patients with T2DM without coronary artery disease, stroke,
peripheral arterial disease or any other form of cardiovascular disease (aortic aneurysm or
dissection), hospitalized in Clinic of Prevention and Rehabilitation, Institute of Cardiovascular
Diseases Timişoara, during July 2018-July 2019. All patients were known diabetics previously
diagnosed according to the consensus report by the American Diabetes Association (ADA) and
the European Association for the Study of Diabetes (EASD) in 2018 based on fasting plasma
glucose >126 mg/dL or 2h plasma glucose >200 mg/dL by oral glucose tolerance test (OGTT)
[11]. The T2DM patients were compared with a control group of 50 healthy age-matched
subjects (CON), selected in the same period from a data base screened for cardiovascular risk
factors delivered by primary care physicians who collaborate with our clinic.
Vitamin D was determined using commercial ELISA kit. 25-OH-D3 values were interpreted
as follows: a value greater than or equal to 30 ng/mL was considered sufficient, values between
21 ng/mL to 29 ng/mL were considered insufficient, the deficit was defined by values under 20
ng/mL. The “gold-standard” measurement of arterial stiffness, c-f PWV, was performed with
the Medexpert Arteriograph device [12]. Determination of IMT in the carotid artery was
performed by the average of three measurements in each of the three successive segments of
the vessel, according to the Mannheim consensus [13]. The presence of the plaque was defined
by an IMT value >0.9 mm.
241
Results
Clinical, biochemical and demographic features of the patients are summarized in Table 1.
No significant difference between the 2 groups was observed regarding age, body mass
index, gender, systolic blood pressure, diastolic blood pressure and heart rate.
There were significant differences between the two groups regarding HbA1c (8.44±2.26 in
T2DM vs. 6.50±1.40 in CON, p<0.001), c-f PWV (9.86±3.09 m/s in T2DM vs. 7.62±1.47 in
CON, p<0.001), IMT (1.9±0.8 mm in T2DM vs. 0.8±0.5 mm in CON, p<0.001) and 25 OH-
D3 (20.49±7.31 mg/dL in T2DM vs. 37.09±4.59 in CON, p<0.001).
Table 1. Characteristics of all patients (n=100)

T2DM Controls p value
n 50 50
Age, y 67.33±9.10 67.44±8.98 0.238
Male sex, n (%) 25(50.00) 24(48.33) 0.991
BMI, kg/m 2
33.12±3.82 32.63±4.76 0.081
SBP, mm Hg 139.95±28.27 129.23±31.22 0.064
DBP, mm Hg 81.84±14.75 76.87±26.33 0.183
HR, bpm 81.68±21.04 78.75±15.11 0.174
T2DM, n (%) 50 (100.00) - -
HbA1c, % 8.44±2.26 6.50±1.40 <0.001
eGFR, ml/min/1,73m 2
55.78±24.27 87.30±13.31 <0.001
c-f PWV, m/s 9.86±3.09 7.62±1.47 <0.001
25-OH-D3, ng/mL 20.49±7.31 37.09±4.59 <0.001
IMT, mm 1.9±0.8 0.8±0.5 <0.001
BMI, Body mass index; SBP, systolic blood pressure; DBP, diastolic blood pressure; HR,
heart rate; T2DM, type 2 diabetes mellitus, HbA1C, glycated haemoglobin, eGFR, estimated
glomerular filtration rate; c-f PWV, carotid-femoral pulse wave velocity; 25-OH-D3, 25-
hydroxyvitamin D; .IMT, intima-media thickness. Values were expressed as mean ± standard
deviation (SD)
Fig. 1. (a) (b) (c)
A strong negative correlation was found between HbA1c values and 25-OH-D3 (r=-0.607,
p<0.001) (Fig. 1a), c-f PWV and 25-OH-D3 (r=-0.711, p<0.001) (Fig. 1b) and a negative,
significant and medium correlation was observed between IMT and 25-OH-D3 (r=-0.473,
p<0.001) (Fig. 1c).
Discussions
In the present study we observed that IMT and c-f PWV as indicators of arterial stiffness
and serum 25-OH-D3 levels as biomarker of arterial stiffness were significantly modified in
242
T2DM patients compared to control. Numerous studies have demonstrated a relationship

between diabetes mellitus and arterial stiffness [3, 14, 15] or with 25-hydroxivitamin D3 [16].
In our study a statistically significant correlation was found between 25-OH-D3 and HbA1c
values (p<0.001), and c-f PWV (p<0.001) and IMT (r= -0.473, p<0.001). These results suggest
that vitamin D may be related to glucose control in diabetes mellitus type 2. In addition,
statistically significantly more T2DM patients than the control group had vitamin D deficiency
(p<0.001). These results are the same with literature [16, 17].
It has been shown that vitamin D deficiency is being related to the development of
autoimmune disease, like rheumatic inflammatory diseases [18-20] or diabetes mellitus type 1
[21].
Aging per se is associated with arterial stiffening, atherosclerosis and endothelial
dysfunction [12], but in the presence of a sedentary lifestyle or insulin resistance [22] and
coexistence of other illnesses [23-25] with increased morbidity, arterial stiffness becomes an
important risk factor for future cardiovascular events in patients with T2DM [26, 27].
Conclusions
This study demonstrated that measurement of c-f PWV and IMT are convenient and reliable
methods in T2DM patients and together with determination of 25-OH-D3 can be used to
monitoring arterial stiffness in these patients. These techniques might play a key role in order
to make appropriate managerial decision for patients with T2DM.
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(Kaunas, Lithuania). 2019; 55(6)
9. Georgescu D, Iurciuc MS, Ionita I, Dragan S, Muntean M, Ancusa OE, et al., Migraine without Aura
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Lithuania). 2019; 55(12).
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(ADA) and the European Association for the Study of Diabetes (EASD). Diabetologia. 2018; 61(12):
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12. Iurciuc S, Cimpean AM, Mitu F, Heredea R, Iurciuc M. Vascular aging and subclinical atherosclerosis:
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24. Jurca-Simina IE, Juganaru I, Iurciuc MS, Iurciuc S, Ungureanu E, Dobrescu AI, et al., What if body fat
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Systematic Literature Review. Medicina (Kaunas, Lithuania). 2019; 55(6).
26. Iurciuc S, Vlad A, Iurciuc M, Avram A, Cioraca G, Avram C, et al., RISK FACTOR AND LIFESTYLE
CHANGES IN HIGH RISK PATIENTS: PP.23.442. Journal of hypertension. 2010; 28: pp. e377-e8.
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244
Maternal Obesity: A New Challenge of Modern-Day Medicine –

Case Report
DIMITRIU Mihai1,2, NICULAE Mihai-Bogdan3, GHEORGHIU Diana-Claudia2,

HARADJA Horatiu2, CIRSTOVEANU Catalin Gabriel4,
DAN Adelina-Loredana2, IONESCU Andra-Maria2
3 Clinical Hospital Colentina, Bucharest (ROMANIA)
4 “Marie Curie” Children’ Clinical Hospital, Bucharest (ROMANIA)
Emails: drmihaidimitriu@yahoo.com, hharadja@gmail.com, danadelina90@gmail.com, ionescu.andra001@gmail.com
Abstract
The significant increase in obesity worldwide has become a real problem that obstetricians
are facing more and more often. This major health problem brings many risks for both mother
and baby, in the short and long term. For this reason, medical personnel should be trained to try
to fight this epidemy and be able to provide medical assistance when needed. We report the
case of a 28-year-old woman morbidly obese pregnant woman, who delivered by Caesarean-
section with favourable evolution.
Keywords: Obesity, Maternal health, Fetal health, Obstetric complications
Introduction
The prevalence of obese people has raised at an alarming pace in the last decades, and this
does not rule out a pregnant woman. In 2016 more than 1.9 billion adult people were
overweight, from which 650 million were obese [1]. Regarding the percentage of
overweight/obese women, this is 39% [1]. Romania is seventh in terms of overweight/obesity
when it comes to women in the European region [2].
Many factors are involved, which include lifestyle, technologic and economic advances
which created a sedentary population with access to cheap and high-calorie foods. The
community moves less and eats more. This is one of the central reasons why this problem is
growing alarmingly in developed and developing countries. Metabolic disorders are proven in
the obese population, being closely linked to the modern food industry [3]. Obesity is a public
health problem and a real challenge in modern medicine from all points of view.
Maternal obesity, pre-existing or developed during pregnancy, brings with its problems
regarding the development of comorbidities, the assistance of the pregnancy, birth and
aftercare, intrauterine development of the fetus as well as the evolution of the child later in life
(Table 1) [4].
The most widely used tool for the classification of obesity is body mass index (individual’s
weight – kilograms divided by the square of his or her height – meters) (Table 2.). This is not
the perfect variant for this representation. As an example of this conclusion are bodybuilders,
who have a high body mass index, given by the high percentage of muscle mass and not fat, as
it happens for obese people. Such people have no metabolic disease risks because this threat
comes with excess adipose tissue [4]. Even though this method is not perfect, it is the most
useful option we have available to classify obesity.
245
Table 3. Obstetric complications of obese pregnant women

Spontaneous conception
Miscarriage
IVF conception
Recurrent miscarriage
Early pregnancy Neural tube defects
Congenital anomalies Spina bifida
Congenital heart disease
Omphalocele
Gestational nonproteinuric hypertension
Preeclampsia
Late pregnancy Gestational diabetes mellitus
Preterm birth
Intrauterine fetal demise (stillbirth)
Caesarean delivery
Decreased VBAC success
Anaesthesia complications
Peripartum Excessive blood loss
Operative morbidity Postpartum endometritis
Wound infection/breakdown
Postpartum thrombophlebitis
Fetal macrosomia (EFW ≥4500 g)
Fetal/neonatal complications
Shoulder dystocia
Childhood obesity
EFW: estimated fetal weight; IVF: In vitro fertilization; VBAC – vaginal birth after caesarean
Table 4. BMI chart

Weight categories BMI (kg/m2)
Underweight <18.5
Healthy weight 18.5-24.9
Overweight 25-29.9
Obese 30-34.9
Severe obese 35-39.9
Morbidly obese ≥40
The standard definition of the non-pregnant population is not accurate for pregnant women,
as they have a physiological weight gain that is lost with the birth (fetus, placenta, amniotic
fluid, blood). This should be considered when evaluating patients.
Excess adipose tissue is an important endocrine organ that can lead to excessive hormonal
release and determine metabolic, inflammatory, vascular pathways imbalances, which affect
many organs [5-7].
Due to the increased risks associated with pregnancy in an obese woman preconception
counselling, evaluation and care thru the pregnancy and after birth should be offered [8]. It
would be ideal to be able to inform patients before conception about the adverse reactions of
obesity on fertility, about complications associated with obesity and pregnancy, evaluation of
comorbidities related to obesity, offering guidance regarding weight loss, bariatric surgery
benefits and risks [9]. Management of such pregnancy should include maternal weight, BMI,
blood pressure, early ultrasound, medication review (stop weight loss medication for example),
diabetes screening, usual blood tests, evaluation of nutritional deficiencies and treating them
for women who undergo bariatric surgery [10-13]. The obese pregnant woman should reduce
their weight gain as much as possible. Exercise should be encouraged, as it has many health
benefits. Screening for fetal aneuploidy should be performed even though it is not at a higher
risk in such patients, it is more challenging to be completed and should be done by experienced
physicians also fetal ultrasound survey [14, 15]. At regular follow-ups screening for gestational
246
diabetes should be performed, assessment of fetal growth and its well-being. For labour and
delivery, the maternity unit should have the right equipment and instruments for such a patient,
continuous fetal heart rate tracing, an anaesthesia consultation.
The risk of stillbirth, pain in the postpartum period and some anomalies are increased in the
obese patient [16-18]. Postpartum contraception in obese and diabetic patients is challenging
[19, 20]. If the delivery comes to a C-section, thromboprophylaxis and antibiotic prophylaxis
are needed. Careful postoperative monitoring dramatically reduces the occurrence of
complications. Intrauterine contraception is preferred in such cases [5, 6].
Case Presentation
We shortly present an extreme case of a 28-year-old pregnant obese woman, who was
admitted in the Department of Obstetrics & Gynaecology “Sf. Pantelimon” Emergency Clinical
Hospital, Bucharest, Romania, on August 2018, after being transferred from a smaller clinic,
for entering labour and high blood pressure. From the patient’s personal history, we take note
of no previous pregnancies, estimated 42 weeks of gestation, no pregnancy evaluation before
the current exam, BMI=57.14 kg/m2 (morbidly obese), smoker (10 cigarettes/day).
At admission in our department, the patient was cooperative; blood pressure was 220/120
mmHg, pulse 130 beats/minute. The obstetrical examination showed a pregnant uterus with
difficult Leopold’s manoeuvres to perform due to excess abdominal fat tissue. Local
examination with the speculum showed no macroscopic lesions on the cervix and no bleeding
or amniotic fluid loss. On the bimanual examination, we could identify dilatation of 2-3 cm,
intact membranes, cranial presentation, birth canal challenging to assess. Laboratory exams
were completed and showed high glucose levels. Transabdominal ultrasound was performed
and revealed one live fetus, cranial presentation, with estimated weight around 4000 g, placental
maturation grade III and AFI = 3.43 cm.
The informed consent was obtained, and C-section was performed under spinal anaesthesia
(spinal block/intradural block/intrathecal block) and followed by extracting a live fetus,
feminine, 3900 g, Apgar 8. After surgery, the patient was transferred to the intensive care unit,
so her vitals were continuously supervised.
The postoperative evolution was favourable, without any complications. On day 15, the
patient was released from the hospital with a good general condition and afebrile.
The patient did not come for the six weeks after birth follow-up.
Discussions
All physicians must address this problem with their patients and explain all the issues that
come with it. It has been showed that a better understanding and the help of doctors could
reduce the number of patients that become obese. Surveillance of such pregnancies is difficult;
most of the times patients encounter health problems, even life-threatening ones.
The birth method is adapted from case to case depending on the associated pathologies and
the risks determined by them.
Conduction of patients after surgery when it comes to obesity is difficult because of the many
complications that may appear regarding the operative wound and its healing, difficult
mobilisation and comorbidities associated with obesity.
Conclusion
In the current context, the incidence of obesity knows new limits, thus having a significant
impact on the evolution of pregnancies.
247
One can sense an increase in births by caesarean section and pre- and postoperative
complications caused by obesity.
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249
Coping Strategies, Illness Perceptions and Optimism:

Contribution to Self-Management and Adherence in Diabetic
Patients
POPA-VELEA Ovidiu1, APOSTOL Alina Florina2,

DIACONESCU Liliana Veronica1
1
Department of Medical Psychology, Faculty of Medicine, “Carol Davila” University of Medicine and
Pharmacy, Bucharest (ROMANIA)
2
“Sf. Ioan” Clinical Hospital, Bucharest (ROMANIA)
Emails: ovidiu.popa-velea@umfcd.ro, liliana.diaconescu@umfcd.ro, alina.apostol@mail.com
Abstract
Aims
Self-management and adherence in diabetes are therapy aims which are consistent with a
higher quality of life of affected patients and with a better prognosis. This study explored the
relationships between these variables and illness perceptions, coping strategies and optimism.
Methods
The design of the study was cross-sectional and included 120 patients (52 men and 68
women) suffering from type 2 diabetes. They were administered Brief Illness Perception
Questionnaire (BIPQ), Brief COPE, Life Orientation Test-Revised (LOT-R), Diabetes Self-
Management Questionnaire (DSMQ) and Medication Adherence Report Scale (MARS).
Pearson’s correlations and multiple regressions were subsequently run, in order to point out
the most significant correlations between the study variables.
Results
In terms of disease perception, the highest negative impact was represented by the duration
of the disease. Across the whole sample, the participants used predominantly functional coping
strategies. Optimism levels were below the recommended threshold, while self-management
and adherence were acceptable. Adherence and global self-management scores were correlated
positively with functional coping strategies and negatively with dysfunctional ones, with men
displaying less significantly this association. Self-management was predicted by active coping
(t=3.107, p<.002), self-distraction (t=-2.472, p<.032) and use of instrumental social support (t=-
2.266, p<.026), while illness perception was the only significant predictor for adherence (t=-
1.963, p<.050). The percentages of variance explained by these predictors were 52.8% for self-
management and 31.5% for adherence.
Conclusions
Our results argue in favour of coping strategies and disease perception being important for
influencing self-management and respectively adherence in diabetic patients. These results
could serve in constructing better counselling and psychotherapeutic strategies in diabetes.
Keywords: diabetes, coping strategies, illness perceptions, optimism, adherence, self-management
250
Introduction
Diabetes is a problem of great public health importance, with an increasing prevalence in the
last decades [1] and potentially creating a considerable burden to affected individuals and
society [2].
Although diabetes is a chronic disease, an efficient self-management can significantly
improve its prognosis, by empowering the patient to be able to make informed decisions, to
know the behaviours necessary to conserve autonomy and to solve the problems that may arise
in within the disease [3]. In contrast to other medical conditions, where patient’s autonomy is
difficult or impossible to realize – this in turn potentially burdening the medical team [4] –,
self-management in diabetes is attainable and can have long standing benefits.
Another key parameter which is indicative for the prognosis of diabetic patients is adherence.
It is generally reported to have wide variations (from 36-85%) [5, 6] and may be influenced
by factors such as the patient’s understanding of the treatment, the perceived risk of adverse
effects, the costs and the complexity of the treatment scheme, although the repertoire of
adherence determinants is reported to be even larger [7].
Psychological variables may play a key role in the development of both self-management
and adherence in chronic diseases. Among them, functional coping strategies can stimulate a
proactive attitude and increased confidence in achieving the therapeutic goals [8], including in
diabetes [9, 10]. Illness perception may modulate adherence and self-management, especially
through dimensions such as timeline and control over the disease [7].
At their turn, affective states, such as optimism, are able to may mediate the relationships
between adaptive coping mechanisms, protective behaviours and motivation of the patient [11].
In the particular case of type II diabetes, many of the abovementioned relationships need
more substantial research, in order to better describe their involvement and importance. In this
sense, this study aims to assess the contributed played by illness perceptions, coping strategies
and optimism in the occurrence of self-management and adherence at diabetic patients.
Methods
The design of the study was cross-sectional and included 120 outpatients (52 men and 68
women) (age range 25-77), from a total of 169 who were offered the study questionnaires for
completion (response rate 71%). They were recruited between November 2018-June 2019 in
the National Institute of Diabetes, Nutrition and Metabolic Diseases – Bucharest. Inclusion
criteria comprised type 2 diabetes, age over 18 years, no associated psychiatric conditions, and
signing the written informed consent form about the participation in the study.
All participants completed the following standardized psychological instruments:
- Brief Illness Perception Questionnaire (BIPQ) [12]: comprises 9 items, from which five
measure cognitive illness representations, two the emotional components of the disease,
one the comprehension regarding illness, and another one the three most probable
factors assumed by the patient as causing the disease. High scores indicate that the
patient perceives the disease as more threatening. The questionnaire has been applied to
patients with various chronic diseases and has been reported to have a good validity
(Cronbach’s alpha .50-.70) [12, 13];
- Brief COPE [14]: contains 28 items measuring 14 coping strategies (self-distraction,
active coping, denial, substance use, use of emotional support, use of instrumental
support, behavioural disengagement, venting, positive reframing, planning, humour,
acceptance, religion, and self-blame). The questionnaire has a good validity
(Cronbach’s alpha .50-.90) [14];
251
- Life Orientation Test-Revised (LOT-R) [15]: contains 10 items, from which three assess
optimism, three assess pessimism, and the others serve as fillers. The questionnaire
score is directly proportional to the level of optimism. It has a good validity (Cronbach’s
alpha .74-.78) [16];
- Diabetes Self-Management Questionnaire (DSMQ) [17]: assesses self-care activities
associated with glucose control. It comprises 16 items grouped in four subscales –
Glucose management (GM), Dietary control (DC), Physical activity (PA), Health Care
Use (HU) – and a global scale. High scores of the sum scale show increased adherence.
The questionnaire has good validity (Cronbach’s alpha .60-.77 for the subscales and .84
for the sum scale) [17].
- Medication Adherence Report Scale (MARS) [18]: is an appropriate measure to detect
patients at risk of non-adherence. It comprises 9 items that rate the frequency with which
the patients engaged in the last month in non-adherent behaviour. Higher scores indicate
better therapeutic adherence. The questionnaire is reported to display good validity
(Cronbach’s alpha .85).
Statistical analysis was run using SPSS© 16.0 and included assessment of central tendency,
computing Pearson’s correlations and multiple linear regression. The statistical threshold of
significance was p<.05.
Results
In what concerned the individual perception of the disease, the highest negative impact was
represented by the duration of the disease (mean = 8.54), while the lowest was represented by
the identification of the disease (mean = 4.74). In the latter sense, one should notice the poor
perception of control over the disease (5.14) and the low perceived level of understanding
regarding it (5.60).
The top three preferred coping strategies were active coping (35%), planning (34.16%), and
acceptance (33.33%), while the least three were behavioural disengagement (83.33%),
substance use (1.66%), and denial (4.16%). Across the whole sample, the participants used
predominantly functional coping strategies to handle their diabetes.
The optimism levels across the whole sample was poor (19.12, range 8-30), with 83.33% of
all patients not reaching the recommended threshold of 24 points.
The mean global self-management score was acceptable (6.84, range 3.13-10), with the
highest mean sub score met at the ability to use medical services (7.52), followed by glucose
management (7.47) and physical activity (6.38).
The participants had a satisfactory adherence score (mean 40.63, from a maximum of 45).
In terms of coping strategies, functional (F) strategies correlated positively to adherence and
global self-management score, while dysfunctional (DF) strategies were placed oppositely. In
men, a much smaller number of coping strategies were significantly associated to dependent
variables, especially with adherence (Table 1).
252
Table 1. Significant correlations between coping strategies and the dependent variables
(adherence and global self-management score)
Global self-
Adherence
Coping strategies management score
Global Women Men Global Women Men
r -.024 -.095 .052 -.236** -.397** -.044
Self-distraction (DF)
Sig. (2-tailed) ns ns ns .009 .001 ns
r .202* .286* .148 .490** .435** .289*
Active coping (F)
Sig. (2-tailed) .027 .018 ns .001 .001 .038
r -.163 -.260* -.062 -.324** -.376** -.271
Denial (DF)
Sig. (2-tailed) ns .032 ns .001 .002 ns
Use of emotional support r .260** .255* .256 .165 .285* .017
(F) Sig. (2-tailed) .004 .035 ns ns .019 ns
Use of instrumental r .391** .470** .293* .236** .257* .210
support (F) Sig. (2-tailed) .001 .001 .035 .010 .034 ns
Behavioural r -.366** -.517** -.145 -.345** -.257* -.485**
disengagement (DF) Sig. (2-tailed) .001 .001 ns .001 .034 .001
r .251** .363** .141 .264** .327** .201
Planning (F)
Sig. (2-tailed) .006 .002 ns .004 .007 ns
r .358** .475** .206 .360** .309* .426**
Acceptance (F)
Sig. (2-tailed) .001 .001 ns .001 .010 .002
r -.200* -.269* -.097 -.278** -.259* -.312*
Self-blame (DF)
Sig. (2-tailed) .028 .026 ns .002 .033 .024
*
p<.05; **p<.01; ns = non-significant; the coping strategies not figured in the table did not significantly correlate
to any of the two dependent variables.
Regarding illness perception, both adherence and self-management scores correlated

negatively to the disease identity (r=-.207; p<.05), its perceived consequences (r=-.223; p<.05)
and the derived emotional response (r=-.192; p<.05). Optimism correlated, in both men and
women, to the ability to run physical activities (p<.003). Among demographic variables, age
had a positive correlation to global self-management score in both men (p<.02) and women
(p<.01), and to diet control in women (p<.02).
For the evaluation of the predictive effect of the independent variables on self-management
and adherence, a series of multiple linear regressions were performed. They included, in a first
step, coping strategies, in a second step, optimism and in a third step, illness perception.
In the final model, which included all independent variables (Table 2), active coping, use of
instrumental support and self-distraction were identified as the significant predictors for self-
management, while illness perception was the only significant predictor for adherence.
The percentage of variance explained by these predictors was 52.8% for self-management
and 31.5% for adherence.
Table 2. Predictors of self-management and adherence

Self-management Adherence
Predictors Unstandard.
t Sig. Unstandard. beta t Sig.
beta
(Constant) 5.195 .001 8.592 .001
Self-distraction (DF) -.212 -2.472 *
.032 .041 .100 .921
Active coping (F) .356 3.107* .002 -.556 -1.164 .247
Denial (DF) -.116 -1.319 .190 .078 .212 .833
Substance use (DF) .065 .644 .521 .135 .319 .750
Use of emotional support
.052 .528 .599 .607 1.481 .142
(F)
253
Use of instrumental support

-.288 -2.266* .026 .697 1.316 .191
(F)
Behavioural disengagement
-.154 -1.224 .224 -.807 -1.534 .128
(DF)
Venting (F) -.019 -.217 .828 -.328 -.876 .383
Positive reframing (F) -.010 -.100 .921 -.386 -.944 .347
Planning (F) .099 .753 .453 .331 .606 .546
Humor (F) -.054 -.722 .472 .211 .673 .502
Acceptance (F) .165 1.336 .185 .777 1.507 .135
Religion (F) -.008 -.116 .908 .048 .159 .874
Self-blame (DF) -.077 -.876 .383 .080 .217 .829
Optimism .133 1.396 .166 -.038 -.257 .798
Illness perception -.122 -1.231 .221 -.474 -1.963* .050
*
p<.05; all predictors included
Discussions
The results of our study point out the significant relationship between several coping
strategies (self-distraction, the use of instrumental social support and active coping) and the
ability of the diabetic patient to self-manage this chronic condition. In what concerns adherence,
this is more connected to the perception of diabetes itself, this leaving open the perspective of
continuously acknowledging and understanding the patient’s perception of their symptoms.
Still, in what concerns the specific group of male diabetics, the association between coping
strategies and adherence/self-management seems more unpredictable, this complicating their
possible prognosis.
Optimism was rather uncommon among the studied sample and proved to be poorly related
to both adherence and self-management. The same applies to demographic variables, with the
exception of age.
The above-mentioned results encourage the role of clinical psychologists in being part of the
multidisciplinary team handling diabetic patients. As both illness perceptions and coping
strategies can be objectives of psychological interventions, the assistance of these patients
should take into consideration targeting these variables for improving the patient’s quality of
life in diabetes.
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Cause or Complication? Psychosocial Aspects of Diabetic Patients

with Cardiovascular Diseases
PÁL Tünde1, PREG Zoltán1, SZABÓ Monica Iudita Maria1, NYULAS Kinga2,
BÁLINT SZENTENDREY Dalma1, GERMÁN-SALLÓ Márta1
1 “George Emil Palade” University of Medicine, Pharmacy, Science and Technology, Targu Mures (ROMANIA)
2 Gedeon Richter, Targu Mures (ROMANIA)
Emails: pal0tunde@gmail.com, preg_zoltan@hotmail.com, sztamo@gmail.com, kinga.nyulas@gedeon-richter.ro,
szentendreydalma@gmail.com, gsmarti66@yahoo.com
Abstract
Diabetes mellitus is a chronic illness with several long-term complications. The development
of psychosocial factors, as potential complications is usually neglected. The aim of this study
was the assessment of psychosocial risk factors in diabetic patients with cardiovascular diseases
using a standardized self-assessment questionnaire, as well as to raise awareness of these
conditions. We conducted a cross-sectional study with the enrolment of 480 patients. All
patients completed the standardized psychosocial questionnaire elaborated by the European
Society of Cardiology. In the study sample, the mean age was 67.69 years (SD 9.85 years, with
female predominance: 51.7%). Disturbances in glucose homeostasis were present in 49.5%,
33.3% of patients had type 2 diabetes mellitus. The most common psychosocial factors were
hostility, work and family stress, and low socio-economic status.
Amongst diabetic patients, the prevalence of type D personality was significantly higher,
whilst social isolation in non-diabetic patients. Depression was more common amongst diabetic
patients, but without significant difference. Patients with diabetes on oral anti-diabetic
medication confessed significantly more common depression and anxiety.
Psychosocial risk factors are common in diabetic patients with cardiovascular diseases. This
can have a contribution to diabetes risk, but also can enhance complications of cardiovascular
diseases in the presence of diabetes.
Keywords: diabetes mellitus, psychosocial factors, cardiovascular diseases, depression
Introduction
Currently, 463 million people, aged between 20 and 79 years are living with diabetes
worldwide, according to the newest data published last year by the International Diabetes
Federation (IDF) [1]. In Europe, the estimated prevalence is 59 million with the highest rates
in Germany. In Romania, based on studies from the last five years the prevalence of diabetes
was between five and seven million patients [2]. According to the World Health Organisation
(WHO) reports, diabetes is the fourth disease worldwide with the highest mortality, preceded
by cardiovascular diseases, cancer and respiratory diseases [3]. Dealing with type 1 and type 2
diabetes mellitus (T1DM, T2DM) is challenging for both patients and healthcare professionals.
A combination of genetic and environmental factors contributes to the development of
T2DM [4]. There is enough evidence to support that some of the psychosocial stress factors
(PsRFs) are associated with the development of the disease in healthy individuals [5]. The
influence of depression and low educational level on diabetes appearance is clearly
demonstrated [6]. The development of cardiovascular diseases (CVD) in diabetic patients is
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also higher in individuals with psychosocial characteristics [7]. An individual’s psychosocial

risk profile is built up from emotional factors and chronic stressors. Emotional factors include
depression, anxiety, hostility, type D personality, anger, and post-traumatic stress disorder
(PTSD) while chronic stressors low social support, low socioeconomic status (SES), work
stress, marital stress and caregiver strain are considered chronic stressors [8]. Probably the most
investigated PsRF is depression, which was associated with increased CVD morbidity in
diabetic patients [7].
Numerous studies emerged the idea that psychological factors trigger and lead to diabetes or
they have an impact on the disease severity and complications [9]. According to the American
Diabetes Association (ADA), diabetic patients should be assessed with standardized and
validated instruments in the terms of psychosocial factors at the very first visit and later
periodically, as well as when complications are detected or in case of treatment modifications.
This statement recommends the screening of diabetes distress, depression and anxiety
routinely [10].
In this study, we aimed to assess the psychosocial profile of diabetic patients with
cardiovascular diseases utilizing a standardized self-administered questionnaire elaborated by
the Prevention Working Group of the European Society of Cardiology (ESC) [11] and
recommended to be used in every cardiovascular patient. Another goal was to raise the
awareness of psychosocial stress factors in these individuals.
Subjects and Methods
We conducted a cross-sectional study at the Cardiovascular Rehabilitation Clinic of Târgu

Mureş. We enrolled 480 patients between December 2016 and November 2019. Apart from
clinical assessment in concordance with the individual’s disease profile, all participants
completed the standardized self-administered ESC psychosocial questionnaire [11] in their
mother tongue, Romanian or Hungarian. The test focuses on psychological and socioeconomic
characteristics which were found to have the most solid evidence concerning CVD. It has
nineteen items in nine topics: low socio-economic status (also one question regarded to the
educational degree), work and family stress, social isolation, depression, anxiety, hostility, type
D personality, post-traumatic stress disorder, and other mental disorders. Data input was
collected in Microsoft Word Excel database and data management and analyses were performed
with the use of SPSS v.20.0 software. The study was accepted by the Ethics Committee of the
Emergency County Clinical Hospital of Targu Mures. Participants signed a consent form to
participate in our research study.
Results
480 patients with different cardiovascular diseases were analysed. Mean age was 67.69 years
(ranged between 22 and 93 years with female predominance: 51.7%), more than half of the
patients came from urban areas, 27.5% of patients attended from five to eight classes and 23.1%
gymnasium. The study population characteristics are summarized in Table 1. Almost half of
the participants (49.5%) had disturbances in the glucose homeostasis, the most prevalent form
was type 2 diabetes (33.3%). Half of the diabetic patients were treated with oral anti-diabetic
agents.
257
Table 1. Study population characteristics N/Mean %/SD

Participants 480 100.00%
Age (y) 67.69 9.85
Sex
Female 248 51.7%
Male 232 48.3%
Heart rate (bpm) 72.23 15.01
Arterial hypertension 462 96.25%
Mean SBP/DBP (mmHg) 135/81 20/11
Hypercholesterolemia (> 5.17mmol/L) 141 35.25%
Total cholesterol (mmol/L) 6.14 0.8
Disturbances in glucose homeostasis 238 49.5%
T2DM 160 33.3%
Newly diagnosed T2DM 31 6.5%
T1DM 2 0.4%
Impaired fasting glucose 16 3.3%
Impaired glucose tolerance 29 6.0%
Treatment
Oral anti-diabetic medication (OAM) 49.4%
Insulin 25.9%
Nutrition therapy 13.0%
Mean BMI (kg/m2) 31.15 6.11
Obesity 264 55%
Mean BMI (kg/m2) 35.28 4.78
Comorbidities
Heart failure 238 49.5%
Coronary heart disease 197 41.04%
Atrial fibrillation 129 26.9%
Peripheral artery disease 84 17.5%
Cerebrovascular disease 70 14.5%
Chronic kidney disease (stages 2, 3, 4, 5) 44 9.2%
Chronic obstructive pulmonary disease 39 8.1%
Y: years; bpm: beats per minute; SBP: systolic blood pressure; DBP: diastolic blood pressure; T2DM: type 2
diabetes mellitus; T1DM: type 1 diabetes mellitus; OAM: oral anti-diabetic medication; BMI: body mass index.
The most common psychosocial factors were hostility, work and family stress, and low
socio-economic status. Amongst diabetic patients, the prevalence of type D personality was
significantly higher, whilst social isolation in non-diabetic patients. Depression was more
common amongst diabetic patients, but without significant difference (Table 2).
Table 2. Total sample Non-diabetic Diabetic pts P-value

Psychosocial risk factors %(n) pts %(n) %(n)
Low SES 63.3 (304) 64.5 (205) 61.1 (99) 0.102
Work & family stress 65.0 (312) 65.4 (208) 64.2 (104) 0.341
Social isolation 43.8 (210) 45.9 (145) 39.5 (64) 0.013
Depression 30.0 (144) 28.6 (91) 32.7 (53) 0.057
Anxiety 44.0 (211) 45.5 (143) 42.0 (68) 0.204
Hostility 65.4 (314) 65.7 (209) 64.8 (105) 0.692
Type D personality 53.1 (255) 51.3 (163) 56.8 (92) 0.009
PTSD 54.6 (262) 53.5 (170) 56.8 (92) 0.075
Other mental disorders 7.7 (37) 7.2 (23) 8.6 (14) 0.286
Pts: patients; SES: socioeconomic status; PTSD: post-traumatic stress disorder
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Patients with diabetes on OAM confessed significantly more common depression and
anxiety (Table 3).
Table 3. OAM %(n) Insulin treatment %(n) P-value

Psychosocial risk factors
Low SES 65.0 (52) 54.8 (23) 0.729
Work & family stress 70.0 (56) 54.8 (23) 0.622
Social isolation 40.0 (32) 35.7 (15) 0.565
Depression 33.8 (27) 23.8 (10) 0.039
Anxiety 41.3 (33) 33.3 (14) 0.030
Hostility 63.7 (51) 73.8 (31) 0.490
Type D personality 53.8 (43) 47.6 (20) 0.129
PTSD 53.8 (43) 54.8 (23) 0.601
Other mental disorders 11.3 (9) 7.1 (3) 0.675
Pts: patients; OAM: oral anti-diabetic medication; SES: socioeconomic status;
PTSD: post-traumatic stress disorder
Discussion
Diabetic patients have an increased risk of developing cardiovascular diseases. Evidence

suggests that traditional cardiovascular risk factors do not fully explain the high cardiovascular
mortality and morbidity in diabetic patients [5]. This finding is also supported by the fact that
interventional approaches in the reduction of cardiovascular risk, such as weight loss, blood
pressure and cholesterol control, in diabetic patients did not reach the expected results [5]. The
coexistence of PsRFs could represent the missing link. Vogelzangs et al., showed an association
between PsRFs, such as depression, anxiety, negative life events, inadequate emotional support
and metabolic syndrome, regardless of cardiovascular pathologies [12], which association was
previously demonstrated by numerous studies [13].
Although the impact of the individual’s psychosocial risk profile on diseases is accepted, its
assessment and management are still lacking. The ESC made efforts to overcome this problem
by introducing assessment strategies and explicit indications [11] in terms of psychosocial risk
factors. Also, the ADA has recognized the importance of this issue and through clear indications
facilitates the assessment of depression, anxiety and diabetes distress [10] in diabetic patients.
Based on the above-mentioned clues we demonstrated in our study, not only that
psychosocial risk factors are common amongst diabetic patients with cardiovascular diseases
but also that their assessment is feasible. Depression was found to be twice as common in
diabetic patients compared to the general population, with female predominance [9]. In
concordance with this, in our survey depression was more common amongst diabetic patients,
with marginal statistical significance. However, the control group does not fit the general
population label. Dealing with psychological disorders in diabetic patients can contribute to
behavioural and life-style anomalies, leading to resistance and disrespect of recommendations
for disease management, thus to uncontrolled glycaemia which results in diabetes-related
complications [14]. Diabetic patients who have depressive symptoms are at a higher risk of
developing macro- and microvascular complications [7] and vice versa, the presence of
macrovascular disease, chronic foot ulceration, proliferative retinopathy was associated with
increased prevalence of depressive symptoms [9].
Unfortunately, diabetes-related complications are scarce in this study. In parents whose child
was diagnosed with T1DM a higher rate of post-traumatic stress disorder (PTSD: determined
as negative emotions such as anxiety, fright, feeling helpless) was observed, compared to the
general population and it was confessed more common amongst mothers [15]. In a Swedish
259
longitudinal study, work stress was associated with the development of T2DM, however only
in female patients who had 60 years at baseline [16]. Chronic stress was associated with
numerous diseases, such as cardiovascular diseases [17, 18], cancer, and a recent review article
detailed the possible mechanism how stress plays a role in the development of T2DM, in disease
progression, as well as in patient adherence [5]. Despite of this, in our study sample, work and
family stress were more common amongst non-diabetic patients. Diabetic patients are prone to
feel anxiety, fear because of insulin administration or hypoglycaemia [10] and it was also
observed the omission of insulin injection due to anxiety [19] which leads to problems in
maintaining good glycaemic control [20]. Post-traumatic stress disorder is mentioned next to
anxiety in this patient group [10]. Our study showed a significantly higher prevalence of anxiety
and depression amongst diabetic patients on medication therapy. The origin of these factors is
difficult to explain as it was a cross-sectional survey.
Diabetes distress is a relatively new term and is used to reflect the emotional state rising
from living with diabetes and the burden of relentless daily self-care and the possible
complications. Patients with diabetic distress can feel stress, guilt or also denial [21] and it was
associated with poorer diabetes-related prognosis. According to recommendations of the ADA
its assessment is essential in diabetes care [10]. However, in this study, we did not evaluate this
factor.
Psychosocial risk factors not only could have a role in the development of diabetes or
influence cardiovascular outcomes in diabetic patients, but they have also a major role in
diabetic care and patient adherence to medication and recommendations. Low socioeconomic
status can hinder patients from the best treatment possibilities, according to Berkowitz et al.,
[22]. The DAWN study [23] (The Diabetes Attitudes, Wishes, and Needs) conducted in 13
countries (also countries from Europe, not including Romania) has pointed out among others,
the importance of the psychosocial aspects of life with diabetes. This study demonstrated, that
psychosocial interventions are needed in order to achieve better patient adherence and diabetic
self-care [23]. Once PsRFs are assessed, multidisciplinary team are warranted with initiation of
individual or psychotherapy, group and family therapy, cognitive behaviour therapy,
pharmacotherapy, recommendations about lifestyle management could be also taken into
account [9]. After the findings of the DAWN, further studies were conducted to assess and to
manage psychosocial factors in diabetes. Diabetes prevention exists, however, preventing
T1DM currently it is not possible [24], primary prevention measurements for T2DM are
feasible and effective, and proper diabetic care and monitoring are effective in the prevention
of diabetes-related complications [25]. As one size does not fit all, and some of the psychosocial
risk factors were related to the development of diabetes, maybe their management before the
diagnosis of diabetes would be reasonable.
The main limitation of the study could be the lack of information regarding glycaemic
control of diabetic patients. Due to the profile of our Clinics, obtaining HbA1c from patients in
order to assess glycaemic control was not feasible. Another limitation could be the absence of
information about diabetes-related complications.
Conclusions
We demonstrated that psychosocial factors are common in diabetic patients with

cardiovascular diseases. This can have a contribution to diabetes risk, but also can enhance
complications of cardiovascular diseases in the presence of diabetes, although it is difficult to
determine “which came first: the chicken or the egg”?
Our study shows that existing recommendations for evaluation of psychosocial stressors in
cardiovascular patients with diabetes can and should be implemented in daily practice.
260
Finally, we consider that future longitudinal studies are needed to clarify which are the best
care models for our patients.
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262
Mild Cognitive Impairment Is More Frequent in Middle Aged

Diabetics and Females
PÁL Tünde1, GERMÁN-SALLÓ Márta1, SZABÓ Monica Iudita Maria1,

NEMES-NAGY Enikő1, NYULAS Kinga2, BÁLINT SZENTENDREY Dalma1,
PREG Zoltán1
1 “George Emil Palade” University of Medicine, Pharmacy, Science and Technology, Targu Mures (ROMANIA)
2 Gedeon Richter, Targu Mures (ROMANIA)
Emails: pal0tunde@gmail.com, gsmarti66@yahoo.com, sztamo@gmail.com, nemesneniko@gmail.com,
kinga.nyulas@gedeon-richter.ro, szentendreydalma@gmail.com, preg_zoltan@hotmail.com
Abstract
Cognitive dysfunction and dementia represent two major concerns of the ageing society.
Amongst several conditions which were associated with the development and acceleration
of cognitive deterioration, diabetes mellitus and arterial hypertension seem to have the most
consistent role. We aimed to study the effect of diabetes mellitus on cognitive function in
hypertensive patients. The cross-sectional study included 428 patients – 181 patients with type
2 diabetes mellitus and 247 patients without, aged between 50-79 years. The detection of
cognitive impairment was performed with the Montreal Cognitive Assessment questionnaire
(MOCA) and the Mini Mental State Examination test (MMSE). We compared cognitive
performance scores of diabetic and non-diabetic patients from the same age group. For
statistical analysis, SPSS v.20.0 was used. The average age of the studied patients was 67.1+/-
7.4 years (51.9% female). Cognitive impairment (MOCA scores under 26 points) were present
in 65.2% of the studied patients, 60.7% in non-diabetic, and 71.3% in diabetic patients
(p=0.015). Cognitive dysfunction was more common amongst female patients and in the 50-59
years age group. Dementia (MMSE score under 24 points) was present in 15.9% of the
participants, 16.6% in non-diabetic, and 14.9% in diabetic patients (p=0.68). Cognitive
impairment is frequently present in diabetic patients, being more prevalent in middle-aged and
female patients. The presence of diabetes enhances neurocognitive decline in patients with high
blood pressure, especially with regard to mild cognitive impairment.
Keywords: diabetes, cognitive impairment, dementia
Introduction
Dementia is one of the most feared chronic diseases in ageing societies. Progressive
deterioration of cognitive functions has a huge impact on individuals, who gradually lose their
independence and may become socially isolated. Furthermore, dealing with neurocognitive
disability represents also an important economic burden which is constantly growing with the
increasing number of persons living with dementia. Thus, according to the recent World Health
Organization (WHO) data currently affects 50 million people and projected to triple by 2050
[1]. They are two main barriers to deal with this devastating illness. One is its subtle onset and
clinically silent evolution for a long term. The other is the lack of effective treatment. However,
an evolving body of literature documents the association of cognitive impairment and dementia
with modifiable risk factors like physical inactivity, smoking, bad eating habits, but also with
treatable conditions like hypertension, diabetes, hypercholesterolemia, obesity. These lifestyle-
263
related risk factors are also well-known hazards for cardiovascular diseases. Amongst these,
hypertension and diabetes mellitus seem to have the most consistent role in the development of
cognitive decline. It is important to keep in mind that in patients with dysglycaemia besides
cardiovascular health, brain health should be also our concern. A better understanding of the
relationship between diabetes and cognitive impairment could have a role in dementia risk
reduction.
The aim of our study was to investigate the consequences of diabetes mellitus on cognitive
function in hypertensive patients.
This cross-sectional study was conducted at the Târgu Mureș Cardiovascular Rehabilitation
Department. A total number of 428 patients were included – 181 patients had type 2 diabetes
mellitus, 247 patients without diabetes. Patients were aged between 50-79 years. All
participants underwent general physical examination, height, weight and blood pressure
measurement. We recorded demographic data, clinical data including personal history of
comorbidities and cardiovascular risk factors. Routine laboratory investigations (complete
blood count, glycaemia, full lipid profile, creatinine, uric acid, liver enzymes, and urine sample)
were done in every patient, as well as ECG. Other paraclinical investigations were completed
according to each patient’s disease profile.
Cognitive impairment was screened using the Montreal Cognitive Assessment questionnaire
(MOCA) and the Mini Mental State Examination test (MMSE) was used for dementia
screening. Cognitive impairment was considered under a MOCA score of 26 points,
respectively dementia was considered under a MMSE score of 24 points. We compared
cognitive performance scores of diabetic and non-diabetic patients from the same age group.
Microsoft Office Excel was used for data input and statistical software SPSS v.20.0 for data
management and analyses. Independent sample t-test was used for comparing means, the chi
square test was used for comparing categorical variables.
The Ethics Committee of the Emergency County Clinical Hospital of Targu Mures approved
the study and patients signed a consent form to participate in our research study.
Results
Average age of the studied patients was 67.1+/-7.4 years (51.9% female).
Table 1. Characteristics of the studied patient populations

Non-diabetic Diabetic p
(mean+/-SD) (mean+/-SD)
Age (years) 66.8+/-7.8 67.5+/-6.8 0.33
SBP (mmHg) 132.2+/-23.1 135.5+/-23.1 0.16
DBP (mmHg) 81.9+/-10.5 80.4+/-11.2 0.09
BMI (kg/m2) 29.9+/-5.1 34.0+/-5.3 0.002
Waist (cm) 100+/-16.7 107.9+/-16.4 0.001
eGFR 77.2+/-25.1 75.1+/-23.2 0.4
Uric acid (μmol/l) 253.4+/-142.7 282.1+/-134.1 0.035
Total Chol. (mmol/l) 4.86+/-1.24 4.45+/-1.18 0.002
LDL Chol. (mmol/l) 2.99+/-0.95 2.51+/-1.04 0.008
HDL Chol (mmol/l) 1.26+/-0.32 1.12+/-0.29 0.01
Triglyceride (mmol/l) 1.54+/-0.88 1.82+/-0.95 0.005
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Cognitive impairment (MOCA scores under 26 points) was present in 65.2% of the studied
patients, 60.7% in non-diabetic, and 71.3% in diabetic patients considered statistically
significant (chi-square test p=0.015). The difference in the prevalence of cognitive impairment
was more evident in females than in males, and in patients from the age group 50-59 years, and
less evident in other age groups – see Table 2.
Table 2. Frequency of cognitive impairment (MOCA score under 26 points) in the studied patients according
to age group and gender
Age group Non-diabetic% Diabetic% Chi-square test p
50-59 years 43.6% 74.1% 0.011
60-69 years 52.8% 58.9% 0.52
70-79 years 76.7% 81.5% 0.47
Sex
Female 59.4% 76.6% 0.009
Male 62.2% 65.5% 0.66
Dementia (MMSE score under 24 points) was present in 15.9% of the studied patients,
16.6% in non-diabetic, and 14.9% in diabetic patients, with no statistically significant difference
(p=0.68). There was no significant difference according to sex and age groups in the prevalence
of dementia in the studied patients.
Discussion
There is abundant evidence in the literature suggesting a strong link between diabetes
mellitus and cognitive impairment. In spite of this, the condition frequently remains
undiagnosed [2, 3]. In our study, mild cognitive impairment was demonstrated in around 70%
of the patients with diabetes and hypertension. Investigating individual’s cognitive deterioration
and especially the ethiology of this condition could be challenging, as patients with type 2
diabetes mellitus frequently have different comorbidities and factors which may have a negative
impact on cognitive abilities. Some of these are cardiovascular risk factors such as arterial
hypertension, dyslipidaemia, obesity, physical inactivity, cerebro- and cardiovascular diseases
[4], but also low socioeconomic status, depressive symptoms [5].
A longitudinal study performed in a Swedish population demonstrated that diabetes and even
prediabetes were independently related to accelerated deterioration of cognitive functions,
objectivized by the MMSE test. In this study researchers also corroborated their findings with
brain imaging, showing reduced global, mainly white matter volume already in the prediabetes
stage while diabetic patients had more white matter hyperintensities [6].
However, there are some other studies which came to a negative result, with respect to the
association of prediabetes to cognitive impairment [7].
There is enough evidence supporting the idea, that dysglycaemia increases the magnitude of
cognitive impairment attributed to other cardiovascular risk factors, making “the brain more
susceptible to other conditions of aging, such as Alzheimer’s disease (AD) or
neurodegeneration in general,” [8]. We demonstrated that mild cognitive impairment is more
common in a hypertensive diabetic population, compared to a non-diabetic hypertensive
population with same age and blood pressure values. A significant difference was noticed in
the middle-aged group.
265
The risk of dementia in diabetic patients is higher compared to non-diabetic patients and
diabetes increases the risk of vascular dementia. In contrast to this, the risk of AD is minor [9],
[10], according to previous studies. In our study, dementia prevalence, although it was much
higher compared to the general population [1] showed no difference whether diabetes was
present or not. This may suggest that clustering of the other cardio-metabolic risk factors, like
hypertension, obesity, high triglycerides and uric acid, low HDL cholesterol level might have
a higher influence on the appearance of dementia regardless of the presence of diabetes.
Gender differences in cognitive abilities are a highly investigated and unanswered topic.
Previous studies showed that the prevalence of AD is higher among females and they have
a higher risk of developing dementia [11]. Studies conducted in Europe and in the United States
have found no significant gender differences in cognitive abilities in contrary to studies
performed in low and middle-income countries, where it was found a significant female
predominance [12]. An experimental study by Sakata et al., found that female diabetic mice
had poorer cognitive functions compared to males [13]. We found significant sex-difference
with a female predominance. As our country is considered a middle-income country, we
consider that our findings are congruent with previous results.
The present study confirmed the findings of earlier cognitive impairment in diabetic patients.
We divided our study sample into different age groups (50-59; 60-69; 70-79 years).
In our study, the most vulnerable patients were those between 50-59 years. A Chinese study
analysing data from 8535 patients concluded that the worst cognitive abilities, measured as
episodic memory and executive function, were found in those patients who had untreated
diabetes in the age group of 45-59 years [14]. A German study came to a similar conclusion,
that mild cognitive impairment was twice more frequent in middle-aged (50-65 years) patients
with diabetes mellitus type 2 [15]. These results are in concordance with our findings in terms
of the age-group, however, we did not have enough information about diabetes control in the
study sample and most of our patients were treated with oral or injectable anti-diabetics, none
of them were untreated diabetes mellitus patients. Relation of untreated diabetes to cognitive
impairment is also supported by the longitudinal study of Yaffe et al., who revealed lower
cognitive scores in diabetic patients who had poorer glucose control assessed by higher HbA1C
[16].
Conclusions
Cognitive abilities are frequently affected in the presence of diabetes mellitus, which
enhances neurocognitive decline in patients with high blood pressure, especially with regard to
mild cognitive impairment. Our results suggest that middle-aged diabetics and females are more
vulnerable. Apparently, the presence of diabetes enhances neurocognitive decline in patients
with high blood pressure, especially in relation to mild cognitive impairment.
Furthermore, damaged cognition could also have an impact on patient-physician
communication and could affect patients’ compliance.
In conclusion, cognitive impairment should be screened in patients with diabetes mellitus
from at least the age of 50. This may present a unique opening for preventing or delaying
cognitive impairment and dementia.
Funding Acknowledgements
Funding for the study was provided by the Hungarian Academy of Science, contract
nr.0346/26.02.2016.
266
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9. van Duinkerken E, et al., (2019). Diabetes mellitus in the young and the old: Effects on cognitive
functioning across the life span. Neurobiol Dis 134, pp. 1-10.
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A systematic review. Can J Diabetes 41(1), pp. 114-119.
11. Gabelli C, et al., (2015). Gender differences in cognitive decline and Alzheimer’s disease. Ital J Gender-
Specific Med 1(6), pp. 21-28.
12. Singh PK, et al., Gender difference in cognitive health among older Indian adults: A cross-sectional
multilevel analysis. SSM – Popul Heal 5(3), pp. 180-187.
13. Sakata A, et al., (2010). Female exhibited severe cognitive impairment in type 2 diabetes mellitus mice.
Life Sci 86(17), pp. 638-645.
14. Zhang L, et al., (2019). Association between diabetes and cognitive function among people over 45 years
old in china: A cross-sectional study. Int J Environ Res Public Health 16(7), pp. 1-12
15. Angela W, et al., (2014). Association of diabetes mellitus and mild cognitive impairment in middle-aged
men and women. Journal of Alzheimer’s Disease, 42(4), pp. 1051-1069
16. Yaffe K, et al., (2012). Diabetes, glucose control and 9-year cognitive decline among non-demented older
adults. Arch Neurol. 69(9), pp. 1170-1175.
267
The Prevalence and Characteristics of Depression in Diabetes

Mellitus Patients. Is Routine Screening of Depression Necessary in
The DM Patients?
LEȚI Maria-Mădălina2, BODNĂRESCU-COBANOGLU Mihaela1,

DOBRESCU Iuliana2,3, POP Anca Lucia1,3, POPA Denisa1,
ROSU Isabella Andreea1, IORGUT Cristina1, ZETU Cornelia1
1 “N. C. Paulescu” National Institute of Diabetes, Nutrition and Metabolic Diseases, Bucharest (ROMANIA)
2 “Prof. Dr. Alexandru Obregia” Clinical Hospital of Psychiatry, Bucharest (ROMANIA)
Emails: ancapop@hotmail.com, mariamadalinaleti@gmail.com, mihaela_bdn@yahoo.com, iulianadobrescu@yahoo.com,

denissa.popa@yahoo.ro, isabellarosu@yahoo.ro, cryiorgut@yahoo.com, corapnc@yahoo.com
Abstract
Diabetes mellitus and depression represent chronic illnesses with major debilitating
consequences. Moreover, these two pathologies – one metabolic, one psychiatric – are
bidirectional interrelated, frequently undiagnosed and untreated [1]. To evaluate the prevalence
and characteristics of depression in diabetes mellitus (DM) patients we performed a hospital
based prospective observational clinical study on 70 patients with ages between 18-80 years
old, with no previously diagnosed depression, admitted in the Clinical Department of the
Diabetes and Metabolic Disease National Institute “N. C. Paulescu” in Bucharest during the
period of 2017-2018. In this purpose we performed the 17 points interview on depressed mood
using the Hamilton Depression Rating Scale (HAM-D) clinician-administered depression
assessment tool, during the usual clinical round evaluation. The severe and very severe
depression grade was present in the 18,6% of the DM patients, predominantly female gender,
with different degrees of obesity, from urban environment, with medium level of education,
single or in a stable relationship, retired, with a duration of 6-10 years of diabetes or over 20
years, that may have insulin therapy or oral antidiabetic therapy, with at least one microvascular
complication, that may or may not have diabetic retinopathy, chronic renal disease, and that
may have a PAD or a major comorbidity. Mild depression was present in 24.3% patients and
moderate depression in 21.4% of the group. Only 35.7% of patients had no active symptoms of
depression according to the HAM-D scale. This study is an initial stage in stepping up towards
the identification and diagnostic of psychological status of DM.
The depression scale Hamilton – D is a good tool for the primary evaluation of the patients
with diabetes mellitus during the periodical evaluations in the diabetes specialised clinics,
emphasising the need of evaluation during the routine check-ups of the DM patients and
immediate referral for intervention with pharmacological and non-pharmacological therapy in
depressed DM patients in order to improve the life quality of the diabetic patient, optimise the
self-care and antidiabetic therapy outcomes.
Keywords: diabetes, depression, Hamilton D, HAM-D, diabetes distress
Introduction
According to International Diabetes Federation (IDF) report in 2019, diabetes mellitus

(DM) is one of the most common chronic disorders, affecting more than 425 million people
268
worldwide in 2017 – with an estimated of almost 700 million cases in 2045 [1, 2]. New IDF
figures shows in present a continue increase in diabetes prevalence across the globe, with almost
half of billion people with diabetes, of which up to half being undiagnosed and more than 10%
of the health expenditure costs [3] that have caused an estimate of four million deaths
worldwide in 2017 [4], half of them young and active (under 60 years old), reiterating the need
for urgent action for prevention, early diagnose, identification of comorbidities and proper
intervention.
10,5%
9,8% 2045
9,3% 2030 700 milions
8,5% 2019 578 milions
4,7% 2014 463 milions
1980
Fig. 1. Dynamics of the global prevalence of Diabetes Mellitus (WHO, IDM 2019)
Of all DM cases worldwide, 59 million of diabetes patients live in Europe, with an estimated
growth percentage of 15%, up to 69 million cases in 2045, with the highest number of children
and adolescents (0-19 years) with type 1 diabetes (297,000) and one in six live births affected
by hyperglycaemia in pregnancy [4]. The number of DM patients in Romania is ranking from
over 810 000 people [5, 6] in 2014, to 1 785 000 patients (extrapolated) in 2019 of which 1 208
300 adults. One in 11 adults have diabetes mellitus and 20,7% are undiagnosed (IDF, 2019) [4].
Multiple comorbidities are interrelated with Type 1 and Type 2 diabetes mellitus:
cardiovascular disease – hypertension, acute myocardial infarction (AMI) and in over 22%
cases – heart failure [7,8]; cerebrovascular diseases – atherosclerosis and dyslipidaemia [9, 10],
stroke [11]. Poor management of DM leads to specific microvascular complications: diabetic
neuropathy, nephropathy, retinopathy; macro-vascular complications – peripheral artery
disease (PAD) [12], diabetic foot syndrome (DFS) and lower extremity amputation (LEA) [13,
14].
There is consistent evidence for the shared psychological and biological mechanisms for
depression and type 2 diabetes [15]. Comorbidity with anxiety and depression in DM are
bidirectional and mutually-reinforcing [16] the depression and anxiety was identified as a strong
predictor of cardiovascular outcomes [17, 18, 19, 20], associated with worsening of the quality
of life, impairment of the diabetes self-management, poor diabetes treatment adherence [21],
poor glycaemic control [22], ignorance of the healthy lifestyle recommendations (diet, physical
activity) [23], food intake disturbances, obesity [24], increased incidence of complications [25,
26], poorer survival and rehospitalisation (in patients with heart failure HF) and increased risk
of mortality [27] the outcomes of both conditions (depression and DM) being worsened by the
presence of the other and also a reduced life expectancy [28]. According to WHO, depression
is the first disability cause at the global level, with an estimate of approximate 350 million of
persons involved, with a 20 fold increased suicidal risk [29], having a high association with
chronically diseases like cardiovascular diseases (CVD), DM, Chronic Obstructive Pulmonary
Disease (COPD) and oncological diseases – and a decrease of the lifespan with 7-11 years [30].
Medication or psychotherapy can reduce depression among people with diabetes mellitus,
in addition to improving diabetes-related health (e.g., HbA1C) and DM treatment adherence
[27-29]. The investigation of the factors that contribute to anxiety and depression in patients
with diabetes, in order to identify the patients at risk can give a better opportunity to reduce the
risk of developing anxiety, depression and reduce the incidence of complications and
comorbidities related to DM. A recent study showed that one third of the patients with severe
depression may present overfeeding disturbances (depression with increased appetite) [31] also
269
showed that above obesity with all the possible metabolic and CVD consequences, depression
is linked to systemic inflammation and a trend toward higher CRP in the severe depressive
patients with increased appetite [32].
- EMOTIONAL TRAITS
- low self esteem
BEHAVIOURS - isolation
- sedentariness/lasiness - social inability/anxiety
- lack of social - chronic pain/fatigue BIOLOGY
interaction - indeciseveness - chronic inflammatory
- chaotic - lack of motivation syndrome
diet/semiprepared - high citokines
food/uncontrolled sugar - mood swings/apathy/depresion
intake -obesity, overweight
- unhealthy behaviour - low HDL
(smoking/binge → poor diabetes self - high LDL TG
drinking/substance medical/nutritional
abuse/anorexia/bulimia) - achantosis nigricans
management
- polychistic ovary sindrome
→ diabetes
complications
Fig. 2. Positive bidirectional correlation between diabetes and depression with negative prognostic (after
Diabetes co.uk, 2019)
Few studies have examined associations between depression and anxiety in diabetes
patients [20-23, 33], fewer studies have examined the utilization of treatment for mental
disorders in diabetics, a clear and attainable target for improving physical and mental patient
outcomes. More than that, lesser is investigated the diabetic distress (diabetes – specific distress
or diabetes – related distress), described as the distinctive emotional state residing in feelings
of worry, stress, frustration, guilt, denial, burnout [34] derived from the coping with diabetes –
affecting one in four type 1 DM, one in five insulin dependent diabetics and one in six OAD
(oral antidiabetic drugs) diabetic patients. Diabetic distress could negatively impact the
management of the disease and onset of complications and is best managed in the context of
diabetes care [35, 36, 37]. Such information could potentially identify important gaps in
diabetes care [38]. Proper recognition, diagnosis and management of the depression and distress
in diabetes mellitus patients (that are treated with non-pharmacologic and pharmacological
therapies) will improve the outcome both of the diabetes (self) care and the therapy compliance
and also will contribute to prevention of metabolic and depression related complications [39].
Less is known about the depressive – diabetic patients’ profile in Romania and the need
of the routine depression screening in the diabetic patients. Consequently, in this study we
analysed the statistical data gathered from The National Institute of Diabetes, Nutrition and
Metabolic Disease “N. C. Paulescu” Bucharest, Romania, gathering clinical data on 70 patients
diagnosed with diabetes mellitus with no previously diagnosed depression – for which we
performed an supplementary evaluation of the depression and anxiety (the anxiety was
presented in a previous paper) during the usual clinical round evaluation. The objective of the
present study was to assess the frequency and factors associated to diabetes mellitus in a cohort
of patients with diabetes mellitus.
270
In the present study we made a hospital based prospective observational clinical study on 70
patients successively admitted in the Clinical department of the Diabetes and Metabolic Disease
National Institute “N. C. Paulescu” in Bucharest during the period of 2017-2018.
Participants
We randomly selected a group of patients with type 1 and 2 diabetes mellitus (T2DM)
diagnosed at any point in their lives, or who had been prescribed diabetes medication during
the evaluation period, based on the order of presentation at the clinical hospital during the study
period, with no previously diagnosed depression. The patients were evaluated using the
Depression scale Hamilton D (Numerical Likert scale with 5 gravity levels) [21]; we collected
demographical, anthropometrical measurements; we performed screening of the diabetes
specific complications but also about associated comorbidities.
The inclusion/exclusion criteria

The inclusion criteria: presence of diabetes mellitus, age over 18 years old, admitted in the
clinic, on oral antidiabetic therapy, insulin or association of regimen – during the study period,
with no previously diagnosed depression. The patients with previous psychiatric treatment,
diagnosis of depression or other psychiatric diseases were excluded. The diagnostic status of
the evaluated subjects was known to the researchers as they were involved in recruitment,
screening and diagnosis, so the investigators were unable to be blinded to participants’
diagnostic status.
Study design
The study was done in conformity with the formal requirements for clinical observational
prospective study. All data was collected in accordance with the Declaration of Helsinki, with
all participants providing written informed consent to participate. For this study, we collected
data from the written and electronic medical records of the patients regarding following
parameters: anthropometric data (weight, height, body mass index – BMI), demographic data
(age, sex), parameters of glucose metabolism (glycaemia, HbA1C%), serum lipid profile (total
cholesterol, HDL-cholesterol, LDL-cholesterol, triglycerides), renal function panel (uric acid,
creatinine, glomerular filtration rate), blood pressure measurements (systolic and diastolic),
diabetes treatment (insulin, oral antidiabetics) and diabetes specific complications (retinopathy,
nephropathy, neuropathy). Routine laboratory tests were performed including fasting glucose
and glycated haemoglobin (HbA1c) for each patient.
The Hamilton scale for depression

The Hamilton Depression Rating Scale (also known as the HAM-D or HDRS) is the most
widely used clinician-administered depression assessment scale, commonly used for assessing
symptoms of depression – was applied according to the current guidelines [40], [21]. We
performed 17 points interview items on depressed mood (gloomy attitude, pessimism about the
future, feeling of sadness, tendency to weep), guilt, suicide, sleep disturbances, work and
interests, delay and retardation (slowness of thought, speech, and activity; apathy; stupor),
agitation (restlessness associated with anxiety), anxiety, psychic, and somatic (gastrointestinal,
indigestion, cardiovascular, palpitations, headaches, respiratory, genitourinary, other somatic
symptoms – heaviness in limbs, back, or head, diffuse backache; loss of energy and fatigability),
genital symptoms (loss of libido, menstrual disturbances), hypochondriasis, loss of weight,
insights, diurnal variation, depersonalization and derealisation, paranoid symptoms,
obsessional symptoms. [41]
271
A total score was calculated by summing the individual scores from each question, results
evaluated according to specifications in Table 1 [22, 42, 43, 44]. The interviews were done after
a previous HAM-D interview training done by psychiatrist, the time to perform an interview
varied between 15 and 20 minutes and the patients were kindly guided during the interview,
with respect to personal intimacy.
Table 5. The Hamilton Depression Rating Scale grades

Grade HAM-D total score Depression
Grade 0 <7 points Normal/Absent/remitted
Grade I 8-13 Mild or trivial depression
Grade II 14-18 Moderate depression
Grade III 19-22 Severe depression
Grade IV More or equal to 23 Very severe depression
Statistical analyses
Data are presented as mean ± standard deviation (±SD). Clinical characteristics were
compared using the t Student Test. Pearson’s and Spearman’s correlation coefficients were
calculated to evaluate relationships between variables. Significance was defined at the 0.05
level of confidence. All calculations were performed using the Statistical Package for Social
Sciences Software (SPSS) version 21.0.
Results
Demographics of the studied group

In the group of 70 patients included in the study 15.7% (n=11) had type 1 Diabetes Mellitus
(DMT1) and 84.3% (n=59) type 2 DM (with an almost equal distribution among sexes – 52.9%
males (n=37) and 47.1% woman (n=33). The median age was 57.3+/-13 years, with a median
of 58 years old, distribution area R-38.6% (27) U-61.4% (43). According to the Hamilton D17
scale (Table 1) 35.7% of the patients (n=25) had no active depression; 64.3% presented overall
scores of depressions (n=45).
Presence of depression in the studied group

The majority of patients were on the mild depression category (24.3%, n=17) with scores
under 14 points on the HAM-D scale, the moderate depression was observed in 15 patients
(21.4%), severe depression in 6 patients (8.6%) and very severe depression in 10% of the
studied (n=7) with scores of over 23 points on the HAM-D scale. From the group of patients
with depression (64.3%, n=45) 17.8% had type 1 DM and 82.2% (n=37) had type 2 DM. The
subgroup of patients with depression had a mean age of 57.9±12.8 yrs. old, predominantly from
urban area: n=27(60%), mostly female gender (64.4%) compared with men (35.6%).
Education level: low (no education or finished education up to middle school)-14 (31.1%);
moderate (high school) – 24(53.3%); high (university) – 7 (15.6%). Occupation: unemployed
– 8 (17.8%); employed – 11 (24.4%); retired – 26(57.8%), Marital status: Single -13 (28.9%),
in a stable relationship – 32(71.1%).
Diabetes duration: mean duration of diabetes in patients with depression subgroup was
12.9±8.6 yrs. There has been observed an equal higher incidence of depression in 2 groups: 6-
10 years and over 20 years duration of diabetes as highlighted in Table 2.
272
Table 6. Diabetes duration in the depression subgroup (n=45)

Duration (years) Frequency Percent
<1 year 7 15.6%
1-5 years 4 8.9%
6-10 years 11 24.4%
11-15 years 7 15.6%
16-20 years 5 11.1%
>20 years 11 24.4%
Prevalence of obesity in the diabetes-depression group

Normal weight (assed by BMI) was observed only in 10 (22.2%) of patients with diabetes
and depression. 44.5% of the depression sub-group were obese in different grades (Table 3) and
33.3% overweight.
Grade III
obesity Normal
Grade II obesity weight
16%
Grade I overweight
obesity 33%
25%
Fig. 3. The prevalence of obesity in the diabetes-depression sub-group
Diabetes treatment: most of the patients – 19 (42.2%), in the diabetes – depression subgroup
was on the mixed antidiabetic regimen with both OAD and insulin with 14 patients (31.1%) on
OAD regimen and n=12(26.7%) depressive diabetic patients on insulin regimen.
Number of severe comorbidities: over half (53.3%, n=24) of patients with diabetes and
depression had at least one severe comorbidity (cancer, liver cirrhosis or CKD).
Diabetes complications in the diabetes depression subgroup: 84.4% of the diabetic with
depression patients had at least one microvascular complication, with 28.9% with two
complications and 17.8% with three coexisting microvascular complications.
Table 7. Number of associated microvascular complications

Microvascular complications Frequency Percentage Cumulative percent
0 7 15.6 15.6
1 17 37.8 53.3
2 13 28.9 82.2
3 8 17.8 100.0
Total 45 100.0
Chronic kidney disease: 42.2% of diabetes – depression patients group had chronic kidney
disease, 57.8% being without CKD (n=26).
273
proliferative
9%
nonproliferative
29% absent
Fig. 4. Diabetic retinopathy in diabetes-depression group
Diabetic neuropathy: most of the diabetic-depression group had distal symmetric

polyneuropathy (DSPN) (16, 35.6%), 14 (31.1%) both (autonomous and DSPN) and 33.3%
(15) had no diabetic neuropathy, DSPN being the most common chronic diabetes complication
according to the American Diabetes Association (ADA). There were no significant differences
between the age of the diabetes depression group compared to non-depressed diabetic group
according to the HAM-D scale (rs=0.04, p=0.7). The degree of depression was not statistically
significant correlated with the age group (rs=0.046, p=0.7).
We observed strong positive correlation of the depression with the female gender, with the
more severe depression stages (rs=0.526, p<0.0001). Interestingly, the shorter the stature of
the patients were the higher the depression association (rs=-0,239, p=0.04). The patients in a
stable relationship had lower depression grades compared to single patients (weak statistical
significance rs=-0.227, p=0.05). Depression grade had a strong negative correlation with the
level of education, the depression grade is higher on the lower level of education patients (rs=-
0,239, p=0.04).
From the total 47,1% of prevalence of peripheral arterial occlusive disease (PAD) prevalence
in the study group, in the DD subgroup 35,5% had PAD, the depression grade, the higher the
prevalence of PAOD (rs=-0.315, p=0.008) (with no analysis on the AOMI grades).
Also, depression grade had a strong positive correlation with anxiety (measured on HAM-A
anxiety scale) (rs=0,814**, p<0.0001). The oral antidiabetic treatment was negatively
correlated with the degree of depression (rs=-0.301*, p=0.01), the oral treatment having a less
negative impact on the psychological status, with no statistical correlation being observed in
our study with the insulin therapy (rs=0.209, p=0.08).
We consider one important study limitation the fact that we performed the study on a limited
sample size (n=70) and without a control group. In the present study we didn’t evaluated the
diabetic distress, antidepressive or other psychiatric interventions as well as association with
complementary therapies. Certain associations were of borderline statistical significance and a
larger study would allow further exploration.
This was an inaugural study done in the National Institute of Diabetes and Metabolic Disease
“N.C. Paulescu”, done together with a study on anxiety on diabetic patients admitted in the
clinic. The HAM-D scale can be a reliable initial tool for the diabetology clinician to identify
the DM patient at risk for developing depressive pathology – that can undergo undiagnosed and
so, to reveal the psychological suffering using a simple scale, available also in the app format
[42]. This scale can be used by a psychologist/psychiatrist or a licentiates diabetes specialist in
order to improve the quality of medical services and promote interdisciplinarity, to direct the
right patients to psychological of psychiatrically support and treatment in order to increase
compliance to diabetes treatment, improve the metabolic outcome, the self-care and lower the
complications rates.
274
Conclusions
In the studied group the severe and very severe depression grade were present in the 18,6%
DM patients predominantly female gender, with different degrees of obesity, from urban
environment, with medium level of education, single or in a stable relationship, retired, with a
duration of 6-10 years of diabetes or over 20 years, that may have insulin therapy or oral
antidiabetic therapy, with at least one microvascular complication, that may or may not have
diabetic retinopathy, chronic renal disease, and that may have a PAD or a major comorbidity.
This study is an initial stage in stepping up towards the identification and diagnostic of
psychological status of DM. The depression scale Hamilton – D is a good tool for the primary
evaluation of the patients with diabetes mellitus during the periodical evaluations in the diabetes
specialised clinics in collaboration with psychological/psychiatrically services.
With a prevalence of total 18,6% of severe and very severe depression in the studied group
during a six-month period, we emphasise the need of depression evaluation in DM patients
during the routine check-ups or annually using a simple interview scale, immediate referral for
intervention with pharmacological and non-pharmacological therapy in depressed patients.
Despite the multiple barriers in screening and treatment of depression and distress in DM,
implementing strategies to inform and train the healthcare providers, inform the patients, screen
and refer the patient to support services and by so, improve the life quality of the diabetic
patient, optimise the self-care and antidiabetic therapy outcomes. Perspectives: evaluation of
diabetes distress comparative to diabetic depression in order to correctly address the need for
further psychological/psychiatric evaluation and intervention.
Acknowledgments
We thank to the management of the clinic that allowed the performance of the study. The
study was performed with no financial burden.
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277
Diabetes Mellitus and Pregnancy
RADU Mihaela Corina1,2, BOERU Adrian Calin1, MARIN Mihaela Liliana1,

MANOLESCU Loredana Sabina Cornelia2
1Hospital of Obstetrics and Gynaecology, Ploiesti (ROMANIA)
Email: radu_mihaela72@yahoo.com
Abstract
Diabetes interacts significantly with the pregnancy, being able to generate complications
with important effects on maternal health, both immediate and subsequent to pregnancy. Our
objective is to evaluate the non-genetic risk factors of diabetes mellitus in pregnant women.
This may lead to an easier identification of pregnant women at risk of diabetes mellitus and
an earlier diabetes mellitus diagnosis, but also in establishing measures to minimize the effects
of the disease on pregnancy and the unborn baby.
A retrospective study was carried out during 2017-2019 period, on a total of 84 pregnant
women who gave birth at Ploiesti Gynaecological Obstetrics Hospital.
We divided the women in two equal groups and analysed them by age, socio-demographic
characteristics, education, risk factors for diabetes mellitus, number of pregnancies,
complications at birth.
The non-genetic risk factors associated with diabetes in pregnancy noticed in our study were;
age over 30 years old, smoking status and higher BMI.
The genetic predisposition of gestational diabetes is confirmed by the results obtained in the
study. Pregnancy complicated by diabetes increases the rate of birth by caesarean section.
Pregnancy with gestational diabetes increases the risk of neonatal metabolic disorders.
Complicated pregnancy with diabetes with poor glycaemic control increases the risk of
postpartum neonatal complications, other than metabolic disorders. More than half of new-
borns born from complicated pregnancies with diabetes have suffered postpartum
complications.
Keywords: diabetes; pregnancy; non-genetic risk factors; gestational diabetes
Introduction
Diabetes mellitus is defined as a group of metabolism diseases characterized mainly by

hyperglycaemia induced by insulin deficient secretion, insulin resistance or both entities to
varying proportions [1].
Chronic hyperglycaemia of diabetics is associated with long-term impairment, dysfunction
and insufficiency of various organs, especially the eyes, kidneys, nerves, heart and blood
vessels. Several pathological processes are involved in the evolution of diabetes, ranging from
autoimmune destruction of beta pancreatic cells that induce insulin deficiency to abnormalities
that cause insulin resistance. Diabetes has become a major issue for the individual, medicine
and society. This is because it is “a common, treacherous, long-lasting and devastating disease
if not well taken care of” [2].
The alarming increase in the number of diabetes patients in the world is already a certainty.
This situation does not evade Romania either [3].
The latest data released by experts provides an alarming prognosis for diabetes mellitus.
278
The latest report of International Diabetes Federation from 2019 reveals that 463 million
people are living with diabetes worldwide [2]. This estimate exceeds all the previous estimates
regarding the number of subjects with diabetes. Almost half of the persons with diabetes remain
undiagnosed. Romania is considered among the top 10 countries in Europe with estimated
persons with diabetes. According to the same data, the number of subjects with diabetes will
overrun 1.7 million by the year 2030 [2]. In Europe in 2017 there were 66 million cases of
diabetes mellitus and in Romania in the same year were registered 1.785.300 cases [4].
However, the exact number of people with diabetes in Romania is still unknown. Despite
several attempts, a National Registry of persons with diabetes could not be yet established in
Romania. Therefore, it is difficult to know the real number of subjects with diabetes. The
growth will occur especially in developing countries.
It is considered that the increased incidence of type 2 diabetes mellitus is one of the worst
situations in human history. The epidemiological dynamics of diabetes also results from the
data of the PREDATORR (PREvalence of DiAbeTes mellitus, prediabetes, overweight,
Obesity, dyslipidaemia, hyperuricemia and chronic kidney disease in Romania) study. This is
the first national study analysing the prevalence of diabetes mellitus (DM) and prediabetes, and
their association with cardiometabolic, sociodemographic, and lifestyle risk factors in the
Romanian population aged 20-79 years. In all, 2728 participants from 101 clinics of general
practitioners were investigated. The overall age- and sex-adjusted prevalence of DM was 11.6%
(95% CI 9.6%-13.6%), of which 2.4% (95% CI 1.7%-3.1%) had unknown DM. The prevalence
of DM increased with age and was higher in men than in women. The age- and sex-adjusted
prevalence of prediabetes was 16.5% (95%CI 14.8%-18.2%), with the highest percentage in
the 60-79-year age group and in women. Obesity, abdominal obesity, dyslipidaemia, low
education level, and a family history of diabetes were associated with glucose metabolism
disorders [3].
The data published by the National Centre for Studies of Family Medicine regarding the
Project “Sentinel Medical Units Network” MEDINET (computerized family medicine research
network) attests that the prevalence of diabetes in the studied group is 0.14% for type 1 diabetes
mellitus and 1.09 % for type 2 diabetes mellitus (compared to 1.8% in the general population)
[5]. The same source indicates that diabetes is the 9th frequent diagnose in general and the 14th
most frequent new diagnose in the family doctor practice. The prevalence of chronic
complications and the conditions associated with diabetes mellitus is impressive. The most
common chronic complications are retinopathy, neuropathy and nephropathy, found in 1/3 of
the population affected by diabetes [3].
The endocrine changes that occur during pregnancy are of a complexity and of an unexpected
magnitude. In the production and metabolism of hormones the placenta plays an essential role.
Women with pregestational type 1 and type 2 diabetes mellitus and gestational diabetes
continue to have poorer pregnancy outcomes than the background population, including a three-
to fourfold higher rate of perinatal mortality [6]. These women can develop a whole range of
complication due to diabetes.
The influence of pregnancy on diabetes can be characterized by the fact that pregnancy is a
diabetic condition with multiple hormonal and platelets factors with diabetic effect. It is a
proved fact that pregnancy worsens pre-existing diabetes. The most representative
complications are: hyperketonaemia with ketoacidosis and hypoglycaemia [7]. In Europe data
showed in the report of International Diabetes Federation from 2019 reveals that the risk for
diabetes in women with prior gestational diabetes mellitus is 11.5 (CI:7.5-17.6) [2].
279
Objective
Diabetes interacts significantly with the pregnancy, being able to generate complications
with important effects on maternal health, both immediate and subsequent to pregnancy.
Diabetes mellitus represents, along with high blood pressure, the most common health
problems in pregnancy.
Our objective is to evaluate the non-genetic risk factors of diabetes mellitus in pregnant
women. This may lead to an easier identification of pregnant women at risk of diabetes mellitus
and an earlier diabetes mellitus diagnosis, but also in establishing measures to minimize the
effects of the disease on pregnancy and the unborn baby.
Material and Method
A retrospective study was carried out during 2017-2019 period, on a sample of 84 pregnant
women who gave birth at Ploiesti Gynaecological Obstetrics Hospital. The study was carried
out in compliance with the rules of medical ethics, the patients expressing informed consent
regarding the use of their data from the observation sheets, the use of the results of the clinical
and laboratory investigations.
The eighty-four pregnant women were divided into two groups, namely:
- Group A, consisting of 42 pregnant women with diabetes mellitus (any form)
- Group B, consisting of 42 pregnant women without diabetes
We analysed the groups by age, socio-demographic characteristics, education, risk factors
for diabetes mellitus, number of pregnancies, complications at birth.
Within group A, of 42 pregnant women, 8 (19.04%) had pre-existing type I diabetes, 11

pregnant women (26.19%) had pre-existing type II diabetes and 23 (54.76%) pregnant women
have been diagnosed with gestational diabetes. The increased prevalence of gestational diabetes
controlled only by changing lifestyle, diet and physical activity is in agreement with the
literature data and we concluded that this is the first line of treatment in balancing the glycaemic
ratio in pregnant women diagnosed with gestational diabetes.
The patients with diabetes were predominantly in the age range between 31 and 35 years,
with an average of 30.83 years, compared with the pregnant women without diabetes where the
age range was between 26 and 30 years with an average of 28.80 years (Table 1).
Table 1. Distribution of pregnant women by age

Age Under 20 21-25 years 26-30 years 31-35 years 36-40 years over 40
years years
Lot A/42 8(19.04%) 6(14.28%) 5 (11.90%) 17(40.47%) 4(9.52%) 2(4.76%)
Lot B/42 7(16.66%) 8(19.04%) 19(45.23%) 4(9.52%) 3/42(7.14%) 1(2.28%)
The data obtained in our study confirms the data from the specialized literature according to
which the age over 30 is an important risk factor for the development of gestational diabetes
[8-14].
Women who have a job that requires them a lot, is stressful and has a lot of work, are at risk
and are 21% more likely to develop type 2 diabetes, according to researchers from Gothenburg
University. The results of our analysis show that 59.52% (N=25) of pregnant women diagnosed
with diabetes claim that work is stressful [8].
280
Table 2. Distribution of pregnant women by occupation

Occupation Student Unemployed Employed Stressful work
Lot A 4/42(9.52%) 9/42(21.42%) 29/42(69.04%) 25/42(59.52%)
Lot B 8/42(19.04%) 13/42(30.95%) 21/42(50.00%) 16/42(38.09%)
Our study shows that the prevalence of diabetes is higher in urban areas than in rural areas.
This would be a consequence not only of a better detection and rapid diagnosis in the urban
area, but also a reflection of the more stressful and thrilling city life, associated with the greater
presence in the city of some risk factors: such as obesity and sedentary lifestyle along with
inadequate diet (Table 3).
Table 3. Distribution of pregnant women by place of birth

Place of birth Rural Urban
Lot A 15/42 (35.71%) 27/42 (64.28%)
Lot B 12/42 (28.57%) 30/42 (71.42%)
These percentages show that, although the people who attended the study graduated from
high school and lived in the urban area, they still had problems in accessing medical services
and administering treatments, due to the low level of education. (Table 4) This is worrying,
considering that diabetes is one of the most psychologically demanding chronic diseases, as
almost 95% of its management is performed by the patient. Thus, patients with low levels of
education have less knowledge about effective disease management and are not sufficiently
prepared to properly manage their disease. All of this ultimately leads to higher costs in the
health care system, as people with poor disease management get to use more medical services,
fail to control their medication and cannot follow the prescribed treatment properly [8].
Table 4. Distribution of pregnant women by education

Education No school education High school education College education
Lot A 12/42(28.57%) 23/42(54.76%) 7/42(16.66%)
Lot B 15/42(35.71%) 14(33.33%) 11/42(26.19%)
Studies have shown the existence of nicotinic receptors in the pancreatic beta cells
responsible for insulin secretion [9]. Thus, nicotine may reduce insulin release through these
receptors. There are also animal studies that have shown that exposure to nicotine in the pre
and neonatal period can lead to abnormal pancreatic beta-cell function and even apoptosis (cell
death). In patients with diabetes that smoke it is more difficult to obtain metabolic control,
requiring higher doses of insulin compared to non-smokers [10]. The results obtained in this
analysis showed that 80.95% (N=34) of pregnant women diagnosed with diabetes are smokers
(Table 5).
Table 5. Distribution of pregnant women by smoking habits

Smoking habits Smokers Non-smokers
Lot A 34/42 (80.95%) 8/42 (19.04%)
Lot B 17/42 (40.47%) 25/42 (59.52%)
The results obtained in this analysis, which took into account the history of diabetes of the
first degree relatives, showed us that 52.38% (N=22) of the pregnant women diagnosed with
diabetes in the current pregnancy have at least one relative of the 1st degree with diabetes, as
opposed to 9.52% (N=4), the percentage of the pregnant women in group B who have at least
one relative of the 1st degree with diabetes. This result confirms the data from the specialized
literature. The genetic predisposition for this disease is confirmed in other studies, the genetic
factor being a risk factor for the development of diabetes in pregnancy [11-12] (Table 6).
281
Table 6. Distribution of pregnant women by heredoc lateral background for diabetes

Heredoc lateral background for Present Absent
diabetes
Lot A 22/42 (52.38%) 20/42 (47.61%)
Lot B 4/42 (9.52%) 38/42 (90.47%)
In the literature there is a controversy regarding the number of pregnancies and diabetes,
some studies claiming that the large number of pregnancies and multi-parity could be risk
factors for gestational diabetes, other studies reject this hypothesis [13-15].
In the present study there was no correlation between the increased number of pregnancies,
multi-parity and increased risk for developing gestational diabetes (Table 7 and 8).
Table 7. Distribution of pregnant women by parity

parity (P) 1P 2P 3P 4P 5P Over 5P
Lot A/42 18(42.85%) 9(21.42%) 4(9.52%) 3(7.14%) 5(11.90%) 3(7.14%)
Lot B/42 16(38.09%) 8(19.04%) 6(14.28%) 5(11.90%) 4(9.52%) 3(7.14%)
Table 8. Distribution of pregnant women by gestation

Gestation 1G 2G 3G 4G 5G Over 5G
(G)
Lot A/42 17(40.47%) 9(21.42%) 5(11.90%) 3 (7.14%) 6(14.28%) 2(4.76%)
Lot B/42 14(33.33%) 7(16.66%) 7(16.66%) 5(11.90%) 5(11.90%) 4(9.52%)
In the present study it is observed that in pregnant women with diabetes, BMI has higher
values, which is in agreement with studies from other university centres and with data from
several meta-analyses that revealed that the risk of developing gestational diabetes is two to
eight times higher in overweight and obese pregnant women with high body mass index [16]
(Table 9).
Table 9. Distribution of pregnant women by BMI

BMI ≤18.5 18.6-25 25.1-30 30.1-40 >40
Lot A 0(0.00%) 9(21.42%) 23(54.76%) 6(14.28%) 4(9.52%)
Lot B 3(7.14%) 28(66.66%) 5(11.90%) 4(9.52%) 2(4.76%)
Analysing the two groups we found an increased incidence of diabetes in women that gave
birth by C-section, 80.95% (N=34) compared to 19.04% (N=8) in the group of pregnant women
without diabetes. This analysis confirms current data showing that caesarean delivery is
frequently encountered in the pregnant population with diabetes. It is known that C-section is
recommended more frequent in other cases, such as infectious diseases that may be transmitted
to the foetus or new-born [17-20]. Although in childbearing women with diabetes mellitus the
recommendations of the clinical guidelines are vaginal birth and diabetes itself is not an
indication of caesarean delivery, the rate of caesarean births is increasing due to the
complications caused by diabetes in pregnancy (Table 10).
Table 10. Distribution of pregnant women by birth mode

Birth mode C-section Vaginal birth
Lot A 34/42(80.95%) 8/42 (19.04%)
Lot B 8/42 (19.04%) 30/42 (71.42%)
In the analysis of postpartum complications, infectious complications, postoperative wound

pathology, postpartum haemorrhagic episodes were considered. The results showed that the
number of postpartum complications was higher in the group of pregnant women with diabetes
compared to the control group (Table 11).
282
Table 11. Distribution of pregnant women by post-partum complications

With/without complications Yes No
Lot A 33/42(78.57%) 9/42(21.42%)
Lot B 12/42(28.57%) 30/42(71.42%)
Table 12. Distribution of pregnant women by complications in pregnancy

Lot A 25/42(59.52%) 7/42(16.66%)
Lot B 11/42(26.19%) 31/42(73.80%)
The analysis of the APGAR score showed that new-borns from complicated pregnancies
with diabetes obtained lower APGAR scores than new-borns from mothers whose pregnancies
were not complicated with diabetes, also APGAR scores with values of APGAR 5 were present
only in the group of pregnant women with diabetes in pregnancy (Table 13).
Table 13. Distribution of pregnant women by Apgar score of new-borns

Apgar 10 9 8 7 6 5 Under 5
Lot A/42 4 (9.52%) 6(14.28%) 14(33.33%) 12(28.57%) 2(4.76) 4(9.52%) 0
Lot B/42 14 (33.33%) 24 (57.14%) 2(4.76%) 2 (4.76%) 0 0 0
The analysis of the groups according to the weight of the new-borns at birth showed that the
new-borns from the mothers with diabetes were overweight; the results confirmed that diabetes
is an important risk factor for fetal macrosomia (Table 14).
Table 14. Distribution of pregnant women by new born weight

New-born weight Under 2500 2500-2990 3000-3490 3500-3990 4000-4490 Over 4500
grams grams grams grams grams grams
Lot A/42 1(2.38%) 2(4.76%) 17(40.47%) 13(30.95%) 8(19.04%) 1(2.38%)
Lot B/42 3(7.14%) 6(14.28%) 24(57.14%) 8(57.14%) 1(2.38%) 0
Carbohydrate metabolism disorders of the new-born that were analysed were hypoglycaemia
and hyperglycaemia. The obtained results show that 54.76% (N=23) of the new-borns of
mothers with gestational diabetes suffered these complications compared with 11.90% (N=5)
new born babies from mothers without gestational diabetes that suffered metabolic
complications (Table 15).
Table 15. Distribution of pregnant women by metabolism disorder of the new-born

Metabolism disorders Yes No
Lot A 23/42(54.76%) 19/42(45.23%)
Lot B 5/42 (11.90%) 37/42(88.09%)
The frequency of new-borns without complications is higher in group B (61.90% vs.

28.57%). When analysing the two groups, other complications of the new-born, other than the
metabolic disorders, were considered. Complications analysed were respiratory distress
syndrome, obstetric trauma, congenital malformations, and postpartum new-born infectious
syndrome. As a result, we found that new-born babies from mothers without diabetes had a
smaller number of complications (Table 16).
Table 16. Distribution of pregnant women by complications of the new-born

Lot A 30/42(71.42%) 12/42(28.57%)
Lot B 16/42(38.09%) 26/42(61.90%)
283
Conclusions
The non-genetic risk factors associated with diabetes in pregnancy noticed in our study were
age over 30 years old, smoking habits and higher BMI.
The genetic predisposition of gestational diabetes is confirmed by the results obtained in the
study. Pregnancy complicated by diabetes increases the rate of birth by caesarean section.
Pregnancy with gestational diabetes increased the risk of neonatal metabolic disorders.
Complicated pregnancy with diabetes with poor glycaemic control increased the risk of
postpartum neonatal complications, other than metabolic disorders. More than half of new-
borns born from complicated pregnancies with diabetes have suffered postpartum
complications.
Rapid change in lifestyle is one of the factors that have a major impact on the onset of
diabetes, obesity or cardiovascular disease. Adopting measures to control the impact of
environmental/social factors on the communities we live in today becomes an effective means
of action for primary and secondary diabetes prevention. Health education should be one of the
areas where the impact of environmental and societal factors is diminished. This condition
(diabetes) is largely generated by the society and not by the health systems and for this reason
the solution must be found at the level of the communities or society.
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285
Incidence and Severity of Sudomotor Dysfunction in Early Stages

of Sensorimotor Neuropathy in Type 2 Diabetes
DOROS Rodica1, STEGARU Daniela1, IACOBINI Andra Evelyn1, MOTOC Razvan1,

BRAILEANU Bogdana1, ANDREI Catalina1, CACEAUNE Elena1, ELIAN Viviana1,
ZETU Cornelia1
1“N. Paulescu” National Institute of Diabetes, Nutrition and Metabolic Diseases, Bucharest (ROMANIA)
Email: dorosrodica@yahoo.com
Abstract
Background and Aim

Diabetic polyneuropathy (DPN) is a heterogeneous nervous dysfunction that associated
sensorimotor and autonomic neuropathy. The study aimed to evaluate the presence and severity
of sudomotor dysfunction in the low severity score of sensorimotor neuropathies.
Methods
A study group of 20 (11 males) patients with type 2 diabetes and low severity score assessed
by Toronto clinical score were evaluated for sudomotor dysfunction using Sudoscan device,
Impeto Medical
Result
The incidence of sudomotor dysfunction was related to the Toronto clinical score severity.
Sudomotor dysfunction was diagnosed in 36% (n=4) patients in group with no sensorimotor
neuropathy (n=11), 57% (n=4) patients in group with mild sensorimotor neuropathy (n=7),
100% (n=2) patients in group with moderate sensorimotor neuropathy (n=2). Severity and
topography of sudomotor dysfunction were not related to Toronto score severity and
sensorimotor DPN topography.
Conclusion
Estimation of sudomotor dysfunction provides a quantitative measurement of autonomic
dysfunction in preclinic stages of DPN giving support to earlier diagnosis and therapeutic
intervention.
Keywords: diabetic polyneuropathy, sudomotor dysfunction, autonomic dysfunction, Sudoscan, type 2 diabetes
Background and Aim
Heterogeneity of nervous lesions produced in long-standing diabetes mellitus associates in

different proportions small unmyelinated and myelinated fibers, large myelinated fibers or both
sensorimotor and autonomic fibers. This morpho pathological heterogeneity of diabetes
polyneuropathy (DPN) is related to distinctive symptomatology, clinical signs, specific
syndromes and neurophysiological tests. The presence of one positive test is sufficient to
diagnose nerve dysfunction as isolated nerve fibers type could exist [1].
Diabetic nerve fibers lesions are more than 50% initially asymptomatic [2] and neuropathic
dysfunction improvement was reported with physical exercise and metabolic optimization [3].
286
In this view, any efforts for detecting and treat preclinical and reversible conditions are
justified.
Several clinical scores and neurophysiologic tests for diabetic polyneuropathy are now
available. For large and small myelinated, nerve biopsy and in special, sural nerve biopsy is the
method of reference [4]. Other methods that could be taken into consideration for myelinated
nerve fibers dysfunction are nerve conduction tests for large fibers [5] and laser Doppler
flowmetry, sensory testing, and autonomic testing for small fibers [4]. These tests measure
different morpho pathologic lesions not always associated. In these circumstances, their results
could not substitute freely one to each other. It was also suggested that in DPN exists a more
complex functional relationship between different nervous fiber than related to simple nervous
lesions classification [2].
Small unmyelinated axons density and diameter reduction were considered the first
manifestation of the autonomic system dysfunction and their regeneration was demonstrated by
quantitative measures as intraepidermal nerve density [6]. A reduced density of sweat glands
nervous fibers and its association with DPN was demonstrated by comparative morphology and
neuropathy impairment score [7].
Measurement of the electrochemical skin conductance (ESC) through reverse iontophoresis
is a method that determines the autonomic nervous alteration of the unmyelinated cholinergic
sympathetic C nerve fibers that activate sweat glands through muscarinic receptors [8]. The
measurement of ESC was considered a reliable method of sweat glands nerve fibers estimation
and a sensitive test in DPN diagnosis. Sudomotor dysfunction was correlated with peripheric
and autonomic neuropathy by many authors [9-11].
Sudoscan device was considered a reliable device in autonomic nervous system dysfunction
assessment [12]. The device used reverse electrophoresis and estimates electrochemical
reaction between electrodes and chloride ions liberated from sweat glands.
Application of small intermittent and gradual electric stimulus on palmar and plantar
activates small nerve fibers that determine the increase of chlorine ions from the sweat glands
duct. The chloride ions increment increase skin conductance. Decreased number of sweat
glands or reduced autonomic nervous fibers determined a low ESC.
The device provides an objective determination of ESC of the palmar and plantar skin and 2
risk score: Nephropathy score (NA) and Cardiac Autonomic Neuropathy score (CAN), that are
computed using ESC, age and/or body mass index (BMI).
Toronto clinical score is a validated method for screening and severity rank estimation of
DPN. This score was validated related to morphological and electrophysiological
characteristics of sensorimotor diabetic neuropathy [13, 14].
The study aimed to analyse the autonomic dysfunction of sympathetic C fibers of sweat
glands as an early modification in diabetic patients in type 2 diabetes.
This pilot-study analysed 20 consecutive patients (55% male) with type 2 diabetes, evaluated
in our clinic with low Toronto clinical score from 0 (no neuropathy) to a maximum of 10 points
(moderate sensorimotor neuropathy). Exclusion criteria were: type 1 diabetes, diagnosed
autonomic neuropathy or severe DPN, HbA1c >9%, excessive alcohol consumption, conditions
that could modify innervations of sweat glands (connective tissue diseases, neurologic or
dermatologic conditions, known vitamins B deficiencies, concomitant anticholinergic
medication). The study was approved by the ethics committee and patients consented to study
procedures. Patients were evaluated in the outpatient clinic according to local procedures.
287
Sudomotor function was measured using Sudoscan device, Impeto Medical, France. Toronto
clinical score sensibility tests were performed using the method described by Perkins and
collab. [15]. Statistical analysis was performed with SPSS 22.
Results
Study group characteristics are presented in Table 1. In this study group patients had a mean
age of 62 years, mean evolution of diabetes 6.46 years, mean BMI 29.7, mean electrochemical
skin conductance (ESC) in the normal interval (more than 70 µS for feet and 60 µS for hands).
Mean value for Nephropathy score (mean NS=57) and Cardiac autonomic neuropathy score
(mean CAN=33.6) were in moderate risk interval (40-60 for NS and 30-50 for CAN). Mean
values are presented in Figures 1 and 2. Similar values with no significant difference were
observed in gender analysis (Table 2). Feet ESC was below the normal range (70µS) in 6
patients (2 males, 4 females) and Hands ESC was reduced below the normal range (60µS) in 6
patients (4 males, 2 females). Four patients (2 males, 2 females) have both palmar and plantar
sudomotor dysfunction, 2 males’ patients have only palmar sudomotor dysfunction and 1
female has only plantar sudomotor dysfunction.
Table 1 Study group characteristics (anthropometric, Table 2 Comparison between genders for
metabolic, years of diabetes, Toronto score, study group
Electrochemical skin conductance, Nefropathy score,
male (n=11) female (n=9)
Cardiac autonomic neuropathy score)
1 2 1
Mean SD Minimum Maximum Mean SD Mean SD p value
Age (years) 62 8.51 50 81 64.91 9.22 58.44 6.31 ns
BMI 29.68 4.37 23.0 40.1 30.29 4.94 28.92 3.71 ns
Height (cm) 168.95 9.85 156 190 174.18 10.22 162.56 4.07 <0.05
Weight (kg) 84.70 17.43 47 116 92.09 17.01 75.67 13.87 <0.05
HbA1c 6.70 0.91 5.0 8.6 6.79 0.81 6.578 1.07 ns
years diabetes 6.46 4.59 1 15 6.09 3.30 7.00 5.87 ns
Toronto score 4.80 3.47 0 10 4.64 4.03 5.00 2.87 ns
Feet ESC1 (µS)2 73.40 10.82 52 87 77.00 5.69 69.00 14.07 ns
Hands ESC (µS) 63.70 18.74 24 84 61.82 15.03 66.00 23.26 ns
Nephropathy score 56.85 16.54 4 78 58.91 12.10 54.33 21.30 ns
Cardiac Autonomic 33.60 10.41 1 46 36.09 7.57 30.56 12.91 ns
Neuropathy score
Right foot ESC (µS) 74.30 10.76 50 86 77.73 5.22 70.11 14.32 ns
Left foot ESC (µS) 73.80 11.50 52 88 76.73 6.59 70.22 15.28 ns
Feet asymmetry % 2.95 2.93 0 10 3.00 3.38 2.89 2.47 ns
Right hand ESC (µS) 63.15 19.32 24 85 60.36 15.44 66.56 23.77 ns
Left hand ESC (µS) 64.60 18.16 24 84 63.82 14.48 65.56 22.79 ns
Hands asymmetry % 4.55 4.51 0 15 5.82 4.64 3.00 4.06 ns
1- stadard deviation; 2- electrochemical skin conductance; 2- µSiemens 1- p value computed with independent T-test, p<0.05 statiscal significant; ns-
not statistical significant; 2- standard deviation; 3- electrochemical skin
conductance µSiemens
288
Table 3a Bivariate correlations indices for analysed parameters

1
Positive correlation
BMI Weight (kg) 0.805*** Cardiac autonomy BMI 0.497*
neuropathy score
Cardiac autonomic 0.497* Right foot ESC (µS) Feet ESC (µS) 0.981***
neuropathy score
Height (cm) Weight (kg) 0.678** Left foot Esc (µS) 0.959***
Weight (kg) BMI 0.805*** Hands ESC (µS) 0.469*
Height (cm) 0.678** Left hand ESC (µS) 0.467*
Toronto score Years of diabetes 0.573** Right hand ESC (µS) 0.466*
HbA1c Left hand ESC (µS)2 0.592** Left foot ESC(µS) Feet ESC (µS) 0.967***
Hands ESC (µS) 0.561* Right hand ESC (µS) 0.492*
Right hand ESC (µS) 0.526* Hands ESC (µS) 0.491*
Feet ESC (µS) Right foot ESC (µS) 0.981*** Left hand ESC (µS) 0.485*
Left foot Esc (µS) 0.967*** Right hand ESC(µS) Hands ESC (µS) 0.995***
Hands ESC (µS) 0.452* HbA1c 0.526*
Right hand ESC (µS) 0.452* Feet ESC (µS) 0.452*
Left hand ESC (µS) 0.446* Left hand ESC (µS) Hands ESC (µS) 0.991***
Hands ESC (µS) Right hand ESC (µS) 0.995*** Right hand ESC (µS) 0.971***
Left hand ESC (µS) 0.991*** HbA1c 0.592**
HbA1c 0.561** Left foot ESC (µS) 0.485*
Left foot Esc (µS) 0.491* Right foot ESC (µS) 0.467*
Right foot ESC (µS) 0.469* Feet ESC (µS) 0.446*
Feet ESC (µS) 0.452* 1- Pearson coefficients in descending order; 2- electrochemical conductance of skin; *-
p<0.05;**- p<0.01; ***-p<0.001
Table 3b Bivariate correlations indices for

analysed parameters
1
Negative correlations
Age (years) Right hand ESC (µS) 0.538*
Hands ESC (µS) 0.536*
Left hand ESC (µS) 0.525*
Feet ESC (µS) Feet asymmetry % 0.473*
Hands ESC (µS) Age (years) 0.536*
Cardiac autonomic 0.464*
neuropathy score
Cardiac autonomy Left foot Esc (µS) 0.488*
neuropathy score Right hand ESC (µS) 0.472*
Hands ESC (µS) 0.464*
Left hand ESC (µS) 0.457*
Left foot ESC (µS) Feet asymmetry % 0.499*
Feet asymmetry % Left foot ESC (µS) 0.499*
Feet ESC (µS) 0.473*
Right hand ESC Age 0.538*
(µS) Cardiac autonomic
0.472*
neuropathy score
Left hand ESC (µS) Cardiac autonomic 0.457*
neuropathy score
1- Pearson coefficients in descending order; 2- electrochemical conductance of skin; *-
p<0.05;**- p<0.01; ***-p<0.001
289
Computed Pearson correlations between analysed parameters are particularized in Tables 3a

and 3b. Mean values for feet and Hands ESC after Toronto clinical score severity are presented
in Table 4.
From 11 patients evaluated with no sensorimotor neuropathy with Toronto score, 36% have
autonomic neuropathy (2 patients have severe autonomic neuropathy and 2 patients’ moderate
autonomic dysfunction). From 7 patients with mild sensorimotor Toronto score, 57% (4
patients) had moderate autonomic dysfunctions. Both patients 100% (n=2) with moderate
sensorimotor neuropathy had also autonomic neuropathy. The most severe 2 cases of autonomic
neuropathy were found in patients with no sensorimotor neuropathy according to Toronto
clinical score. Sudomotor dysfunction values positive for autonomic neuropathy and their
relation with Toronto score are detailed in Table 5.
Table 5 Abnormal electrochemical skin conductance (ESC) related

to Toronto clinical score subgroups
Toronto clinical score Feet ESC (µS)<70µS Hands ESC
subgroups N value µS n value µS
no neuropathy (n =11) N=2 53, 54 N=2 24, 29
mild neuropathy (n=7) N=3 52, 60, 68 N=2 40, 52
moderate neuropathy (n=2) N=1 69 N=2 36, 56
n- total number of patients in subgroup; N- number of ESC abnormal value (positive values
for autonomic neuropathy)
Table 6 Distribution of patients in severity neuropathy subgrous groups by
gender
male female
(n=11) (n=9)
Toronto severity no neuropathy (n=11) 6 5
1
groups mild neuropathy (n=7) 3 4
moderate neuropathy (n=2) 2 0
ESC Feet score 2 no sudomotor dysfunction (n=14) 9 5

moderate sudomotor dysfunction (n=6) 2 4
severe sudomotor dysfunction 0 0
ESC Hands score3 no sudomotor dysfunction (n=14) 7 7
moderate sudomotor dysfunction (n=3) 3 0
severe sudomotor dysfunction (n=3) 1 2
290
Toronto clinical score diagnose neuropathy in 45% of patients, Feet ESC score in 30% of
patients and Hands ESC score in 20%. Distribution detailed in Figure 3 and by gender in Table
6.
A positive correlation was found between Feet and Hands ESC. The normal values of HbA1c
were not correlated with the decrease of ESC. Age was negatively correlated with Hands ESC
and positively with Toronto score.
Discussions
For this study group, sensorimotor neuropathy and autonomic neuropathy do not completely
overlap sustaining the idea that they represent different aspects of nerve dysfunction.
Autonomic dysfunction for this study group was not related to obesity and was not
negatively affected by increasing HbA1c. A positive correlation between Hand ESC values and
HbA1c seems to be a statically finding related to the small number of cases and low to moderate
HbA1c. However metabolic optimization did not prove to be equally effective in type 2 diabetes
polyneuropathy as in type 1 diabetes polyneuropathy suggesting that other factors as a slow and
prolonged increment of glycaemic value, associated comorbidities as dyslipidaemia, arterial
hypertension, increased adipose tissue or drugs involved in their treatment are implicated in
nervous lesions [16].
Autonomic neuropathy is not axon length-dependent and it is not appearing primarily at feet
level. In this study, hands autonomic neuropathy was an early and more severe modification.
Relationship between age and feet ESC was not demonstrated in this study group possible
because of the small number of analysed cases or a more complex relationship between
autonomic dysfunction localization and neuropathological factors. This aspect is important for
the clinician who has to explore patients with diabetes.
Conclusion
Sudomotor dysfunction is an early modification in type 2 diabetes that appears in a

significant proportion of patients before the Toronto clinical score detects sensorimotor
neuropathy. Regardless of the fibers type involved in DPN, the potential nervous regeneration
for pre-symptomatic stages could be a successful target in the treatment of diabetic neuropathy.
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292
Hypercortisolism Is Associated with Increased Cardiovascular

Mortality and Morbidity: A Retrospective Study
DUMITRIU Roxana1, DUȘCEAC Roxana1,2, SALMEN Teodor3,

POIANĂ Cătălina1,2
1 “C. I. Parhon” National Institute of Endocrinology, Bucharest (ROMANIA)
2 Department of Endocrinology, “Carol Davila” University of Medicine and Pharmacy, Bucharest (ROMANIA)
3 “Prof. Dr. N. C. Paulescu” National Institute of Diabetes, Nutrition and Metabolic Diseases, Bucharest (ROMANIA)
Email: roxanaeu@gmail.com
Abstract
Objective
Cushing’s syndrome is known to be associated with increased cardiovascular risk. Chronic
hypercortisolism, independently of its ethiology, can determine metabolic impairment,
increased mortality and morbidity. The objective of our study was to determine the predictors
of cardiovascular disease and the most frequent metabolic disorders. The primary end point of
our study was defined as the occurrence of major cardiovascular events: myocardial infarction,
stroke, deep venous thrombosis or pulmonary embolism, ischemic heart disease.
Methods
25 patients with Cushing’s syndrome (15 with Cushing’s disease and 10 with adrenal
adenomas) were included. Anthropometric (weight, height, BMI) and laboratory parameters
(including lipid profile, hormonal profile) were measured in patients. A retrospective analysis
was conducted and the patients were divided in two groups (first with Cushing’s disease and
the second one with Cushing’s syndrome caused by adrenal adenoma).
Results
In the first group there was observed a higher prevalence of dyslipidaemia (56.25% versus
40%), the percentage of strokes was higher (12.5% versus 0%) and diabetes mellitus was
diagnosed in 43.75% of patients. Overall a high percentage of the patients presented
thromboembolic events (20% in the both groups), with a slightly higher occurrence in the
second group (patients with adrenal adenomas).
Keywords: Cushing, hypercortisolism, metabolism, cardiovascular risk
Introduction
Cushing’s disease is the most frequent cause of endogenous hypercortisolaemia, it is 5 to 6

times more frequent than adrenal Cushing’s syndrome [1] and it occurs more frequently in
women. ACTH-independent Cushing’s syndrome is found in 20% of cases and it is more
frequently due to adenomas (10%) or adrenal carcinomas [1]. Bilateral micronodular or
macronodular hyperplasia are rare and represent less than 10% of all cases of ACTH-
independent Cushing’s syndrome.
The signs and symptoms of Cushing’s syndrome are a result of long-term exposure to
excessive glucocorticoids. Hypertension, central obesity, diabetes mellitus, osteoporosis,
thromboembolic events, insulin resistance are highly prevalent compared to the general
293
population. In this scenario the metabolic syndrome plays the most important role in
determining the increased cardiovascular risk with a mortality rate four times higher than in the
general population [2]. It is not known if the remission of the disease reverses these
complications. Some studies showed that in patients in remission the high prevalence of
metabolic syndrome, atherosclerosis, cardiovascular and cerebrovascular risks remain.
In the literature there are few studies that compare the metabolic and cardiovascular risks in
patients with Cushing’s disease versus adrenal cortisol-secreting adenomas. Also, if we
evaluate if other concomitant hormone deficiencies present in Cushing’s disease impact the
metabolic impairment the data is controversial [3].
Methodology
A retrospective review of 25 patients diagnosed with Cushing’s syndrome or Cushing’s

disease. We evaluated the metabolic and cardiovascular profile.
Anthropometric evaluation (height, weight, BMI), fasting glucose, HbA1c, total cholesterol,
HDL (high density lipoprotein), LDL cholesterol (low density lipoprotein), triglyceride levels,
ACTH and cortisol levels (preoperatively and postoperatively) were measured.
The statistical analysis was done using SPSS 19.0 software, quantitative data was expressed
as mean ± SEM and categorical data as percentage. Logistic regression was used to analyse the
dichotomic response variables. Statistical significance was set at p<0.05.
Results
Mean age of the patients enrolled was 55.56±13.29 years, mostly females, 12 (48%) of them
were active smokers. Mean BMI was slightly higher in the group of patients with Cushing’s
disease, the majority were obese (60% versus 50% in the second group), with no significant
difference. Overweight was present in 2 of 10 the patients with adrenal adenomas (20%) and in
9 of the 15 patients with Cushing’s disease (40%) (Fig. 1).
10 of the 15 patients with Cushing’s disease underwent transsphenoidal resection, 8 of them
were treated by radiotherapy. All patients with residual pituitary deficiency underwent hormone
replacement.
All patients with Cushing’s syndrome caused by adrenal adenomas underwent unilateral
adrenalectomy (9 of 10 patients) and 1 patient had bilateral adrenalectomy with good outcomes
postoperatively.
Dyslipidaemia was present in 52% of the cases, no significant differences between the two
groups.
Hypertension was found in 21 patients (72%) most of them had mild hypertension (grade II
according to WHO/ISH 2003) (Table 1).
Type 2 diabetes had a higher prevalence in Cushing’s disease (43.75%) than in patients with
adrenal adenomas. 8 patients were under oral glucose-lowering drugs and 3 were under insulin
therapy.
294
Cushing’s syndrome Group 1 (Cushing’s disease) Group 2 (Cushing’s syndrome)

Age 55.56±13.29 years 50.75±14.01 years 64.11±5.75 years
BMI 29.34±5.96 kg/m 2
50.75±14.01 kg/m 2
26.36±4.56 kg/m2
Dyslipidaemia 52% 56.25% 40%
Hypertension 72% 68.75% 70%
PAD 24% 25% 20%
Stroke 8% 12.5% 0%
TE 20% 12.5% 30%
Type 2 diabetes 36% 43.75% 20%
Table 1. Main characteristics of the groups (BMI=body mass index, PAD=peripheral artery disease,
MI=myocardial infarction, TE=thromboembolic event )
ACTH (pg/ml) Cortisol 8 a.m (µg/dl)

General 51.37 21.54±8.16
Group 1 (CD) 78.85 25.17±8.47
Group 2 (CS) 6.64 17.17±5.74
Table 2. Baseline hormonal profile-cortisol, ACTH (CD=Cushing’s disease, CS=Cushing’s syndrome)
Baseline hormonal profile with mean values of cortisol: 21.54±8.16 µg/dl and mean ACTH
values: 51.37 pg/ml (normal range: 3-66 pg/ml). Mean exposure to hypercortisolaemia was of
1 year.
100%
80% 40% 20%
16%
60% 16% 20%
40%
48% 60% 50%
20%
0%
General CD CS
Obesity Overweight Normal
Fig. 1. Prevalence of obesity and overweight in patients with Cushing’s disease and Cushing’s syndrome
(adrenal adenoma)
Mean BMI did not change significantly after therapy. It was observed a persistence of central
fat distribution in both groups after remission (Fig. 1). Glucocorticoid normalization induced a
decrease of hypertension in both groups. Lipid metabolism improvement was not observed,
most of the patients were under long standing statin therapy.
Fig. 2. Major cardiovascular events (CHD=coronary heart disease, TE=thromboembolic events)
295
The composite primary end-point was observed in 72% of the patients. 8% of the patients
experienced a stroke, 44% were diagnosed with ischemic coronary disease and 20% had
thromboembolic events (mostly patients with Cushing’s syndrome after surgical treatment –
unilateral or bilateral adrenalectomy) pulmonary embolism or deep vein thrombosis (Fig. 2).
A positive correlation was found between the value of basal cortisol and ischemic coronary
disease (p<0.05) (two tailed Pearson correlations).
The percentage of cured patients was 25%, 20% of them were patients with Cushing’s
syndrome caused by adrenal adenomas cured after unilateral adrenalectomy.
Discussions
The results of our study confirm that patient’s with Cushing’s syndrome have a high
prevalence of obesity, dyslipidaemia, hypertension. Also, if we compare between the ethiology,
patients with Cushing syndrome present a higher prevalence of risk factors for major
cardiovascular events. Also, the prevalence of diabetes mellitus was higher in these patients.
Cortisol excess is known to affect glycaemic metabolism through many mechanisms:
increased gluconeogenesis, inhibition of glucose uptake by the peripheral tissues and impaired
insulin function [4]. Also, the lipid metabolism is involved, in our study we found a higher
prevalence of dyslipidaemia in patients with Cushing’s disease. This feature results from a
glucocorticoid induced increased lipolysis, liponeogenesis and adipogenesis. Altered adipokine
release, vascular alterations, atherosclerosis and hypertension are some of the factors that
contribute to the increased cardiovascular morbidity and mortality [5, 17]. Also, these patients
are in a prothrombotic state. The risk of major cardiovascular events is estimated considering
the combined effect of several risk factors: hypertension, diabetes, dyslipidaemia etc), organ
damage (proteinuria, left ventricular hypertrophy) [6, 14].
Even in cured patients from Cushing’s syndrome, after a long remission, there is observed a
high prevalence of atherosclerosis and several risk factors (present when the disease was active)
remain present, probably due to visceral obesity along with insulin resistance.
Cushing’s syndrome represents an archetype of metabolic syndrome in several clinical trials
the different clinical signs and symptoms showed that the patients were obese (95%), they had
facial plethora (90% of cases), hypertension (75% of cases) and glucose intolerance (60% of
cases) [7, 16]. The visceral obesity can be explained by glucocorticoids that stimulate the
appetite and the lipoprotein lipase activity. The metabolic syndrome can also persist in a big
number of patients, despite the normalization of cortisolemia. Cortisol excess affects GH/IGF-
1 increasing visceral fat and insulin resistance.
Studies show that approximately 20% of patients with Cushing’s syndrome experience at
least one thrombotic event, mostly of them occurring after pituitary or adrenal surgery [8].
The highest risk was observed in patients that underwent bilateral adrenalectomy.
Hypertension has a multifactorial pathogenesis and rises the odds for a major cardiovascular
event to occur [9, 13, 15]. Mineralocorticoid mimetic activity, alterations of renovascular
resistance, vascular remodelling are complex mechanism that determine high blood pressure
values. The molecular basis of mineralocorticoid-induced hypertension is linked to the
overactivity of the epithelial sodium channel via the activation of glucocorticoid receptors. It
causes glomerular hyperfiltration.
Endothelin-1 (ET-1) is a potent vasoconstrictor with mitogenic and atherogenic effects on
smooth muscular cells. ET-1 has been found elevated in patients with Cushing’s syndrome, that
probably promote early atherosclerosis [10, 11, 12]. The production of vasodilators is down
regulated, glucocorticoids inhibit nitric oxide synthase (NOS), prostacyclin, prostaglandins and
kallikreins. An increased sensitivity to catecholamines is also present [10].
296
Conclusions
Our study confirms that chronic hypercortisolism contributes to metabolic impairment and
increased cardiovascular risk. The two groups we analysed in this study showed a slight
difference between patients with Cushing’s disease and those with Cushing’s syndrome caused
by adrenal adenoma, the first group presented a higher percentage of dyslipidaemia,
thromboembolic events after pituitary surgery and stroke. The patients with adrenal cortisol
secreting adenomas seem to have a better clinical outcome, 20% of these patients were cured
after surgery. We found a positive (strong correlation) between basal cortisol and coronary heart
disease.
REFERENCES
1. Lahera vargas M. et al., (2009). Prevalence, ethiology and clinical findings of Cushing’s syndrome.
Endocrinol Nutr. 56 (1). Pp. 32-39.
2. Extabe J., Vazquez J.A. (1994). Morbidity and mortality in Cushing’s disease: an epidemiological
approach. Clinical Endocrinology (Oxford), 40, pp. 479-484.
3. Barahona M.J. et al., (2009). Persistent body fat mass and inflammatory marker increases after long-
therm cure of Cushing’s syndrome. Journal of Clinical Endocrinology and Metabolism. 94. pp. 3365-
3371.
4. Andrews R.C. Walker B.R. (1999). Glucocorticoids and insulin resistance: old hormones, new targets.
Clinical Science. 96. Pp. 513-523.
5. Ferrau fF. Korbonits M. (2015). Metabolic comorbidities in Cushing’s syndrome. European Journal of
Endocrinology. 173 (4), pp. 133-157.
6. Mancini T. et al., (2004). High cardiovascular risk in patients with Cushing’s syndrome according to
1999 WHO/ISH guidelines. Clinical Endocrinology. 61 (6), pp. 655-801.
7. Chanson P. Salenave S. (2010). Metabolic Syndrome in Cushing’s Syndrome. Neuroendocrinology. 92
(1), pp. 96-101.
8. Suarez M.G. et al., (2019). Risk for thrombotic events high after Cushing’s syndrome surgery.
9. Pivonello R. et al., (2016). Complications of Cushing’s syndrome: state of the art. 4 (7), pp. 611-629.
10. Igaz P., Seccia T.M. (2019). Editorial: Endocrine Forms of Hypertension: Clinical and emerging
Molecular aspects. Front. Endocrinol.
11. Giordano R. et al., (2011). Metabolic and Cardiovascular outcomes in patients with Cushing’s syndrome
of different aetiologies during active disease and 1 year after remission. 75, pp. 354-360.
12. Lambert J.K. et al., Predictors of Mortality and Long-term Outcomes in Treated Cushing’s Disease: A
study of 346 patients. 98 (3), pp. 1022-1030.
13. Barbot M. et al., (2018). Diabetus Melittus Secondary to Cushing’s Disease. 284 (9).
14. Colao A. et al., (1999). Persistence of increased cardiovascular risk in patients with Cushing’s disease
after five years of successful cure. J Clin Endocrinol Metab. 84, pp. 2664-2672.
15. Graversen D. et al., (2012). Mortality in Cushing’s Syndrome: a systematic review and meta-analysis.
Eur J Intern Med. 23, pp. 278-282.
16. Swearingen B. et al., (1999). Long-term mortality after transsphenoidal surgery for Cushing disease. Ann
Intern Med. 130, pp. 821-824.
17. Barahona M.J. et al., (2009). Persistent body fat mass and inflammatory marker increase after long-term
cure of Cushing’s syndrome. Journal of Clinical Endocrinology and Metabolism. 94, pp. 3365-3371.
297
Heart Failure in Patients with Diabetes Mellitus: Paraclinical

Findings
SALMEN Teodor1, STEGARU Daniela1,2, PIETROSEL Valeria-Anca1,

ZARZU Alexandra1, ALEXE Vlad2, MILICESCU Alexandra1, DOROS Rodica1,
MIHAI Andrada1,2
1“N. Paulescu” National Institute for Diabetes Mellitus, Nutrition and Metabolic Disorders, Bucharest (ROMANIA)
Email: stegaru.daniela@gmail.com
Abstract
Introduction
Diabetes Mellitus (DM) and glycaemic control increase the risk of developing heart failure
(HF). DM cardiomyopathy is associated with both diastolic and systolic dysfunction, and
eventually by clinical HF. Furthermore, obesity is responsible for structural and functional
changes in the heart that consequently lead to an increase in the incidence and prevalence of
HF in DM patients.

A cross-sectional study was carried out in NIDNMD N.C. Paulescu from Bucharest during
January-November 2019. Demographic, clinical and paraclinical data were collected from
inpatients admitted to the Cardiology Department and analysed them using Microsoft Excel
2013 and PSPP software.
Results
269 inpatients with a mean age of 61.8±11.55 years, 48.32% females, a mean A1c
8.12%±1.9, mean duration of diabetes of 13.07±4 years were included. All patients had DM,
while 53.9% of patients were obese, and 54.57% of patients had HF.
Discussions
DM patients have an increased risk of cardiovascular events and, also, DM can determine
HF by direct microvascular inflammation of the myocardium. HF prevalence is increasing and
the patients that request hospitalization for HF have a worse survival, with an increase in
mortality rates. DM can lead to diastolic dysfunction in the absence of ventricular hypertrophy,
through myocardiopathy.
Conclusions
HF, DM and obesity prevalence continues to rise, leading to an increased financial burden
for the healthcare systems worldwide. SGLT2 inhibitors are the only medication efficient both
in prevention and amelioration of HF symptoms, and, also, in decreasing the rate of
hospitalization for HF. Despite that DM treatment has clear indications, there is still need of a
consensus for a recommendation of nutritional intervention and decrease weight for overweight
and obese patients.
Keywords: diabetes mellitus, heart failure, obesity, A1c haemoglobin
298
Introduction
Patients with diabetes mellitus (DM) have over twice the risk of developing heart failure
(HF) than patients without DM [1, 2]. The Framingham Heart Study suggests that DM
independently increases the risk of HF up to 2-fold in men and 5-fold in women compared with
age-matched controls [3, 4]. The increased incidence of HF in diabetic patients persists
independently of other risk factors such as age, high blood pressure (HBP),
hypercholesterolemia, and coronary artery disease (CAD). Cardiovascular outcomes and
prognosis are worse for patients with DM in comparison to those without [5].
Complex structural and functional changes underlie diabetic cardiomyopathy. In people with
DM, the presence of myocardial dysfunction in the absence of overt clinical CAD, valvular
abnormalities, and other conventional cardiovascular risk factors, such as hypertension and
dyslipidaemia, has led to the descriptive terminology, diabetic cardiomyopathy. The prevalence
of diabetic cardiomyopathy is increasing in parallel with the increase in DM. Diabetic
cardiomyopathy is initially characterized by myocardial fibrosis, dysfunctional remodelling and
associated diastolic dysfunction, later by systolic dysfunction, and eventually by clinical heart
failure [6].
Further data [7] demonstrated differences in left ventricular (LV) mass and wall thickness
and increased diastolic and systolic dysfunctions between diabetic patients and normal
individuals. It is reported that in type 1 DM, each 1% increase in glycated haemoglobin A1c
was linked to a 30% increase in the risk for HF (Lind et al.,), whereas in type 2 DM (T2DM),
each 1% rise in haemoglobin A1c level was associated with an 8% increase in risk, independent
of other risk factors, including obesity, smoking, HBP, dyslipidaemia, and coronary heart
disease (Stratton et al.,), suggesting that progressive increases in glycemia are a powerful
promoter of HF in patients with DM [5].
Obesity is associated with structural and functional changes in the heart and has adverse
effects on hemodynamic and left ventricular (LV) structure (dilation and hypertrophy) and
function. Furthermore, left atrial enlargement is frequently seen as a consequence of both an
increase in blood volume and changes in LV diastolic filling. Summing up, it may be assumed
that obesity leads to an increase in the incidence and prevalence of HF [8]. Moreover, Aune D
et al., report a strong association between body mass index (BMI) and waist circumference with
HF incidence [9].
A cross-sectional study was carried out in NIDNMD N.C. Paulescu from Bucharest during
January-November 2019. We collected demographic, clinical and paraclinical data from
inpatients admitted to the Cardiology Department and analysed them using Microsoft Excel
2013 and PSPP software.
To diagnose HF, signs and symptoms as well as patient’s medical history were reviewed, a
physical examination was performed, and paraclinical investigations were run: ECG and
echocardiography.
Results
We included 269 inpatients with a mean age of 61.8±11.55 years, 48.32% females. DM
distribution was 20.44% type 1 DM, 79.55% T2DM of which 66% were insulin-treated.
Further characteristics are shown in Table 1 and Table 2.
299
Table 1. Baseline characteristics of the study population

Parameter Mean Value
Mean age (years ± SD) 61.8±11.55
Females (%) 48.32
Mean A1c 8.12±1.9
Mean duration of diabetes (years ± SD) 13.07±4
Obesity (%) 53.9%
Mean BMI (kg/m2±SD) 31.27±7.28
High Blood Pressure (%) 82.39
Dyslipidaemia (%) 77.46
Mean eGFR (ml/min/m2 ± SD) 86.76±31.44
Mean ACR (albumin-to-creatinine ratio) (mg/g 553.94 (1, 6473)
creatinine)
Table 2. Baseline comorbidities of the study population

Parameter Mean Value
Confirmed ASCVD (%) 93.3%
Coronary artery disease (%) 6.31%
Peripheral artery disease (%) 24.53%
Carotid atherosclerosis (%) 71.74%
Heart failure (%) 54.57%
Left ventricular hypertrophy (%) 61.71%
Diastolic dysfunction (%) 69.5%
Confirmed ASCVD refers to presence of Coronary artery disease, Peripheral artery disease, Carotid
atherosclerosis or history of cerebral or cardiac acute events (Ischemic stroke, Myocardial infarction)
DM complications presented in the study group were both microvascular – diabetic

peripheral neuropathy (55.01%), autonomic neuropathy (4.83%), retinopathy (32.34%),
chronic kidney disease (30.85%), and macrovascular – coronary artery disease (6.31%),
peripheral arterial disease (24.53%), carotid atherosclerosis (71.45%). Other metabolic
syndrome components were also present: 53.9% obesity, HBP (82.89%) and dyslipidaemia
(78.06%).
More than half of patients (54.64%) already had HF and 19.33% had atrial fibrillation.
After cardiac echography examination was performed, several parameters were particularly
considered: LV ejection fraction, presence of LV hypertrophy and diastolic dysfunction. LV
hypertrophy (LVH) as appreciated by measuring the LV wall thickness during cardiac
ultrasound examination was detected in 61.71% of the studied group.
Distribution of the left ventricular ejection fractions (LVEjF) values and the degrees of
diastolic dysfunction are depicted in the figures below (Fig. 1 and Fig. 2).
300
Fig. 1. Distribution of Ejection Fractionin the study group Fig. 2. Distribution of Diastolic Dysfunction in the
study group
While assessing the association between glycaemic control as featured by the A1c
haemoglobin values and cardiac ultrasound parameters such as the presence of diastolic
dysfunction and left ventricular hypertrophy, significant correlations were observed [OR of
1.09 CI (1.06, 1.13) for DD and 5.15 CI (3.03, 8.77), for LVH respectively].
We further divided the patients into two groups by taking into consideration the BMI – a
non-obese group with BMI <30 kg/m2, and, respectively, an obese group for BMI >=30 kg/m2.
Thus, it was noticed that the frequency of atherosclerotic cardiovascular disease, HBP and left
ventricular hypertrophy was significantly greater in the obese group (p=0.001, p<0.001 and
p=0.001 respectively). Also, a correlation between BMI and macrovascular complications of
diabetes showed a positive correlation with Carotid atherosclerosis OR 1.03 CI (1.02, 1.04).
Nonetheless, given the possibility that DM can lead to diastolic dysfunction in the absence
of ventricular hypertrophy, we evaluated the patients that according to the cardiac echographic
examination presented parameters suggesting diastolic dysfunction, despite normal ventricular
wall thickness. Hence, 46 of 179 patients, amounting up to 25.69%, were discovered to be in
this category.
The study population was further analysed based on the LVEjF values: preserved ejection
fraction (>=50%) and modified ejection fraction (<50%). Macrovascular complications
(Peripheral arterial disease, Carotid atherosclerosis, Atherosclerotic cardiovascular disease) did
not significantly correlate to the degree of ventricular systolic dysfunction (Table 3).
Regarding other cardiovascular risk factors, it was observed that the presence of chronic
kidney disease (as diagnosed using KDIGO 2012 criteria) had a strong association with the
LVEjF value (p=0.05), although between ejection fraction value and albumin to creatinine ratio,
no statistically significant results were obtained.
Table 3. Correlation between LVEjF and macrovascular complications of diabetes

Carotid atherosclerosis OR 0.97 CI (0.41, 2.29)
Peripheral arterial disease OR 0.98 (0.4, 2.41)
Atherosclerotic cardiovascular disease OR 0.57 CI (0.26, 1.26)
Discussions
DM can determine HF by both atherosclerosis-dependent and independent mechanisms.

Type 2 DM patients have an increased risk of atherosclerotic cardiovascular events such as
myocardial infarction leading to myocardial wall abnormalities and consequent reduced
ejection fraction HF. Moreover type 2 DM patients have an increased risk of HF given the direct
microvascular inflammation of the myocardium [10-11].
301
Factors such as aging, growing survival after myocardial infarction and increment in the
prevalence of HF risk factors (age, gender, obesity, poor glycaemic and blood pressure control)
are responsible for rising the HF prevalence [12], reports estimating that HF will affect 23
million persons world-wide [13] and that 1 in 5 persons will develop HF anytime during their
lifetime [14].
DM can lead to diastolic dysfunction in the absence of ventricular hypertrophy, through
myocardiopathy [6]. There is also, a U-shape relationship between HF and HbA1c [15].
Another aspect related to the global prevalence of HF revealed from the analysis of Clinical
Practice Research Datalink from Great Britain that worth to be mentioned is that a patient with
HF which requests hospitalization for HF will have a worse survival than the patients with HF
p<0.001 [16-17]. Even more, there are data that states that after the first hospitalization for HF,
the mortality rates increase [18].
Conclusions
HF prevalence continues to rise, in parallel with the increase in both DM and obesity
prevalence; this leads to an increasing financial burden for the healthcare systems worldwide
through costs generated by the medical care as well as the decreased quality of life of these
patients.
The only efficient therapies, both in prevention and amelioration of these symptoms, and
also in decreasing the rate of hospitalization for HF are represented by SGLT2 inhibitors,
medications that are recommended by the actual guidelines in treatment of HF and chronic
kidney disease [19-20]. Our study demonstrated that albumin to creatinine ratio tightly
correlates with the presence of HF.
Despite the fact that in DM treatment are clear indications both in the guidelines of
diabetologists and cardiologists, there is still need of a consensus for a recommendation of
nutritional intervention and decrease weight for overweight and obese patients.
REFERENCES
1. Nichols GA, Hillier TA, Erbey JR, Brown JB. Congestive heart failure in type 2 diabetes: prevalence,
incidence, and risk factors. Diabetes Care 24: pp. 1614-1619, 2001.
2. Dei Cas A, Khan SS, Butler J, et al., Impact of diabetes on epidemiology, treatment, and outcomes of
patients with heart failure. JACC Heart Fail 23: pp. 136-145, 2015.
3. Kannel WB, Hjortland M, Castelli WP. Role of diabetes in congestive heart failure: the Framingham
study. Am J Cardiol 34: pp. 29-34, 1974.
4. Kannel WB, McGee DL. Diabetes and cardiovascular disease. JAMA 241: pp. 2035-2038, 1979.
5. Helena K, Abel D. Heart Failure in Type 2 Diabetes Mellitus – Impact of Glucose-Lowering Agents,
Heart Failure Therapies, and Novel Therapeutic Strategies. Circulation Research.124(1): pp. 121-141,
2019.
6. Jia G, Hill M, Sowers J. Diabetic Cardiomyopathy – An Update of Mechanisms Contributing to This
Clinical Entity. Diabetic Cardiomyopathy. 122(4): pp. 624-638, 2018.
7. Bertoni AG, Goff DC, D’Agostino RB, et al., Diabetic cardiomyopathy and subclinical cardiovascular
disease: The Multi-Ethnic Study of Atherosclerosis (MESA). Diabetes Care. 29: pp. 588-594, 2006.
8. Hamzeh N, Ghadimi F, Farzaneh R, Hosseini SK. Obesity, Heart Failure, and Obesity Paradox. J Tehran
Heart Cent 12(1): pp. 1-5, 2017.
9. Aune D, Sen A, Norat T, et al., Body mass index, abdominal fatness, and heart failure incidence and
mortality: a systematic review and dose-response meta-analysis of prospective studies. Circulation. 133:
pp. 639-649, 2016.
10. de Simone G, et al., Diabetes and incident heart failure in hypertensive and normotensive participants of
the Strong Heart Study. J Hypertens. 28(2): pp. 353-360, 2010.
11. Redfield MM. Heart Failure with Preserved Ejection Fraction. N Engl J Med 375: pp. 1868-1877, 2016.
12. Hunt SA, et al., 2009 Focused update incorporated into the ACC/AHA 2005 Guidelines for the Diagnosis
and Management of Heart Failure in Adults A Report of the American College of Cardiology
302
Foundation/American Heart Association Task Force on Practice Guidelines Developed in Collaboration

with the International Society for Heart and Lung Transplantation. J Am Coll Cardiol 53: pp. e1-e90,
2009.
13. Bui AL, et al., Epidemiology and risk profile of heart failure. Nat Rev Cardiol 8: pp. 30-41, 2011.
14. Heidenreich PA, et al., Forecasting the impact of heart failure in the United States: a policy statement
from the American Heart Association. Circ Heart Fail 6: pp. 606-619, 2013.
15. Matsushita K, et al., The association of haemoglobin a1c with incident heart failure among people without
diabetes: the atherosclerosis risk in communities’ study. Diabetes 59: pp. 2020-2026, 2010.
16. Taylor CJ et al., Trends in survival after a diagnosis of heart failure in the United Kingdom 2000-2017:
population-based cohort study. BMJ 364: p. l223, 2019.
17. Tuppin P et al., Two-year outcome of patients after a first hospitalization for heart failure: A national
observational study. Arch Cardiovasc Dis 107: pp. 158-168, 2014.
18. Das SR. 2018 ACC Expert Consensus Decision Pathway on Novel Therapies for Cardiovascular Risk
Reduction in Patients with Type 2 Diabetes and Atherosclerotic Cardiovascular Disease: A Report of the
American College of Cardiology Task Force on Expert Consensus Decision Pathways. J Am Coll Cardiol.
72: pp. 3200-3223, 2018.
19. Atherton JJ et al., National Heart Foundation of Australia and Cardiac Society of Australia and New
Zealand: Guidelines for the Prevention, Detection, and Management of Heart Failure in Australia 2018.
Heart Lung Circ 27: pp. 1123-1208, 2018.
20. Piepoli MF et al., 2016 European Guidelines on cardiovascular disease prevention in clinical practice:
The Sixth Joint Task Force of the European Society of Cardiology and Other Societies on Cardiovascular
Disease Prevention in Clinical Practice (constituted by representatives of 10 societies and by invited
experts) Developed with the special contribution of the European Association for Cardiovascular
Prevention & Rehabilitation (EACPR). Atherosclerosis. 252: pp. 207-274, 2016.
303
Glucose Control in Patients with Acromegaly Resistant to First-

Line Somatostatin Receptor Analogues Monotherapy
GĂLOIU Simona1,2, MĂRGĂRIT Emma1, BACIU Ionela1,2, NICULESCU Dan1,2,

TRIFĂNESCU Raluca1,2, RADIAN Șerban1,2, CĂPĂȚÎNĂ Cristina1,2,
CARAGHEORGHEOPOL Andra1,2, POIANĂ Cătălina1,2
2“C.I. Parhon” National Institute of Endocrinology, Bucharest (ROMANIA)
Emails: endoparhon@gmail.com, simona.galoiu@gmail.com
Abstract
Growth hormone (GH) receptor antagonist (Pegvisomant) has been shown to obtain IGF1
normalization in 60-90% of patients with acromegaly and amelioration of glucose metabolism
in clinical trials. The aim of the study is to evaluate the effect of GH receptor antagonist in
monotherapy versus combined therapy with somatostatin receptor analogues (SSRA) with or
without dopamine agonist (DA) therapy on glucose metabolism related to acromegaly control.
Patients and Methods

37 patients with GH secreting pituitary adenoma previously operated and not cured, totally
or partially resistant to SSRA therapy, treated with pegvisomant as monotherapy or combined
with SSRA with or without DA therapy, consecutively evaluated in Neuroendocrinology Clinic
from Institute of Endocrinology between January2001-March2019. Baseline and at 6 months,
at 1, 2, 3- and 5-years serum IGF1, glucose, HbA1c, and pituitary tumour diameters were
recorded.
Results
IGF1 levels were lowered after 6 months and 1year of treatment and then remained stable at
2,3 and 5 years of treatment with Pegvisomant (from 2.3±0.2 xULN to 1.94±0.18 and 1.43±0.16
xULN at baseline, 6 months and 1 year, respectively, p=0.06 baseline versus 6 months and
0.001 baseline versus 1 year). We obtained control of acromegaly in 20% of patients at 6
months, and in 38% and 54.5 % after 1 and 3 years of treatment. 11 patients had diabetes
mellitus (29.72%) and 2 (5.4%) patients had dyskinesia at baseline. After the first 6 months of
therapy glucose level significantly decreased only in patients treated with Pegvisomant
monotherapy (p=0.02) and not in patients with combined therapy (p=0.4), while glycated
haemoglobin did not change significantly.
Conclusion
In patients with acromegaly resistant to somatostatin receptor analogues monotherapy, GH
receptor antagonist therapy is efficient in controlling IGF1 levels. Glucose levels improved after
the first 6 months of Pegvisomant monotherapy, but not in patients with combined therapy.
Keywords: acromegaly, diabetes
304
1. Introduction
Acromegaly is a disease characterized by a chronic hypersecretion of GH and IGF1, mostly

due to a pituitary adenoma. This disease which untreated is associated with high mortality ratio
(2.5-3 times compared to general population) [1], is accompanied by multiple cardiovascular
and metabolic comorbidities, which contribute to a reduced survival.
Secondary diabetes mellitus is one of the most frequent metabolic complications of
acromegaly, with a prevalence of 19-56% [2].
First generation of somatostatin receptor analogues (SSRA), Octreotide and Lanreotide are
usually the first line therapy in medical treatment of patients with acromegaly not cured after
neurosurgical excision of pituitary tumour, but can be used also as primary therapy. They bind
the somatostatin receptor type 2 (SSTR-2) and type 5 (SSTR-5). SSTR-2 is involved in
glucagon regulation, whereas SSTR-5 is highly expressed in pancreatic beta-cells and is
involved in modulating insulin secretion [3]. Also, incretins are modulated by SSTRA.
Second generation of SSRA, pasireotide, has a high affinity for SSTR-5 (40 times when
compared to Octreotide), followed by SSTR-2, SSTR-3, and SSTR-1 and as a consequence, is
associated with a higher incidence of hyperglycaemia (60-88%) [4].
GH receptor antagonist Pegvisomant is used in patients with acromegaly inadequately
controlled by SSRA, either in combination with SSRA and/or dopamine agonist (DA) either as
monotherapy. Studies have shown that Pegvisomant improves insulin sensitivity in patients
with acromegaly [5].
Aim
To evaluate the effect of GH receptor antagonist in monotherapy versus combined with
STRA with or without DA therapy on glucose metabolism related to acromegaly control.
2. Patients and Methods
We included 37 patients with GH secreting pituitary adenoma previously operated and not
cured, totally or partially resistant to SSRA therapy, treated with pegvisomant as monotherapy
or combined with SSTRA with or without DA therapy, consecutively evaluated in
Neuroendocrinology Clinic from Institute of Endocrinology between Jan2001-Mar2019.
Baseline and at 6 months, at 1, 2, 3- and 5-years serum IGF1, glucose, HbA1c, and pituitary
tumour diameters were recorded.
Diabetes mellitus and disorders of glucose tolerance were diagnosed using ADA 2018
criteria [6]. Serum IGF-I was measured by different commercial assays in different laboratories.
IGF-I data are shown as the ratio with the upper limit of the normal range for sex and age in
each assay (ULN; normal ≤1).
Data were presented as mean ± standard deviation. Comparison of means between two
continuous variables with a normal distribution was performed by the Student’s t-test, and
comparison between variables with abnormal distribution was performed with Mann-Whitney
test. A p value <0.05 was considered statistically significant.
3. Results
Patient’s characteristics
There were 21 females and 16 males; average age was 42.37±11.21 years. Average follow-
up of patients was 7.95±3.61 years (range 0.8-15.89 years). 8 patients were treated post-surgery
with radiotherapy and with Pegvisomant monotherapy and 29 patients were treated with
combined treatment, as follows: 3 patients received post-surgery SSRA and Pegvisomant, 9
305
patients SSRA, Cabergoline and Pegvisomant, 12 patients received pituitary radiotherapy,

SSRA, Cabergoline and Pegvisomant, 4 patients were irradiated and treated with SSRA and
Pegvisomant, and 1 patient was irradiated and treated with Pegvisomant and Cabergoline.
Efficiency of therapy
IGF1 levels were lowered after 6 months and 1year of treatment and then remained stable at
2, 3 and 5 years of treatment with Pegvisomant (from 2.3±0.2 xULN to 1.94±0.18 and
1.43±0.16 xULN at baseline, 6 months and 1 year, respectively, p=0.06 baseline versus 6
months and 0.001 baseline versus 1 year) (Fig. 1). We obtained control of acromegaly in 20%
of patients at 6 months, and in 38% and 54.5% after 1 and 3 years of treatment. Tumour
diameters no significantly rose from 15.8±2.28 mm at baseline to 17.8±2.83mm at 6 months,
18.43±3.62 mm at 1 year and 23.74±4.83 mm at 3 years. There was no difference of IGF1
efficacy or tumour diameters between patients with monotherapy versus combined treatment.
Fig. 1. IGF1 levels evolution after pegvisomant monotherapy or combined with somatostatin receptor analogues
with or without cabergoline
Glucose metabolism
Before Pegvisomant therapy, average glucose level was 122.94±57.72 mg/dl (80-331
mg/dl). 11 patients had diabetes mellitus (29.72%) and 2 (5.4%) patients had dysglicemia, but
not fulfilling the criteria for diabetes mellitus diagnosis. Glucose level was slightly lower in
306
patients with Pegvisomant monotherapy versus combined therapy (p=0.054) at baseline. After
the first 6 months of therapy glucose level significantly decreased only in patients treated with
Pegvisomant monotherapy (p=0.02) and not in patients with combined therapy (p=0.4), while
glycated haemoglobin did not change significantly. There was no difference between
Pegvisomant doses in patients with diabetes mellitus in comparison with patients without
diabetes mellitus.
Table 1. 1-year evolution of glucose levels and glycated haemoglobin in patients with acromegaly treated with
Pegvisomant in monotherapy or combined therapy
Therapy with N Mean Standard Significance (p)
Parameter
Pegvisomant Deviation
monotherapy 7 100.38 17.15
Glucose (mg/dl) base 0.054
combined 28 128.58 62.97
Glucose (mg/dl) monotherapy 7 85.98 19.68
0.01
after 6 months combined 24 116.62 38.70
Glucose (mg/dl) monotherapy 6 88.00 15.59
0.17
after 1 year combined 19 117.91 83.64
HbA1c base % 0.12
combined 22 6.73 1.97
Hba1c 6 months % 0.006
combined 17 6.59 1.68
Hba1c 1 year % 0.03
combined 15 6.67 1.86
4. Discussion
The results of this clinical study show that in patients with acromegaly totally or partially
resistant to somatostatin receptor analogues, GH receptor antagonist Pegvisomant is efficient
in controlling IGF1 levels in 54.5% of patients after 3 years of treatment and this control
remains stable after that. The largest international observational study that showed the long-
term (12 years) evolution of 2090 patients with acromegaly treated by Pegvisomant,
ACROSTUDY, showed a similar control of acromegaly. In this global surveillance study, the
percentage of patients with normal IGFI levels increased from 53% at year 1, to 58% at year 3
and to 73% at year 10 [5]. Similar with this study, in our patient’s tumour diameters did not rise
significantly during Pegvisomant therapy.
Regarding the effect of medical treatment of acromegaly on glucose metabolism, a
metanalysis of 47 prospective interventional studies demonstrated that somatostatin receptor
analogues reduced insulin and increased afterload glucose levels and HbA1c when utilized as
first-line or second-line therapy for acromegaly [7]. The results of this metanalysis suggested
that this medication could worsen glucose metabolism, especially postprandial glucose in
patients with acromegaly, but this effect is nonsignificant when compared with beneficial effect
on GH levels. Opposing effects were observed during GH receptor antagonist therapy, which
has been shown to have a positive effect on glucose metabolism, by decreasing fasting and
postprandial glucose levels. In ACROSTUDY, diabetic patients needed higher Pegvisomant
doses to achieve control of acromegaly than patients without diabetes.
In our study performed on limited number of patients, we observed glucose level
improvement only in patients treated by Pegvisomant as monotherapy after the first 6 months
and not in patients with combined treatment. This could be due to higher glucose levels at
baseline in patients with combined therapy versus monotherapy and maybe these patients need
higher doses of Pegvisomant in order to control IGF1 and glucose levels, as has been showed
in ACROSTUDY results.
307
5. Conclusion
In patients with acromegaly resistant to somatostatin receptor analogues monotherapy, GH

receptor antagonist therapy is efficient in controlling IGF1 levels. Glucose levels improved after
the first 6 months of Pegvisomant monotherapy, but not in patients with combined therapy.
REFERENCES
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Nationwide Survey of Mortality in Acromegaly (2005). J Clin Endocrinol Metab; 90: pp. 4081-6.
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term treatment with pegvisomant: observations from 2090 acromegaly patients in ACROSTUDY
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Physical Activity Level and Patterns of Patients with Type 2

Diabetes
SZABO Monica1,2, MÁTÉ Beáta1, BALOGH Rozália3, GERMÁN SALLÓ Márta1

1 “George Emil Palade” University of Medicine, Pharmacy, Sciences and Technology, Targu Mures (ROMANIA)
2 Department of Internal Medicine, Department of Diabetology, Emergency Hospital, Targu Mures (ROMANIA)
3 Marienhausklinik St. Elisabeth Wadern (GERMANY)
Emails: sztamo@gmail.com, beatamate@yahoo.com, baloghlia@yahoo.com, gsmarti66@yahoo.com
Abstract
Introduction
The beneficial effect of physical activity (PA) on type 2 diabetes is well established. Our
goal was to evaluate the exercise patterns of type 2 diabetes patients in our region and the
relation to sociodemographic data to develop a specific exercise strategy.

In our descriptive cross-sectional study, we obtained sociodemographic (gender, residence,
ethnicity, income, education) and PA data were from 332 type 2 diabetes patients. Exercise
habits were recorded in detail using the International Physical Activity Questionnaire. The
mean physical activity level (PAL) was calculated. Effort was divided into 4 categories:
walking (a), gardening or farming (b), profession-related activities (c) and sports or leisure
activities (d).
Results
Mean age was 63.17 (SD 10∙05) years, 54.3% were women. Only 30% of patients had a PAL
equivalent to or greater than the recommended guidelines. Activity types chosen by patients
were: walking, 55.1%; gardening, 31.3%; professional activities, 10.3%; sports 3.3%.
The PAL was highest in groups b and c (p<0.001). Men and rural inhabitants had a higher
PAL.
Conclusion
Only few patients are achieving the outlined goals. Compliance is far better with domestic
or professional activities than with sports; women, and individuals living in town are less active.
There is a need to develop specific, individualised exercise strategies for our patients.
Keywords: physical activity, diabetes mellitus type 2, Transylvania
1. Background and Aims
The prevalence of type 2 diabetes mellitus is increasing faster than predicted [1, 2]. The
Diabetes Prevention Program Research Group Study [3] and the Finnish Diabetes Prevention
Study [4]. Found that participants in lifestyle intervention plans including dietary changes and
increased physical activity (PA) sharply (by 50%) reduced their chances of developing diabetes
and had better BMIs (body mass index) and lipid profiles. Physical activity is a complex human
behaviour that is modified by cultural, social, environmental, and political factors and thus
considerably varies across different countries and areas. Using a representative Transylvanian
patient cohort selected from a district diabetes care centre, the present study aimed to examine
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the exercise patterns of type 2 diabetic patients (quantity, habits and quality) in relation to
sociodemographic data to identify impediments to PA and develop a specific exercise strategy
for our patients. To our knowledge, no such study has previously been performed.
2. Methods
This community-based cross-sectional study was conducted in Mureş County, Transylvania,

Romania. Data were obtained from patients who were selected randomly from the Central
Diabetes Registry of Mureş County (which included 21632 patients at that time in 2010).
Sample sizes were determined using a probability proportional to size technique. The
distribution of gender and area reflected the distribution of the entire cohort of patients
registered in Mureş County. A representative cohort of 332 type 2 diabetes patients was selected
that fulfilled the inclusion and exclusion criteria and consented to participate. We included only
type 2 diabetes patients who had been diagnosed with diabetes at least one year prior. Patients
with physical or mental disabilities that affected their exercise capability or habits were
excluded. General data such as age, gender, residence (rural or urban) and employment status,
were recorded. Anthropometric measurements (height, weight, and waist circumference) were
collected, and BMI was calculated as weight (kg)/height(m)2. All procedures involving human
subjects/patients were approved by the local Ethical Committee.
All participating patients provided signed informed consent. The International Physical
Activity Questionnaire Long Form was used to assess the characteristics of the patients’
physical activities [5-10]. The Hungarian and Romanian versions of the IPAQ were used and
administered by the interviewer (not self-assessed). The questionnaire recorded the frequency,
intensity, and duration of physical activity and the activity type during the previous one-week
period. Regularity was assessed by asking if the patient had regular PA (at least 60 min in every
48-hour period). Patients were presented with a list of activities, including walking, bicycling,
domestic activities, gardening, farming, job-related work including moderate and intense
activity (e.g., carpentry or construction), and leisure and sports activities (patients reported what
type, e.g., fitness, dancing, jogging, or swimming). The patients were asked to outline the
number of hours each type of activity was performed per day and the number of days per week
the activities were performed in the previous week.
To collect exact measurements and to compare the different exercises, the continuous score
for physical activity level (PAL) was calculated using a special formula to provide the average
activity level per hour (considering the duration and energy cost of each activity). The number
of hours spent performing each PA type per week was multiplied by the MET value of the
activity, and the obtained values were summed for one week and divided by the total number
of hours spent performing all activities. The remaining hours were considered to comprise
sitting or sleeping and were assigned an average MET of 1.5. If a job-related activity was
deskwork, it was included as sitting. The physical activity level (PAL) was calculated using the
formula below, considering 168 hours in one week and 1.5 as the average MET for the
“unspecified activity” hours:
PAL = ((168-∑Ah) x1∙5 +∑ Ec x Ah)/168 (Ec = specific energy cost of activity in MET,
Ah = activity hours/week).
Effort was divided into four categories: walking (a), gardening/farming-related activities (b),
job-related moderate or vigorous activities (c), and physical exercise related to sports or leisure
(d). The four categories included PAs with different intensities. A PAL of 1.7 was considered
the cut-off point for a moderate activity level because it was equivalent to the guideline
recommendation of a minimum of 150 minutes of moderate activity/week.
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According to the PAL, patients were divided into five activity levels as follows: inactive
(PAL less than 1.4), sedentary (PAL of 1.4-1.69), moderately active (PAL of 1.70-1.99),
vigorously active (PAL of 2.0-2.4), and extremely active (PAL greater than 2∙4) [6].
Statistical Analyses
Statistical analyses were performed using SPSS statistical software, version 17.0. Data are
presented as the means and standard deviations or as frequencies. The relationship of the
physical activity level as a dependent variable was assessed with age, BMI, and
sociodemographic data such as gender, ethnicity, income, and education level as independent
variables using linear regression and binary logistic regression analyses (with a cut-off point of
1∙7 for the PAL) and cross-tabulation to assess frequencies; risk estimates were calculated.
3. Results
In total, 332 type 2 diabetes patients were included in this study, with only 300 patients
completing comprehensively the questionnaires. The mean BMI was 29.08 (SD 5.52) kg/m2,
68.3% had a lower education level, and 35.3% had a low-income status. The mean PAL was
1.68 (SD 0∙24), and 50.9% of patients reported regular PA. The distribution of patients
according to the different activity types was: 55% of patients were mainly walking, 31.3% were
engaged in gardening and farming, 10.3% reported mostly profession-related activities, and
only 3.3% chose leisure-type activities. 70% of patients performed less than the recommended
level of physical activity (PAL<1.7). Patients were divided into two groups depending on
whether their PAL was less than or greater than the 1.7 threshold. Differences in
sociodemographic characteristics between the two groups are shown in Table 1.
Table 1. Binary logistic regression analysis comparing inadequately and adequately active adults in relation to
sociodemographic characteristics and physical activity type
PAL<1.7 PAL≥1.7 OR P 95%CI

Number 209 91
Age (years, 64. 60.31(9.1) 0.97 0.24 0.93-1.01
mean±SD) (SD 10.1)
Gender: 2.4 0∙01 1.21-5.05
- Men, % 32.1 56
- Women, % 67.9 44
Residence: 3.8 0∙001 1.76-8.46
- Rural, % 42.5 67
- Urban, % 57.5 33
Regular activity
- Yes % 34 90 27.64 0∙001 11.64-65.60
- No % 66 10
Activity type
- Walking 73.6% 10% -0.038 0.001 0.009-0.161
- Gardening 21.2% 57% 1.21 0.27 0.32-4.57
- Professional 3.3% 26.4% 0.43 0.35 0.09-2.00
activity
- Sports, leisure 1.9% 6.6%a
activity
BMI (kg/m2, mean+ SD) 29.54 28.04 0.98 0.23 0.90-1.07
(SD 5.6) (SD 5.04)
RON, Romanian currency, leu
a – sports and leisure activities were considered as referent category
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Using linear regression analysis age was negatively and weekly correlated with activity level
(CI: -0.008 to -0.002 for B -0.005, p=0.000), and individuals with an adequate PAL were
significantly younger (60.31 SD 9.1 vs. 64.19 SD 10.1 years, p<0.001). Men had a higher mean
activity level than women (1.42 SD 0.33 vs. 1.3 SD 0.17, 95% CI 0.05 to 0.17, p<0.000) and
were present in a higher proportion in the PAL >1.7 group (56% vs. 44%, p<0.01, OR 2.4 for
men to be adequately active).
Rural residents had a higher PAL (1.75 vs. 1.61, p<0.000, 99% CI 0.09-0.20), and 67% of
patients with a sufficient activity level resided in a rural area (OR 3.8, 95% CI 1.76-8.46). As
expected, regularly performed activities achieved a higher PAL, and 90% of patients with a
PAL above 1.7 were active daily. Walking could not achieve an efficient activity level, whereas
gardening and professional activities were the best ways to achieve at least a moderate activity
level. Using linear logistic regression BMI weekly and negatively correlated with the PAL,
even following adjustment for age (p<0.002, CI 0.013-0.003 B 0.008), and was higher in the
inactive group, but not in a statistically significant manner (29.54 vs. 28.04, OR 0.9 p=0.2). The
sociodemographic characteristics related to the activity types most often chosen by patients are
summarised in Table 2.
Table 2. Multinomial logistic regression analysis comparing PA type, using sports and leisure activities as
reference and sociodemographic variables
Walking Gardening Professional Sports and leisure
activity activitiesa
Number/% 165/55 94/31 31/10.3 10/3.3
Age (years, mean±SD) 64.2(SD 9.9) 64.9 (SD 9.2)* 53.6 (SD 8.5) 58.0 (SD 5.1)
Gender (for men)
OR 0.61 0.69 1.2
CI 95% 0.1-2.7 0.15-3.1 0.2-6.9
Distribution of activity type
- in men 48% 30% 18% 4%
- in women 59% 31% 6% 3%
Residence (rural resident)
OR 1.9 47.48*** 2.3
CI 95% 0.3-11.8 6.6-338.9 0.3-17.9
Distribution of activity type
- in rural areas 31% 58% 9% 1%
- in urban areas 78% 5% 12% 5%
PAL 1.55 1.82** 1.96** 1.69
(SD 0.08) (SD 0.3) (SD 0.28) (SD 0.09)
Regularity
OR 0.2* 1.8* 0.9
CI 95% 0.04-1.1 0.3-10.6 0.1-6.1
- Distribution of activity
type in patients active
- regularly 36% 46% 13% 3%
- not regularly 74% 15% 8% 3%
BMI (kg/m2) 29. 27.6* 29.0 31∙0
(SD 5.6) (SD 4.7) (SD 6.2) (SD 6.4)
Waist circumference (cm) 105.0 98.0* 100.7 108.4
(SD 17.2) (SD 17.1) (SD 19.9) (SD 15.4)
HbA1c% 7.7 (SD 1.4) 7.8 (SD 1.4) 7.8 (SD 1.7) 7.03 (SD 2.3)
RON Romanian currency, leu
*p<0.05, ***p<0.0001,
a
was used as reference value
b
not calculated because of the low number of cases
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In half of the patients, the most often (or only) performed physical activity was walking, and
one third of patients were involved in gardening activities. Sports and leisure activities were
very unpopular, as they were chosen in only 3.3% of cases. Patients choosing sports or leisure
activities were younger than those who reported gardening or walking (58.6 SD 5.1 vs. 64.9 SD
9.2 years; 95% CI 0.38 to 12.22, p=0.03). There was a very significant age difference (p<0.000)
between the professional activity group and the other three activity groups because the mean
retirement age was 56 years in this cohort.
Gender differences were obvious in profession-related activities, with men being involved
three times more frequently than women. There was a significant relationship between area of
residence and PA type. Gardening activities were performed by 92% of rural residents. Rural
residents did not choose leisure activities. Although walking was the most popular form of
physical activity, gardening and profession-related physical activities achieved the highest PAL
and were practiced the most regularly. HbA1c did not differ significantly between the groups,
BMI, and waist circumference were the most favourable in the gardening group (27.6 SD 4.7
vs. 29.7 SD 5.6, 29.0 SD 6.2, 31.0 SD 6.4, p<0.05, and 98.0 SD 17.1 vs. 105.0 SD 17.2, 100.7
SD 19.9, 108.4 SD 15.4, p<0.05).
4. Discussion
There is a clear “East-West Health Gap” regarding health outcomes that is primarily caused
by socioeconomic differences and unhealthy behaviours [11, 12]. There are differences even
among Eastern European countries regarding PA, as shown by Biddle et al., in a study
conducted among Hungarian, Romanian, and Slovakian youth [12]. To encourage the
recommended amount of physical exercise, the specific local PA patterns must first be
understood. The present study explored the PA pattern in Transylvania for the first time in a
representative sample of diabetes patients. Romania, particularly Transylvania, is a primarily
rural area, with 51.1% of the population of Mureş County living in villages and being active in
gardening, farming, and raising animals. Leisure activities and hobby sports, such as jogging,
biking, swimming, and fitness, are not part of the health behaviour of the general population.
Despite repeated efforts to engage patients in individual or organised physical activities
taking place at the day care centre, only 3% of patients responded positively to the call. In the
studied cohort, 20% of patients were inactive, and an adequate PA level was attained by only
30% of the patients. Compared with reports from different countries and the average population,
this is a very low level of PA [13, 14, 15]. This low level of PA is similar to that of East Asian
countries, such as Japan, China, and Taiwan [13]. The low level of PA in the present study
could also be attributed to the higher mean age, the high percentage of retired persons (where
retirement was the result of age but premature retirement caused by disease was also present),
or the near lack of sport and leisure activities. One small Romanian study [16] reported 30%
inactivity in the general population in a cohort with a lower mean age. PAL was strongly
associated with area of residence and was significantly higher in rural residents (1.75 vs. 1.61).
Two thirds of the patients with an adequate PAL were rural residents, which is in contrast to
most European countries and the USA [17, 18] but similar to reports of Japanese rural residents
[19]. The higher PAL in these patients was related to their involvement in gardening and
farming (activities performed by 92% of patients living in a rural area). Rural life may provide
“bio-psycho-motor” model for sustaining an active lifestyle [20, 21, 22]. Leisure and sports
activities were not practiced by rural residents. In total, 78% of the urban residents chose
walking as their main form of physical activity. Even in urban conditions, only 5% of the
patients were practicing any sports. Sporting facilities are poor, with few available activities,
and few bicycle-friendly roads. Gender differences were as expected, as men were more active;
only one third of the inadequately active population was male [23]. Individuals engaged in
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profession-related activities were typically men. This difference could be partly explained by
the fact that women in rural areas do not take part in professional activities as often as men. The
“beneficial” type or “must” type activities (e.g., gardening, farming, professional activities) had
the highest PAL and were performed regularly. Gardening and farming were primarily
performed in rural areas and were almost non-existent in urban communities. Sports and leisure
activities were almost non-existent (3%), which was confirmed by a European survey of the
general population of Romania that reported regular sports activities in 6% of the population;
this value did not differ significantly from that in Hungary, where the proportion was 5%.
Romania was at the bottom of the survey regarding the percentage of people choosing to use
fitness centres and sports clubs. A low proportion of people in Romania (43%) and Hungary
(49%) said they were motivated to perform PA by health issues or to improve their fitness level.
Romanians listed lack of time (top of EU countries) and lack of sport facilities as the main
causes of inactivity [24].
The strengths of the present study include the investigation of a homogenous, representative
cohort of diabetic patients in a restricted region. In addition to using the well-defined IPAQ, the
questions in the present study focused on different types of activities.
Additionally, direct questioning was used to diminish the limitations of a self-assessment
questionnaire and the risk of under- or overestimating the PA or of patients not fully
understanding the questions.
This study is limited by the estimation of PA and the MET values for certain activities, which
is a bias that occurs in cases of self-reported physical activity and can produce variation of
+10% from the objective energy expenditure measurements. However, the same measure was
used for all patients, which indicates that the study has comparative power.
5. Conclusion
Adult patients with type 2 diabetes had very low PALs, and only 30% of patients achieved
an adequate exercise level. The relatively high percentage of rural inhabitants and the
characteristics of local rural life, as well as the lack of a physical exercise culture and exercise
facilities, may explain the differences. Rural residents and men had higher PALs and reported
more regular PA. Walking and gardening were the most frequent types of activities, but only
the latter was performed regularly, and for insufficient durations. In conclusion, education in
our region must focus on urban inhabitants, women. Walking and gardening should be
encouraged, but walking habits must be discussed to achieve an adequate PAL. The control and
reassessment of the PAL are indispensable and are necessary to address PA in an individualised
manner.
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Fast-food and Diabetes in Paediatrics – a Road to Disaster
MIHAI Cristina Maria1, PINZARU Anca Daniela1, CHISNOIU Tatiana1

1Clinic of Pediatry, Faculty of Medicine, ‘Ovidius’ University, Constanta (ROMANIA)
Email: tatiana_ceafcu@yahoo.com
Abstract
A balanced nutrition during life is the key of a healthy existence. It becomes more important
if it involves children with type 1 diabetes. The lack of medical and nutrition education as well
as poverty could lead to excessive food eating, unbalanced diets, obesity, poor metabolic
control, hyperglycaemia. The connection between fast-food eating in children with type 1
diabetes and a poor glycaemic control was not yet established in our country.
During a 1-year retrospective study (January 1st 2019-January 1st 2020) we evaluated 38
patients already diagnosticated with type 1 diabetes (aged 7-16-year-old) admitted in the
Pediatric Department of the Clinical County Emergency Hospital of Constanta for
hyperglycaemia during insulin treatment. The main inclusion criteria remained unproper meals
(fast-food type) followed by abdominal pain, vomiting, diarrhoea and hyperglycaemia. The
nutrition evaluation underlined that: at least once a week the analysed patients elated a fast-
food meal containing French fries, hamburgers, acidulates beverages. 89% consumed fizzy
drinks daily, 63% had a meal daily consisting in pastry, 26% did not respect the meal hours or
the insulin injection program. Combined with unproper meals it could lead to an early
complication onset, increasing the risk of death among teenagers with type 1 diabetes.
Keywords: obesity, insulin, control, pediatric
1. Aims and Background
The main purpose of nutrition management and eating habits it to create a balance between
food intake – exogenous insulin dosage and glycaemic control. A balanced nutrition during life
is the key of a healthy existence. It becomes more important if it involves children with type 1
diabetes. The lack of medical and nutrition education as well as poverty could lead to excessive
food eating, unbalanced diets, obesity, poor metabolic control, hyperglycaemia.
Medical nutrition therapy (MNT) is an important aspect of diabetes self-care management
and diabetes self-care education [1]. The American Diabetes Association position statement on
MNT includes the following goals:
• maintaining optimal metabolic outcomes (glucose and lipid levels);
• prevention and treatment of chronic complications of diabetes;
• improvement of health through healthy food choices and physical activity;
• taking individual nutritional needs into consideration [1].
Choosing appropriate foods and food portion sizes is integral to achieving these goals, as is
meeting the individual needs of clients.
Junk foods may contribute to diabetes in the following ways:
• Rapid effect on blood sugar levels: Highly processed foods that are high in calories and
low in vitamins, minerals, and fiber break down quickly in the body and can cause a
rapid rise in blood sugar levels;
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• Inappropriate portion size: Junk foods are usually not very filling and frequently come
in large portion sizes. Both these factors may lead people to overeat junk foods. This
can have a negative impact on diabetes, including blood sugar spikes and weight gain;
• Weight gain: Due to its poor nutritional qualities and ability to encourage overeating,
people who eat junk food may gain weight. Excess weight and body fat are major risk
factors for developing type 2 diabetes, which accounts for 90-95 percent of all cases of
diabetes;
• High blood pressure. Junk food is usually very high in sodium (salt), which contributes
to high blood pressure. High blood pressure is linked to an increased risk of type 2
diabetes;
• Triglyceride levels. Junk foods are high in trans and saturated fats, which can raise levels
of triglycerides, a type of fat that is present in the blood. High levels of triglycerides
increase the risk of developing type 2 diabetes.
Romania is still a developing country and the dietary influences of the modern civilization
could be notice at every corner of every street. We eat more for less money. But we stopped
eating healthy. The main concern appears in chronic patients thar require calculated
carbohydrates intake. The fatty composition of the food influents the carbohydrates absorption
rate, therefor hyperglycaemic pick could be delated. The conscience: silent hyperglycaemia or
the short path to complications
During a 1-year retrospective study (January 1st 2019-January 1st 2020) we evaluated 38
patients already diagnosticated with type 1 diabetes (aged 7-16-year-old) admitted in the
Pediatric Department of the Clinical County Emergency Hospital of Constanta for
hyperglycaemia during insulin treatment. There were exclude the ketoacidosis episode
determined by other factors, such as: underlying diseases (celiac disease, flu-like episodes).
The main inclusion criteria remained: unproper meals (fast-food type) followed by
abdominal pain, vomiting, diarrhoea and hyperglycaemia, unproper insulin intake and
glycaemic monitoring. In order to respect the well going of our study mandatory documents
were offered and signed before starting the study: parents inform consent, criteria for
inclusion/exclusion, anamnesis, clinical examination, hospital approval, the proof of type 1
diabetes mellitus already established.
A male predisposition could be notice (28 males: 10 females) as well as the predominant
involvement of the teenagers (age distribution 13-16-year-old: 30 cases). What is important to
underline is the involvement of teenagers. During the first period of life the parents have a
greater control. During time, especially during adolescence, this control becomes weaker.
This is the moment when they want to fell free. A chronic disease such as diabetes requires
glycaemic control, calculated carbohydrates intake, monitoring dietary combinations,
excluding bad habits (such as smoking) and so on. Teenagers want to fell normal, to eat like
their friends, to stay up all night, to party. They start a “war” against their disease and their
parents. Most often they start by eating in large amounts than usual and secretly increasing their
insulin dosage. 79% admitted increasing their insulin dosage in order to eat more, especially
during their time out with friends. 3% of them refused to announce friends about their disease.
The consequences of this type of behaviour is a chaotic meal schedule, with high blood
sugar, weight gain and abnormal metabolic control.
317
The second interesting discovery is related to living environment. Urban area is represented
by 98% of the patients. Poverty leads to consuming large amounts of cheap food (potatoes,
bread). It seems that in rural areas even if the economic status is very low the parents of the
patients could provide more healthier opportunities (vegetables, meat) produced in their own
homes. In the cities the situation is more complex. In every corner they could find a fast-food
restaurant or pastries. We could underline that the urban area is more likely to provide unhealthy
food.
Dietary habits play an essential role in glycaemic control in children with Type 1 Diabetes.
The results of the Diabetes Control and Complications Trial (DCCT) show that, among
patients undergoing intensive glycemic control, consumption of low-carbohydrate, high-fat
(particularly high-saturated fat) diets is associated with poorer glycemic control, regardless of
exercise and serum triglyceride levels [2].
We completed nutrition forms in which every patient had to write down their favourite type
of food. 56% admitted that they would like to eat every day French fries or some sort of pastries
products. The nutrition evaluation underlined that: at least once a week the analysed patients
eated a fast-food meal containing French fries, hamburgers, acidulates beverages. 89%
consumed fizzy drinks daily, 63% had a meal daily consisting in pastry, 26% did not respect
the meal hours or the insulin injection program.
Dyslipidaemia is a common finding in T1DM, possibly due to increased cholesterol
absorption [3], [5]. In patients who follow dietary recommendations, the relevance of metabolic
control in the management of dyslipidaemia goes beyond mere reductions in lipid intake.
Targets for dietary management should ensure adequate intake of fiber and carbohydrates,
which have an impact on glycated haemoglobin (HbA1c) levels and, consequently, on
metabolic control [4].
In our study, the glycemic control was mediocre to poor in all cases, with a median of HbA1c
9.5%. The insulin exogenous apport had a median value of 0,9 UI/Kg/day/patient.
A review article published in 2016 suggested that trans fats could have a negative impact on
insulin sensitivity, but the study author notes that more research is necessary [6].
Sources of trans fats include:
• crackers and chips;
• cookies;
• fast food items, including fries;
• hydrogenated oil or partially hydrogenated oil;
• margarines;
• muffins and cakes;
• shortening.
4. Conclusions
Exogenous insulin dosages should be increased during fatty meals with high caloric and
large amounts of carbohydrates. Insulin is an anabolic hormone that leads to weight gain.
Combined with unproper meals it could lead to an early complication onset, increasing the
risk of death among teenagers with type 1 diabetes.
Diabetes educators need to learn where they should focus their efforts in nutrition education.
Diabetes educators have limited interaction time with patients to teach the numerous aspects
of diabetes care. It is essential that diabetes educators focus on the important and useful aspects
of nutrition that will be helpful to improving diabetes control [7].
318
REFERENCES
1. American Diabetes Association, 2004: Nutrition principles and recommendations in diabetes. Diabetes
Care 27(Suppl. 1): pp. S36-S46.
2. Delahanty LM, Nathan DM, Lachin JM, Hu FB, Cleary PA, Ziegler GK et al., Association of diet with
glycated haemoglobin during intensive treatment of type 1 diabetes in the Diabetes Control and
Complications Trial. Am J Clin Nutr 2009; 89: pp. 518-24.
3. Wiltshire EJ, Hirte C, Couper JJ. Dietary fats do not contribute to hyperlipidaemia in children and
adolescents with type 1 diabetes. Diabetes Care 2003; 26: pp. 1356-61.
4. Gylling H, Tuominen JA, Koivisto VA, Miettinen TA. Cholesterol metabolism in type 1 diabetes.
Diabetes 2004; 53: pp. 2217-22.
5. Sheylle Almeida S. Teles; Nélida Schmid Fornés. Food consumption and metabolic control in children
and adolescents with type 1 diabetes mellitus, Rev Paulista de Pediatria, ISSN 0103-0582, vol 29, no 3,
Sept 2011
6. Ulf Riserus, Trans fatty acids, insulin sensitivity and type 2 diabetes, Scandinavian Journal of Food and
Nutrition 2006; 50(4): p. 161.
7. Nedra K. Christensen, PhD, RD, Elaine Boswell King, MSN, APRN, CDE, Sherrie Hardy, RD, MS, CDE
and Roxane Pfister, MS Food Practices and Preferences in Youth with Diabetes, Diabetes Spectrum 2007
Jan; 20(1): pp. 33-39.
319
Chemical – Toxicological Research on the Evaluation of Serum

Malondialdehide Levels in Patients with Diabetic Arteriopathy
IONIȚĂ Ana Corina1, NICOLESCU Teodor Octavian2*, IONIȚĂ Elena Iuliana3,

NICOLESCU Florica4, MITITELU Magdalena1
1 Department of Clinical Laboratory and Food Hygiene, Faculty of Pharmacy, “Carol Davila” University of Medicine and
2 Department of Organic Chemistry, Faculty of Pharmacy, “Carol Davila” University of Medicine and Pharmacy, Bucharest
(ROMANIA)
3 Department of Pharmacognosy, Phytochemistry, Fitotherapy, Faculty of Pharmacy, “Carol Davila” University of Medicine
and Pharmacy, Bucharest (ROMANIA)

4 Department of Toxicology, Faculty of Pharmacy, “Carol Davila” University of Medicine and Pharmacy, Bucharest
(ROMANIA)
Email: nicolescu.teodor@gmail.com
Abstract
Oxidative stress is associated with numerous chronic diseases. Increased oxidative stress as
well as reduced antioxidant capacity may be complications of type 2 diabetes. This study
highlights the role that oxidative stress can play in diabetes and cardiovascular disease and how
it can be measured in plasma. The purpose of the work is to evaluate and interpret the plasma
level of malondialdehyde (MDA), which is a marker of lipid peroxidation, in patients suffering
from diabetic arteriopathy, but also in healthy patients. The aim was also to identify the
correlation between the level of biomarker studied (MDA) and the classical biochemical
parameters, which illustrate the general metabolic status of the patient (triglycerides,
cholesterol, glucose).
Keywords: oxidative stress, diabetic arteriopathy, malondialdehyde, lipid peroxidation
1. Introduction
Diabetes is an important cause of morbidity, mortality and economic costs in society.

Patients with diabetes have the risk of developing acute metabolic complications such as:
diabetic ketoacidosis, hypoglycaemia. In addition, diabetes presents the risk of chronic
complications, for example: coronary heart disease, retinopathy, nephropathy, neuropathy and
foot ulcers. Many factors influence diabetic pathologies [1].
Insulin resistance that results from obesity and physical inactivity, works as a substrate for
genetic susceptibility. Because food intake influences the amount of insulin required to meet
the goals of blood glucose, it, especially carbohydrates, can contribute to diabetic pathologies.
Carbohydrates most influence postprandial blood glucose level, which is the major
determinant of insulin requirement as a result of food ingestion. The intermediate or long action
of insulin usually covers the effect of proteins and fats [2, 3].
Ketogenic diets with a low carbohydrate content have been shown to be helpful and
ameliorate a number of harmful consequences of diabetes. It has also been observed that insulin
secretion decreases with age and this decrease may be accelerated by genetic factors [4]. Insulin
resistance usually occurs at the onset of type 2 diabetes and is frequently accompanied by other
cardiovascular risk factors: dyslipidaemia, hypertension and prothrombotic factors [5].
Expenditures are four times higher for a diabetic patient compared to the healthy population.
320
Prospective studies have demonstrated the crucial role of hyperglycaemia in the chronic
development of diabetic complications.
Of considerable interest was the harmful role of free radicals in many pathologies, especially
in cardiovascular diseases, diabetes and cancer. Free radicals are represented by atoms or
molecules that have one or more electrons unmatched in their structure and are therefore highly
reactive species [6].
Oxidative stress is intensively studied and can be a key factor not only in diabetes, but in
many other diseases, with implications throughout the body.
The body tries to protect itself from oxidative stress through antioxidants (glutathione,
catalase, superoxide dismutase, plasma antioxidant proteins).
High glucose concentrations can stimulate the production of free radicals, and a low defense
of the body will not be able to counteract the effect of ROS (reactive oxygen species), as a result
there is an imbalance between the amount of ROS and the compounds that protect the body,
leading to a domination determined by ROS with important consequences [7, 8].
Of the most biologically important chemical species, oxygen is ubiquitous and is a major
source of reactive oxygen species (ROS). It has been estimated that up to 5% of inhaled oxygen
becomes a reactive species.
Oxidative stress occurs as a result of an imbalance between the production of oxidizing
compounds and antioxidants with a defense role [9].
Within a certain range (physiological limits), ROS are required at the cellular level for
normal functioning, and exaggerated production leads to oxidative stress that contributes to the
aging and development of pathologies [10].
Oxidative stress plays an extremely important role in cellular damage due to
hyperglycaemia. High concentrations of glucose can stimulate the production of free radicals,
and a low defense of the body will not be able to counteract the effect of ROS (reactive oxygen
species), as a result there is an imbalance between the amount of ROS and the compounds that
protect the body, leading to a domination determined by ROS with important consequences.
Several studies have shown that hyperglycaemia is associated with an increase in oxidative
stress and therefore may lead to a possible link between diabetes and atherosclerosis. This fact
is supported by the observation of the plasma of diabetes that will contain increased levels of
finished oxidative products, including reactive thiobarbituric acid (TBARS) and lipid peroxides
[11, 12].
A genetic study of a family whose members suffer from hypomagnesemia, hypertension and
hypercholesterolemia, shows that the disease has a maternal lineage and the mutation is
mitochondrial-encoded in tRNA, providing a direct link between mitochondrial function and
hypertension [13, 14].
Malondialdehyde (MDA) is one of the most widely used biomarkers evaluating the effect of
lipid peroxidation, as MDA results from this reaction. Following its production, the biomarker
can bind to amino groups of proteins, phospholipids and/or nucleic acids, causing numerous
structural changes and also inducing, as a final effect, cell dysfunction.
The MDA measurement is performed according to the Esterbauer and Cheeseman method
[15, 16, 17, 18], being easy, fast, economical. It involves the impairment of the erythrocyte
membrane, which is subjected to lipid peroxidation and occurs by cleavage of polyunsaturated
fatty acids from the double bonds, thus forming MDA in the plasma, which is easily identified
and then measured [19, 20, 21].
The MDA level can be correlated with Total Antioxidant Status (TAS), which represents
plasma antioxidant capacity, but also with age, as aging leads to a decrease in plasma
antioxidant capacity thus increasing the MDA level, confirming the hypothesis that aging
occurs with decreasing antioxidant capacity of the body and implicitly with an increase in
oxidative stress [22, 23, 24, 25].
321
2. Methodology
43 Patients from the National Institute of Diabetes and Metabolic Diseases were selected
“Prof. Dr. N. Paulescu”, which were divided into two study groups:
• group 1: patients without diabetic arteriopathy (n=18)
• group 2: patients with diabetic arteriopathy (n=25).
Patients underwent venous blood test, à jeun, and then the following determinations were
made:
• classical biochemical parameters, which will highlight the overall metabolic status of
patients (triglycerides, total cholesterol, glycemia),
• the level of malondialdehyde in the plasma, which will indicate the degree of lipid
peroxidation.
Principle of the method

Serum malondialdehyde (MDA) reacts with thiobarbituric acid (TBA) resulting in the
formation of a pink coloured adduct, which will be appreciated spectrophotometric intensity at
a wavelength of 532 nm (scheme 1)
Scheme 1. The reaction of MDA (malondialdehyde) and TBA (thiobarbituric acid)
Reagents Used
• 10% aqueous trichloroacetic acid (ATA) solution
• 0.67% aqueous thiobarbituric acid (TBA) solution
Working Technique
Over 700 μL human serum, 700 μL of distilled water will be added and the mixture is
centrifuged for 10 minutes at 3,500 rotations/minute.
Over 700 µL of the supernatant obtained will be added 700 µL of 0.67% TBA, and the
solution obtained is boiled for 17 minutes in a water bath.
The pink adduct is formed, the intensity of which is determined spectrophotometrically at a
wavelength equal to 535 nm, using as a 0.67% TBA control solution.
3. Results
Table 1 shows the obtained values. Values include biochemical parameters for patients who
have been selected.
322
Table 1. The values of the biochemical parameters obtained in the patients

that were included in the study
Group of patients with

The biochemical Group of patients without
diabetic arteriopathy p
parameters investigated diabetic arteriopathy (n=18)
(n = 25)
Total cholesterol
189.98±38.72 171.83±35.88 SI
(mg/dL)
Glycemia (mg/dL) 139.29±41.56 186.56±94.03 0.017
Triglycerides(mg/dL) 112.22±82.49 136.23±101.86 SI
MDA (µM) 4.55±2.89 6.43±3.08 0.001

SI = statistically insignificant
Following the analysis of the data it was found that both groups of patients (with and without
diabetic arteriopathy) had values within normal limits of the parameters of evaluation of lipid
metabolism: cholesterol, triglycerides, respectively.
At the same time, high values are observed in both groups of patients, regarding the value of
glucose, respectively of malondialdehyde, their concentrations exceeding the normal values
indicated by the specialized literature.
In order to better observe the data presented above, we have presented in the following
figures the dynamics of lipid parameters (cholesterol, triglycerides), in the two groups of
patients in the study (Fig. 1), as well as the dynamics of serum MDA in patients with diabetic
arteriopathy, in comparison with those without diabetic arteriopathy (Fig. 2).
Dynamics of Lipid Parameters

From the analysis of the data presented in Fig. 1, there are no notable changes in lipid
metabolism. Because of this, the parameters cannot be a predictive method for diagnosing
diabetic arteriopathy in patients who are in the early stages of diabetes.
Fig. 1. Dynamics of lipid parameters

(blue-group of patients without arteriopathy; red-group of patients with arteriopathy)
Dynamics of lipid parameters

200
189,98 171,83
136,23
100 112,22
0
Cholesterol Triglycerides
Dynamics of malondialdehyde (MDA)

From the analysis of the data obtained, there is an increase of the serum MDA values in the
group of patients with diabetic arteriopathy (Fig. 2).
323
Fig. 2. Dynamics of malondialdehyde (MDA)

(blue-group of patients without arteriopathy; red-group of patients with arteriopathy)
From the analysis of the presented data, it was observed that the level of triglycerides and
cholesterol did not change significantly in the two groups of patients. The consequence of this
fact has determined us to appreciate the inability to use these parameters to identify diabetic
arteriopathy.
On the other hand, it is observed from the data presented that the level of MDA increases
progressively, with the installation of diabetic retinopathy, and the obtained results can support
this claim.
Thus, we believe that serum MDA determination could be used for early detection of
vascular disturbances that occur in diabetic patients.
4. Conclusions
Diabetes is an important pathology nowadays, which can lead to complications in the various
organs (blood vessels, myocardium, eyes, foot, kidneys, brain), making the patient's life more
difficult.
Oxidative stress is intensively studied and can be a key factor not only in diabetes, but in
many other diseases, with implications for the entire body. Increased oxidative stress as well as
reduced antioxidant capacity may be complications of type 2 diabetes.
MDA is considered the first biomarker of free radicals that produce lipid damage and cause
oxidative stress.
The level of MDA increases progressively, with the installation of diabetic arteriopathy, the
results obtained in the experimental part supporting this claim.
Determination of serum MDA could be used for early detection of vascular disturbances that
occur in diabetic patients.
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1. Varashree, B. S., Bhat, P. G. (2011). Correlation of lipid peroxidation with glycated haemoglobin levels
in diabetes mellitus, Online Journal of Health and Allied Sciences 10(2), pp. 1-4.
2. Paneni, F., Beckman, J. A., Creager, M. A., and Consetino, F. (2013). Diabetes and vascular disease:
pathophysiology, clinical consequences, and medical therapy: part I. European Heart Journal 34(31), pp.
2436-2443.
3. Tsai, C.J., Hsieh, C. J., Tung, S. C., Kuo, M. C., and Shen, F. C. (2012). Acute blood glucose fluctuations
can decrease blood glutathione and adiponectin levels in patients with type 2 diabetes. Diabetes Research
and Clinical Practice 98(2), pp. 257-263.
4. Drăgoi, C. M., Moroșan, E., Dumitrescu, I.B., Nicolae, A. C., Arsene, A. L., Drăgănescu, D., Lupuliasa,
D., Ioniță, A. C., Pantea A., Stoian, C. Nicolae, Rizzo, M., Mititelu, M. (2019). Insights into
324
stroke: a rewiew of pharmacokinetics and pharmacodynamic studies. Nutrients 11 (7), pp. 1479-1513.
6. Ceriello, A., Motz, E. (2004). Is oxidative stress the pathogenic mechanism underlying insulin resistance,
diabetes, and cardiovascular disease? The common soil hypothesis revisited. Arteriosclerosis,
Thrombosis and Vascular Biology 24(5), pp. 816-823.
7. Chiang, J.L., Kirkman M.S., Laffel L.M., et al., (2014). Type 1 diabetes through the life span: a position
statement of the American Diabetes Association. Diabetes Care 37(7), pp. 2034-2054.
8. Grădinaru, D., Borșa, C., Ionescu, C., and Margină, D. (2013). Advanced oxidative and glycoxidative
protein damage markers in the elderly with type 2 diabetes. Journal of Proteomics 13, pp. 181-184.
9. Evans, J.L., Goldfine, I.D., Maddux, B.A., Grodsky, G.M. (2002). Oxidative stress and stress-activated
signaling pathways; a unifying hypothesis of type 2 diabetes. Endocrine Reviews 23(5), pp. 599-622.
10. Korkmaz, G., Konukoglu, D., Kurtulos, E. M., Irmak, H., Bolayirli, M., and Uzun, H. (2013). Total
antioxidant status and markers of oxidative stress in subjects with normal or impaired glucose regulation
(IFG, IGT) in diabetic patients. Scandinavian Journal of Clinical and Laboratory Investigation 73(8), pp.
641-649.
11. Pandey, K.B., Rizvi, S.I. (2011). Biomarkers of oxidative stress in red blood cells. Biomedical Papers
155(2), pp.131-136
12. Morrow, J.D. (2005). Quantification of isoprostanes as indices of oxidant stress and the risk of
atherosclerosis in humans. Arteriosclerosis, Thrombosis, and Vascular Biology 25(2), pp. 279-286
13. Singh, D.K., Winocour, P., Farrington, K., (2011). Oxidative stress in early diabetic nephropathy: fuelling
the fire”, Nature Reviews. Endocrinology 7(3), pp. 176-184.
14. Wang, X., Tao L., and Hai, C. X. (2012). Redox-regulating role of insulin: the essence of insulin effect.
Molecular and Cellular Endocrinology, 349(2), pp. 111-127.
15. Esterbauer, H., Cheeseman, K. H. (1990). Determination of Aldehydic Lipid Peroxidation Products:
Malonaldehyde and 4-Hydroxynonenal. Methods in Enzymology 186, pp. 407-421.
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Polluants From Some Fish of Danube River. Journal of Enviromental Protection and Ecology 13(2), pp.
869-874.
17. Iova, A., Micle, O., Vicaș, L., Micle, L., Iova, S., Muresan, M., Ionita, C. A. (2014). Oxidative stress in
Alzheimer’s Dementia. Farmacia 62(3), pp. 538-546.
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synthesized prostamides on human hepatocellular carcinoma cell line (hepg2), Farmacia. 59(5), pp. 611-
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Farmacia 67(4), pp. 684-690.
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Black Sea. Farmacia 62(3), pp. 625-632.
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products. Farmacia 63(1), pp. 118-122.
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de Chimie 54 (8), pp. 676-680.
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some new quinolone ruthenium (III) complexes with potential cytostatic activity. Journal of Thermal
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325
Research on the Hypolipidemic Action of Almond Oil and

Almond Seeds
IONIȚĂ Ana Corina1, MITITELU Magdalena1*,

NICOLESCU Teodor Octavian2, NICOLESCU Florica3, MOROȘAN Elena1,
OZON Emma Adriana4, IONIȚĂ Elena Iuliana5
(ROMANIA)
(ROMANIA)
4 Department of Pharmaceutical Technology, Faculty of Pharmacy, “Carol Davila” University of Medicine and Pharmacy,
Bucharest (ROMANIA)
5 Department of Pharmacognosy, Phytochemistry, Fitotherapy, Faculty of Pharmacy, “Carol Davila” University of Medicine
and Pharmacy, Bucharest (ROMANIA)

Email: magdamititelu@yahoo.com
Abstract
The therapeutic approach of the disorders of lipid and cardiovascular metabolism starts with
an adequate diet and with the change of lifestyle. Because of this, dyslipidaemia is trying to use
different products obtained from plants to normalize the increased plasma levels of lipids. In
the experimental part of the paper, the hypolipemic action of almond oil and almond seed was
investigated in batches of white male mice, the Wistar breed, in which an increase of serum
lipids was induced by the hyperlipidic diet. The determinations were made in comparison with
a control group. The following biochemical parameters were determined: glucose, serum
triglycerides, total cholesterol and LDL-cholesterol.
Keywords: dyslipidaemia, serum lipids, almond oil, almond seeds
1. Introduction
Dyslipidaemia is one of the conditions that contribute significantly to the development or

exacerbation of cardiovascular disease, especially atherosclerosis. Dyslipidaemia is
characterized by increased blood cholesterol and/or triglycerides. Also, it can be characterized
and too low HDL cholesterol, which contributes to the initiation and development of
atherosclerosis [1, 2, 3].
Dyslipidaemia not cause, usually no obvious symptoms. Many people are alarmed only
when you get to the doctor for routine investigations and is observed by specific analyses, a
high cholesterol level. In very advanced cases, however, the large amount of lipids (fats) and
higher levels of triglycerides in the blood can cause fat deposits in the skin, enlarged liver or
spleen, various neurological disorders (manifested by tingling in hands and feet, for example)
and even pancreatitis [4, 5].
People with high levels of serum lipids, which are associated with other diseases (diabetes,
kidney disease, genetic history of atherosclerosis) have an increased risk of heart attack or
stroke [6, 7].
In the first instance, the therapy involves reducing the weight (body weight), a saturated fat
diet and exercise. If such changes do not work, they resort to drugs to reduce blood lipid levels.
326
The diagnosis of dyslipidaemia is given by a doctor following a set of blood tests.

Blood test called lipid profile is the one that helps the doctor to diagnose properly. The lipid
profile includes total cholesterol, triglycerides, HDL and LDL cholesterol, so all the
information the doctor needs [8, 9, 10].
Dyslipidaemia can cause fat deposits in the arteries, which can cause serious problems such
as heart attacks or stroke, so that patients suffering from this disease should do everything
possible to reduce cholesterol and triglycerides in the blood [11].
Nutrition for patients with dyslipidaemia should be based on:
• Soluble fiber-soluble fiber-rich foods such as grapefruit or oat bran necessarily have to
do necessarily part of the diet of those with dyslipidaemia;
• Raw fruits and vegetables low in carbohydrates – spinach, broccoli, onions and all green
leafy vegetables are recommended for patients with dyslipidaemia, they reduce “bad”
cholesterol;
• Omega-3 – one dose of 6-12 grams of omega-3 fatty acids per day can reduce blood
triglycerides by 40%. These fatty acids are found in foods such as marine lipids.
Adopting a healthy diet and an active lifestyle, from sport to take part, they are essential
for lowering cholesterol.
The patient suffering from dyslipidaemia should limit consumption:
• Foods high in cholesterol, such as meat or dairy products;
• Saturated fat is found in foods such as butter, ice cream, dairy products high in fat, meat,
fat, confectionery and pastry;
• Alcohol is known for its negative effect on triglycerides, so it is crucial that patients
who are diagnosed with dyslipidaemia limit alcohol consumption.
The use of natural treatments to reduce serum lipids should take into account the degree of
pollution of the environment from which the plants are harvested to avoid possible poisoning
with contaminants [12, 13, 14, 15].
Dietary advice should be incorporated into the patient's lifestyle in order to gain and maintain
adequate body weight and to perform sufficient regular physical activity [16, 17, 18].
Regardless of the type of dyslipidaemia, correction of plasma lipid level is not the only
dietary factor we should focus on. It is important that the overall diet be balanced and help
reduce cardiovascular risk [19, 20, 21, 22]. Therefore, the advice should be given in the context
of a balance between a healthy diet and a cardioprotective diet. In this context, the things to be
followed in the diet should be emphasized according to the particular type of dyslipidaemia.
2. Methodology
Hypolimemiant action of almond oil and almond seed was investigated on batches of male
white mice, the Wistar breed, in which an increase of serum lipids was induced by the
hyperlipidic diet. The determinations were made in comparison with a control group.
For the study, batches of white male Wistar rats, with an average weight of 177.5 g, were
used. The clinically healthy animals brought from the farm were kept in our laboratory
conditions for 2 days in order to get used to with the new habitat and the diet. The feeding
regime consisted of feeding the animals at 8.00 and 17.00 and water “ad libitum” from bottles.
During the first two days, feeding was done only with special food for rodents. The animals
were kept in mobile cages in accordance with the requirements of the European Convention on
the Protection of Animals.
During the experiment, the animals were fed for 3 weeks with butter and white chocolate,
and at 2 days intervals specific food for rats. These foods have been used to increase their body
fat. After the 3 weeks, before 3 hours of the first administration, 3 groups of animals, of 8 rats
each were made. Lot I – the lot treated with almond seeds Lot II – the lot treated with almond
327
oil Witness batch Two batches, the test batches, were given almond oil (for each rat given the
equivalent of 10 ml/100g) and seeds of almond (for each rat given the equivalent of 10
mg/100g), being administered in both p.o. cases, daily, for 21 days.
One batch was the control group that received only butter and white chocolate. The treatment
lasted 3 weeks, and during it, continued administration of butter and white chocolate and at
intervals of 2 days specific food for rats. Two hours after the last administration, the animals
were sacrificed by guillotine, blood was collected, and the serum separated by centrifugation
was used to determine the biochemical parameters (glucose, total cholesterol, serum
triglycerides and LDL-cholesterol). The effect variations were calculated at each moment,
compared to the initial moment, according to the formula:
E% = (Ex-E0)/E0 x 100
where:
E0 = the initial effect of the witness
Ex = the measured effect for each component studied
Determination of glucose, serum triglycerides and total cholesterol was done by the
enzymatic Cormay method. The determination of LDL-cholesterol was done by direct method
[23].
3. Results
The results of the clinical investigations are presented in tables 1-4 and fig.1-4.
Table 1. The influence of almonds on serum glucose

Glucose (mg/dL)
Tested product Effect % related to the control
Average group
Control group 120.92 -
Group (batch) I 113.66 -6.00
Group (batch) II 112.26 -7.16
Fig. 1. The influence of almonds on serum glucose (mg/dL)
0
Lot I Lot II
-5
-10
Table 2. The influence of almonds on serum triglycerides

Serum triglycerides (mg/dL)
Tested product Effect % related to the
Average control group
Group (batch) I 80.19 -5.58
328
Fig. 2. The influence of almonds on serum triglycerides (TG, mg/dL)

The influence of almonds on serum
triglycerides
0
Lot I Lot II
-2
-4
TG mg/dL
-6
-8
Table 3. The influence of almonds on total serum cholesterol

Cholesterol (mg/dL)
Tested product Effect % related to the
Average control group
Group (batch) I 77.23 -5.98
Fig. 3. The influence of almonds on total serum cholesterol (mg/dL)

The influence of almonds on total
0 serum cholesterol
-2 Lot I Lot II
Colesterol
-4
mg/dL
-6
-8
-10
Table 4. The influence of almonds on LDL-serum cholesterol

Tested product LDL-c (mg/dL)
Effect % related to
Average the control group
LOT I 4.52 -3.62
LOT II 4.48 -4.47
LDL-c = LDL-serum cholesterol
Fig. 4. The influence of almonds on LDL-serum cholesterol (LDL-c, mg/dL)
The influence of almonds on LDL-

0 serum cholesterol
Lot I Lot II
-2
-4 LDL-c mg/dL
-6
Due to the omega-3 fatty acids contained, almonds are beneficial for the cardiovascular
system: regulating cholesterol and lowering blood pressure.
329
From the analysis of the data obtained, a decrease in the value of all the parameters followed
during the experiment (glucose, total cholesterol, LDL-cholesterol, serum triglycerides) was
observed, both for the almond seeds and for the almond oil.
The experimental results show a more intense hypolipidemic effect for almond oil than for
almond seeds. Almonds specifically contain flavonoid-based antioxidants, which act together
with vitamin E (an active antioxidant that is abundantly found in almonds) to strengthen the
arteries and reduce inflammation.
Almonds have been shown to have a constant effect of lowering LDL-cholesterol (“bad
cholesterol”), both in healthy people and in people who have high cholesterol or who have
diabetes.
4. Conclusions
Dyslipidaemia is often asymptomatic and therefore often remains undiagnosed. Specific

signs and symptoms often occur only when the increase in blood lipid level is quite severe.
Diet plays a central role in the management of dyslipidaemia as an adjunctive method for
drug treatment and is sometimes even used as a first-line treatment.
Clinical investigations in laboratory mice fed a hyperlipidic diet and then treated with
almond oil and almond seeds revealed a certain hypolipidemic effect for the plant products
used. A decrease in the value of all parameters followed during the experiment (blood glucose,
total cholesterol, LDL-cholesterol, serum triglycerides) was observed, both for the almond
seeds and for the almond oil. Almond oil showed the most intense hypolipidemic effect.
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1. Ballantyne, C. M, Grundy, S. M, Oberman, A, et al., (2000). Hyperlipidaemia: diagnostic and therapeutic

perspectives. J Clin Endocrinol Metab 85(6), pp. 2089-2112
2. Packard, C. J., Ford, I. (2015). Long-term follow-up of lipid-lowering trials. Curr Opin Lipidol 26(6), pp.
572-579.
3. Yusuf, S., Bosch, J., Dagenais, G., et al., (2016). Cholesterol Lowering in Intermediate-Risk Persons
without Cardiovascular Disease. N Engl J Med 374 (21), pp. 2021-2031.
4. Denke, M. A., Adams-Huet, B., Nguyen, A. T. (2000). Individual cholesterol variation in response to a
margarine- or butter-based diet: A study in families. JAMA 284(21), pp. 2740-2747.
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better? A randomized, controlled trial. Am J Med 109(7), pp. 549-555.
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of inclusion complexes formed between simvastatin and hydroxypropyl-β-cyclodextrin. Farmacia 59(4),
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7. Sbora, R., Budura, E. A., Niţulescu, G. M., Balaci, T., Lupuleasa, D. (2012). Preparation and
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cyclodextrin and hydroxypropyl-α-cyclodextrin. Farmacia 63(4), pp. 548-555.
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D., Ioniță, A. C., Pantea A., Stoian, C. Nicolae, Rizzo, M., Mititelu, M. (2019). Insights into
stroke: a rewiew of pharmacokinetics and pharmacodynamic studies. Nutrients 11 (7), pp. 1479-1513.
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obesity effect of herbal active and endogenous lipids co-loaded lipid nanocarriers: Preparation, in vitro
and in vivo evaluation. Materials Science and Engineering. C 99, pp. 12-24.
11. Mititelu, M., Ioniţă, A.C., Moroşan, E. (2014). Research regarding integral processing of mussels from
Black Sea. Farmacia, Vol. 62(3), pp. 625-632.
12. Mititelu, M., Moroşan, E., Neacsu, S. M., Ioniţă, E. I. (2018). Research regarding the pollution degree
from romanian Black Sea coast. Farmacia, 66(6), pp. 1059-1063.
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13. Ioniţă, A.C., Mititelu, M., Moroşan, E. (2014). Analysis of heavy metals and organic pollutants from
some Danube river fishes. Farmacia 62(2), pp. 299-305.
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in soil on the composition of some wild edible mushrooms. Farmacia 67(3), pp. 398-404.
15. Mititelu, M., Moroşan, E., Iosif, M., Ioniţă, E. I. (2018). Analysis of quality of different types of honey
from various sources, Proceedings of The Romanian National Congress of Pharmacy – 17th Edition, “21st
Century Pharmacy – Between Intelligent Specialization and Social Responsibility”. Filodiritto Editore –
Proceedings, pp. 84-87.
16. Ioniță, A. C., Ghica, M., Moroșan, E., Nicolescu, F., Mititelu, M. (2019). In vitro effects of some
Farmacia 67(4), pp. 684-690.
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24-63, 70-84, 87-103, 114-116, 121-123, 129-130, 137-138, 141-143, 219-220, 245-248, 253-255.
331
Research on the Influence of Some Sweeteners on Body Weight

and Blood Glucose in Laboratory Mice
MITITELU Magdalena1, NICOLESCU Teodor Octavian2*,

UDEANU Denisa Ioana1, NICOLESCU Florica3
(ROMANIA)
(ROMANIA)
Email: nicolescu.teodor@gmail.com
Abstract
Sugar and sweet products occupy an important place in the nutrition of adults, but especially
of children. Most of the time, the daily ration is outdated and it reaches a consumption that is
not compatible with human metabolism. In the experimental part, we studied the change of the
weight of the white laboratory mice treated with different sweeteners (white sugar, stevioside,
xylitol, brown sugar) for 14 days. At the end of the experiment, blood samples were collected
to determine the blood glucose levels.
Keywords: white sugar, sativoside, brown sugar, xylitol, glycaemic index
Introduction
Excess sugar and the use of artificial sweeteners such as aspartame or sucrose contribute to
serious health problems such as diabetes, heart disease, tooth loss, obesity and various types of
cancer. Exacerbated consumption of sweet products is a major issue in the global health system
at the present time [1, 2, 3].
The metabolizable ones are those that assimilate quickly and easily, providing the body with
the necessary energy. In turn, they are of two types: fast assimilation (sucrose, fructose) and
slow assimilation (starch). Over the last decade, researchers’ interest in studying the effects of
sugar and its products has grown steadily. This is due to the increasing number of cases of
obesity, type 2 diabetes, high blood pressure or the incidence of dental caries in people who
consume excessive amounts of sugar and sugary products [4, 5, 6].
Sugar is a food obtained from sugar beet or cane, with a high content of sucrose, which gives
it a pronounced sweet taste. It is used in food to sweeten foods and drinks, but also as a
preservative. Sugar in the form of refined sugar (white) is the main component in confectionery
products, accounting for 30-40% of the total ingredients. Sugar marketed as brown sugar can
be raw, unrefined, or refined white sugar, which has been coloured after refining with molasses
or caramel. The abuse of sweet food products leads to the occurrence of diabetes, but also
obesity among frequent consumers. These two diseases are direct constituents of some
associated pathologies, which, at present, are causes of morbidity. It is also worth mentioning
that excessive consumption of sugar rich products leads to complications such as high blood
pressure, dental caries and dental complications, but also abnormal hormonal development [7,
8].
Xylitol is a natural carbohydrate that can be extracted from any plant with woody material.
332
Commercially, it is obtained from renewable sources of environment, for example maize

chips, but also from less sustainable sources, such as hardwood. The low glycaemic index
makes it ideal for consumption, even by diabetics.
Stevia is a plant native to South America and has natural sweet compounds. The steviosides
are processed from the Stevia rebaudiana plant. When refined in a white powder extract, they
are 200-300 times sweeter than sugar.
The steviosides is a natural sweetener, but it is also used because of its potential to prevent
or combat some chronic diseases. Stevia extract increases insulin sensitivity in diabetes, lowers
blood pressure, may have diuretic action and has anti-inflammatory effect.
Choosing a natural sweetener can be a healthy alternative, and nutritionists encourage their
consumption as they do not contain harmful additives or dyes to the body. Obviously, it is very
important that natural products come from low pollution environments, in order to avoid
contamination with different pollutants [9, 10, 11, 12, 13]. Natural sweeteners can be an
alternative solution to make up for the lack of sugar in your diet. In sweets, juices or lemonades,
they give flavour and, in some cases, bring real health benefits [14, 15, 16, 17].
Methodology
The change in the weight of the white laboratory mice treated with different sweeteners
(white sugar, stevioside, xylitol, brown sugar) for 14 days was studied. At the end of the
experiment, blood samples were collected to determine the blood glucose levels. The animals,
white laboratory mice, males, weighing 18-25 g, were brought from the farm and allowed to
get used to the new habitat for 2 days. The food was administered in the morning at 8:00 and
in the evening at 17:00. Water received ad libitum from bottles. Five batches of 10 mice each
were made. The first batch (the control batch) was fed normally, and the other batches received
different types of sugar, in addition to normal feed, as follows: Batch 2: 0.5 mL/10 g body, 20%
white sugar solution; Batch 3: 0.5 mL/10 g body, stevioside solution (keep in mind that 5 drops
stevioside solution correspond to 25 g white sugar); Batch 4: 0.5 mL/10 g body, xylitol solution
30%; Batch 5: 0.5 mL/10 g body, 20% brown sugar solution. The administration was done once
a day, in the morning, oral. Body weight was tracked every 2 days, by weighing the animals in
the morning, before the meal.
The effect variations were calculated at each moment, compared to the initial moment,
according to the formula:
E% = (Ex-E0)/E0 x 100
where:
E0 = the initial effect of the witness; Ex = the measured effect for each component studied.
At the end of the experiment, blood was collected to determine blood glucose levels. Blood
glucose was determined in the laboratory using an Omron A2 Easy glucometer. A single drop
of blood was used to determine the value of fasting blood sugar (à jeun).
The data were statistical processing by student t test.
x ± SD = average ± standard deviation
Results
Experimental results regarding the evolution of body weight are shown in Tables 1-5 and
Fig. 1-5. Experimentally, we notice an obvious increase in body weight compared to the initial
moment in mice treated with different types of sugar compared to the control group, depending
on the motor activity and the individual metabolism. The highest increase occurred in the case
333
of administration of white sugar and brown sugar (3-5 g mouse), and in descending order we
mention xylitol (2-4 g/mouse) and stevioside (2-3 g/mouse).
Table 1. Change in body weight (B. W.) in the animals in the control group
B. W. 1
B. W. 2 (g) B. W. 3 (g) B. W. 4 (g) B. W. 5 (g) B. W. 6 (g) B. W. 7 (g)
(g)
The 3rd The 5th The 7th The 9th The The 13th
The 1st
Day Day Day Day 11thDay day
Day
x ± SD 21±1.5 20,3±1.12 21,1±0.19 21,4±1.61 21,7±1.76 22,1±2.11 22,6±1.1
Effect % - -3,3333 0,4761 1,9047 3,3333 5,2380 7,6190
E%
8 7,61
5,23
3,33
6
4 1,9
0.4761
2
-3.33
0
-2
-4
2 DAYS 4 DAYS 6 DAYS 8 DAYS 10 DAYS 12 DAYS
Fig. 1. Change in body weight in the animals in the control group
Table 2. Change in body weight (B. W.) in animals treated with white sugar
B. W. 1
B. W. 2 (g) B. W. 3 (g) B. W. 4 (g) B. W. 5 (g) B. W. 6 (g) B. W.7 (g)
(g)
st The 3rd The 5th The 7th The 9th The The
The 1
Day Day day Day 11thDay 13thDay
Day
x ± SD 21,3±1.3 21,9±1.25 22,5±1.44 23,1±1.61 23,9±0.48 24,7±1.71 25,2±1.82
Effect % - 2,8169 5,6338 8,4507 12,2065 15,9624 18,3098
E%
20 18,3
15,96
15 12,2
8,45
10 5,63
2,81
5

Fig. 2. Change in body weight in animals treated with white sugar
Table 3. Change in body weight (B. W.) in stevioside treated animals

B. W. 2
st (g)
The 1 The 5th The 7Th The 9th The The
The 3rd
Day Day Day Day 11thDay 13thDay
Day
x ± SD 21,4±1.22 21,7±1.6 22,4±1.81 22,4±1.37 23,2±1.42 23,4±1.18 23,4±0.85
Effect % - 1,4018 4,6728 4,6728 8,4112 9,3457 9,3457
334
E%
10 8,41 9,34 9,34
8
6 4,67 4,67
4
1,4
2
Fig. 3. Change in body weight in stevioside treated animals
Table 4. Change in body weight (B. W.) in animals treated with xylitol
B. W. 2
(g)
The 1st rd The 5th The 7th The 9th The The
The 3
Day
x ± SD 21,5±1.14 22±1.3 22,9±1.28 23,5±1.56 23,9±1.73 24,4±1.86 24,7±1.28
Effect % - 2,3255 6,5116 9,3023 11,1627 13,4883 14,8837
E%
15 13,48 14,88
11,16
9,3
10 6,51
5 2,32

Fig. 4. Change in body weight in animals treated with xylitol
Table 5. Change in body weight (B. W.) in animals treated with brown sugar
B. W. 3
st rd (g)
The 1 The 3 The 7th The 9th The The
The 5th
Day
x ± SD 23,7±1.14 24,2±1.39 25±1.63 25,4±1.29 26,1±1.15 26,8±1.33 27,4±0.73
Effect % - 2,1097 5,4852 7,1729 10,1265 13,0801 15,6118
E%
20
13,08 15,61
15 10,12
10 7,17
5,48
5 2,1
0
Fig. 5. Change in body weight in animals treated with brown sugar
The results obtained for the average glucose levels in the batches of animals treated with
different sweeteners are presented in Table 6 and Fig. 6.
335
Table 6. The average glucose levels in the batches of animals treated

with different sweeteners
Glucose levels (mg/dL) ( x ± SD)

Control The type of sugar
group White sugar Stevioside Xylitol Brown sugar
89±1.12 127±0.42 93±1.35 98±0.22 115±1.67
E%
140 127 115
120 93 98
89
100
80
60
40
20
0
Control group White sugar Stevioside Xylitol Brown sugar
Fig. 6. Modification of the glycaemic index in animals with sweeteners
Maintenance of blood glucose within normal limits (70-110 mg/dL) is observed in normal
fed mice and those given stevioside and xylitol. Mice that received white sugar and brown sugar
have elevated values above the upper limit of serum glucose.
Research has shown an average glycaemic increase of: 38 units for white sugar; 4 units for
stevioside; 9 units for xylitol; 26 units for brown sugar.
Conclusions
In the experiment performed on laboratory mice it was found that the natural sweeteners
introduced into the diet for 14 days caused an increase in body mass, but with significant
differences between groups. The highest increase occurred in the case of administration of white
sugar and brown sugar (3-5 g/mouse), and in descending order we mention xylitol (2-4
g/mouse) and stevioside (2-3 g/mouse). Maintenance within the normal limits of glucose (70-
110 mg/dL) was observed in normal fed mice and those who were given stevioside and xylitol.
Mice that received white sugar and brown sugar showed increased values above the upper
limit of serum glucose.
REFERENCES
1. Goldstein, D., Mintz, S., Krondl, M., Mason, L. (2015). The Oxford Companion to Sugar and Sweets.
Oxford University Press
D., Ioniță, A. C., Pantea A., Stoian, C. Nicolae, Rizzo, M., Mititelu, M. (2019). Insights into chrono
nutrition: the innermost interplay amongst nutrition, metabolism and the circadian clock, in the context
of epigenetic reprogramming. Farmacia 67(4), pp. 557-571.
stroke: a rewiew of pharmacokinetics and pharmacodynamic studies. Nutrients 11(7), p. 1479.
4. Forshee, R.A., Storey, M. L., Allison, D.B., et al., (2007). A Critical Examination of the Evidence
Relating High Fructose Corn Syrup and Weight Gain. Crit Rev Food Sci Nutr.,47(6), pp. 561-582.
5. Phillips, K.M., Carlsen, M.H., Blomhoff, R. (2009). Total antioxidant content of alternatives to refined
sugar. J Am Diet Assoc., 109(1), pp. 64-71.
6. Ajibola, A., Chamunorwa, J.P., Erlwanger, K.H. (2012). Nutraceutical values of natural honey and its
contribution to human health and wealth. Nutr Metab., 9, p. 61.
7. Bogdanov, S., Jurendic, T., Sieber, R., Gallmann, P. (2008). Honey for Nutrition and Health: A Review.
J Am Coll Nutr., 27(6), pp. 677-689.
336
8. Mititelu, M., Ioniţă, A. C., Moroşan, E. (2014). Research regarding integral processing of mussels from
Black Sea. Farmacia, 62(3), pp. 625-632.
9. Mititelu, M., Moroşan, E., Neacsu, S. M., Ioniţă E. I. (2018). Research regarding the pollution degree
from romanian Black Sea coast. Farmacia 66(6), pp. 1059-1063.
10. Ioniţă, A. C., Mititelu, M., Moroşan, E. (2014). Analysis of heavy metals and organic pollutants from
some Danube river fishes. Farmacia 62(2), pp. 299-305.
Century Pharmacy – Between Intelligent Specialization and Social Responsibility”, Filodiritto Editore –
13. Ioniță, A. C., Ghica, M., Moroșan, E., Nicolescu, F., Mititelu, M. (2019) In vitro effects of some
Farmacia 67(4), pp. 684-690.
de Chimie 54 (8), pp. 676-680.
337
Study of Hypolipemiant Activity of Some Mussel Extracts
MITITELU Magdalena1, NICOLESCU Teodor Octavian2*, MOROȘAN Elena1,

POP Anca Lucia1, NICOLESCU Florica3
(ROMANIA)
(ROMANIA)
* Corresponding author: nicolescu.teodor@gmail.com
Abstract
The aim of this work is to study the hypolipemiant activity of some biological active extracts
from mussels on rats treated with TRITON WR 1339 for to induce the hyperlipidaemia. For the
experiment it used five groups of 10 rats. For 14 days one group get suspension of mussel meat
10%, another group get total lipid extract from mussel and a group get simvastatin. Were
determined following biochemical parameters on blood serum: cholesterol, HDL-cholesterol,
triglycerides and total blood lipids. Because of their complex composition (especially due to
polyunsaturated fatty acids) specifically marine lipids, the total lipid extract from mussel
present an important hypolipemiant activity as we can see from the experimental data obtained.
Keywords: Mussel extracts, hypolipemiant activity, polyunsaturated fatty acids, marine lipids, obesity
Introduction
A healthy diet includes a variety of foods that provide the body with all the necessary
nutrients. Most people do not consume enough nutrients because they do not include in the diet
an adequate amount from each food group [1, 2, 3].
Many vegetable oils are found in many dietary supplements on the pharmaceutical market
and are useful in many conditions. Thus, supplements with omega 3-6-9 from fish oil, from
hemp oil and flax are supplements indicated as adjuvants in the treatment of cardiovascular
disease, dyslipidaemia and inflammatory diseases [4, 5].
Long chain omega 3 fatty acids have the most favourable effects on the human body. Their
presence in the diet contributes to diminishing the negative consequences of stress, balancing
the optimal ratio between omega-type fatty acids, reducing “bad” cholesterol and fluidizing
blood. Through their action in the human body, these compounds play a special, protective role,
especially over the entire circulatory system.
The effects of long chain omega 3 fatty acids are also felt psychologically. It has been found
that people who regularly consume fish rich in such substances, are better prepared, more
tolerant, without violent starts and much less exposed to depression.
Clinical studies have shown that long-chain omega 3 fatty acids sensitize insulin receptors,
they decrease insulin resistance, preventing the manifestation of the consequences of this
disorder (diabetes, obesity, dyslipidaemia).
Eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) found in marine fish, are life-
sustaining substances. Thus, the inhabitants who live near the temperate or cold sea coasts (the
338
shores of the Japanese islands, the coasts of the Korean peninsula, the Scandinavian fjords,
etc.), form the longest human populations [6, 7, 8].
Marine lipids, especially EPA and DHA, have largely demonstrated their bioactivity in
human health [9].
The mussels (Mytilus galloprovincialis L.) are an important source of biological active
substances and calcium, therefore:
- their meat can be used as food, because of its remarkable alimentary values, with high
assimilation degree – 85% (the content in essential amino acids is almost 30%) [10];
- the lipids have a great biological value because are compose of poly-unsaturated fatty
acids and fat-soluble vitamins (A, D, E).
Mytilus galloprovincialis is one of the most common invertebrates in the Black Sea, whose
rich content in valuable nutrients has brought it to researchers’ attention. The particular nutritive
quality of mussel flesh is due to the proportion of vital components necessary for human
nutrition: protein, fat, carbohydrates, macro- and micro-nutrients, enzymes, vitamins etc.
Mussels biochemical composition is influenced by geographical area, depth, nourishment,
gender, age, reproductive status, climate factors and environmental pollution level [11, 12].
Regarding the marine environment, the quality of the marine products is strongly influenced
by the degree of water pollution [13, 14, 15]. This is why marine products should be harvested
from the areas with the least pollution, to avoid heavy metal or microplastic poisoning [16, 17,
18, 19].
The purpose of the present work is to study the hypolipemiant activity of some biological
active extracts from mussels on rats treated with TRITON WR 1339 for to induce the
hyperlipemia. Were determined following biochemical parameters on blood serum: cholesterol,
HDL-cholesterol, triglycerides, total serum lipids.
Methodology
For the experiment it used 5 groups of 20 white male rats Wistar line, with a weigh of
190±10g. For 7 days one group (the control group) get only the food, a group get also only the
food (the group will be treated with only the TRITON), a group get 10 mL/kgc suspension of
mussel meat 10% p.o., a group get 0.5 mg/kgc total lipid extract from mussel p.o. and a group
get 1.5 mg/kgc simvastatinum (reference substance with hypolipemiant activity) p.o. Before to
start the experiments, the animals were cheap without food for 12 hours. The five groups were
treated with the following substances:
- the control group – the rats were treated with10 mL/kgc NaCl solution 0.9% i.p.;
- the others were treated with 10 mg/kgc solution of TRITON WR 1339 10% i.p.
After two hours the animals were kild and on the blood serum were determined following
biochemical parameters by Cormay method [20, 21]: cholesterol, HDL-cholesterol,
triglycerides, total serum lipids.
The effect variations were calculated at each moment, compared to the initial moment,
according to the formula:
E% = (Ex-E0)/E0 x 100
where:
E0 = the initial effect of the witness
Ex = the measured effect for each component studied
The data were statistical processing by student t test and ANOVA [22].
339
Results
The experimental data are presented in Tables 1-4 and Fig. 1-4 below:
Table 1. Statistical significance of the results regarding the influence of the analysed extracts
on triglycerides, in rats
Triglycerides mg/dL
The analysed product Dose
(mL/Kgc, oral) p/control group Effect % related
x ±SD. to control group
Control group - 51.40±3.09 - -

Triton 10 90.60±4.41 0.002047 +76.26
Triton + mussel meat 10 58.62±2.08 0.003129 -35.29
Triton + total lipid 55.24±5.21 0.002415 -39.02
0.5
extract
Triton + Simvastatinum 1.5 48.3±1.79 IS -46.68
F (Anova) 62.16609
P (Anova) 0.001036
x ± SD= average ± standard deviation; IS = insignificant statistical
Fig. 1. The influence of analysed extracts on triglycerides, in rats
Table 2. Statistical significance of the results regarding the influence of analysed extracts
on total serum lipids, in rats
Total serum lipids mg/dL
(mL/Kgc, oral) x ±SD p/control group Effect %
related to
control group
Control group - 192.02±7.83 - -
Triton 10 278.54±11.22 0.002754 +45.05
0.5
extract
Triton + Simvastatinum 1.5 224.37±9.98 0.003324 -20.50
F (Anova) 121.1431
P (Anova) 0.003051
x ± SD= average ± standard deviation
340
Fig. 2. The influence of analysed extracts on total serum lipids, in rats
Table 3. Statistical significance of the results regarding the influence of analysed extracts on cholesterol, in rats
Cholesterol mg/dL
(mL/Kgc, oral) p/control group Effect % related
x ±SD. to control group
Triton 10 156, 00±2.34 0.002754 +63.33
0.5
extract
F (Anova) 128.947
P (Anova) 0.002178
x ± SD= average ± standard deviation
Fig. 3. The influence of analysed extracts on cholesterol, in rats
Table 4. Statistical significance of the results regarding the influence of analysed extracts
on HDL-cholesterol, in rats
HDL-c mg/dL
Product analysed Dose
(mL/Kgc, oral) x ± SD p/control group Effect % related
to control group
Triton 10 71.34±1.35 0.002461 +95.23
0.5
extract
F(Anova) 57.543
P(Anova) 0.001998
x ± SD= average ± standard deviation; HDL-c = HDL-cholesterol
341
Fig. 4. The influence of analysed extracts on HDL-cholesterol, in rats
From data presented in the tables above it can be observed that the total lipid extracts from
mussel meat has a good hypolipemiant effect. The mussel meat has presented an effect more
reduced (approximatively at half from total lipid extract effect).
Conclusions
Because of their complex composition (especially due to poly-unsaturated fatty acids)

specifically marine lipids, the total lipid extract from mussel present an important
hypolipemiant activity as we can see from the experimental data obtained. All the serum
parameters analysed (cholesterol, HDL-cholesterol, triglycerides, total serum lipids) on rats
after administration of TRITON (hypolipemiant agent) were reduced by the diet with total lipid
extract from mussel.
Clinical studies have shown that long-chain omega 3 fatty acids sensitize insulin receptors,
they decrease insulin resistance, preventing the manifestation of the consequences of this
disorder (diabetes, obesity, dyslipidaemia), so mussels are a good food for the diet of patients
with diabetes, obesity or dyslipidaemia.
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nutrition: the innermost interplay amongst nutrition, metabolism and the circadian clock, in the context
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343
Comparative Assessment of Continuous Hospitalization and Day

Hospitalization in Patients with Diabetes Mellitus
DAINA Lucia Georgeta1, VENTER Alina Cristiana1, POPA Amorin Remus1,

VESA Cosmin Mihai1, BUHAȘ Camelia Liana1, SABAU Monica1,
BONTA Marinela1, DAINA Cristian Marius1
1Faculty of Medicine and Pharmacy, University of Oradea (ROMANIA)
Emails: lucidaina@gmail.com, alinaventer@gmail.com, popa_amorin@yahoo.com, v_cosmin_15@yahoo.com,
cameliabuhas@yahoo.com, sabau.monica@yahoo.com, bontamarinela@yahoo.com, cristi_daina@yahoo.co.uk
Abstract
The rationalization of the health system aims at increasing the efficiency of the medical
services by reducing the costs at the level of the hospital units, increasing the quality of the
medical act and access to services. In this study were analysed the cases with the main diagnosis
of diabetes mellitus discharged in continuous hospitalization and in day hospitalization, in an
emergency clinical hospital. In the analysed period (6 months) the number of cases discharged
in day hospitalization is 10 times higher than the number of cases discharged in continuous
hospitalization. Women predominate in both continuous hospitalization and day
hospitalization, and by origin, in day hospitalization, we find more patients in the urban area.
The mean age was significantly lower in patients with day hospitalization compared to those
with continuous hospitalization. On types of diagnosis, in the continuous hospitalization we
have no cases of type 2 diabetes without complications, while type 1 diabetes and diabetes
mellitus with microvascular complications, is statistically significantly discharged in
continuous hospitalization (the difference being significant in type 1 diabetes). The average
length of hospitalization was 4.13±2.07 days. Given the group and the diagnostic code, as well
as the average length of hospitalization, 22 cases discharged in continuous hospitalization could
have been treated in day hospitalization. For the 22 cases treated in continuous hospitalization,
the hospital lost 1650 RON (EUR 345.20) and if these cases were treated in day hospitalization,
it would have earned 3482 RON (EUR 728.47).
Prevention of continuing hospitalizations considered avoidable can be achieved through
appropriate monitoring in day hospitalization.
Keywords: diabetes, hospital, continuous hospitalization, day hospitalization, average duration of hospitalization, costs
Introduction
The role of the hospital, as a public institution, is to provide the highest quality public
products and services, in order to satisfy the general and individual needs of society, under
conditions of economic efficiency [1, 2]. The financing of the hospital is realized through
various payment mechanisms, the most used being the contracts on types of services. In this
type of contracts are specified the services provided by the hospital and the payment method
for each type of services [3]. The contracting of the hospital services is found in a common
normative act the National Health Insurance House (NHIH) and the Ministry of Health (MH)
called the Framework Contract, which regulates the conditions of the provision of medical
assistance, medicines and medical devices within the social insurance system. [4]. For the
hospital medical care are available: the minimum package of medical services (for the patient
344
who cannot prove the insured quality) and the package of basic medical services (for the insured
patient). Depending on the length of hospitalization, hospital medical care is provided under
continuous hospitalization and day hospitalization [5]. Day hospitalization is an alternative to
hospitalization for patients who cannot be managed in the outpatient or specialized offices,
where evaluation and treatment services are provided. For the hospital, the main advantages of
day hospitalization are: lower long-term care costs, relieving hospital sections of acute cases in
remission or during the stabilization period. For the patient, the advantages are: the possibility
of intensive and multidisciplinary care, flexible approach of the patient with alternatives of care
programs and their planning. The multidisciplinary approach of the patient and the respect of
the diagnostic and therapeutic protocols result in lower costs [6, 7].
The patient diagnosed with diabetes requires lifelong surveillance and monitoring, benefiting
from multiple and complex medical services, with significantly increased costs [8, 9]. The types
of medical services accessed by the patient with diabetes are multiple, from the family doctor,
to the hospital (specialized offices, day hospitalization, continuous hospitalization). Emergency
hospitalization, for this type of patient, most often demands a decompensation of the basic
disease, but a scheduled hospitalization must be very well established by the specialist doctor.
The purpose of the study is to analyse the cases and patients discharged from the hospital,
with the main diagnosis of diabetes, compared in continuous hospitalization and in day
hospitalization, in order to identify potential problems and to implement specific measures.
Methods
A retrospective study was conducted in the period 01.01.2018-30.06.2018, analysing the

patients discharged from the Oradea County Emergency Clinical Hospital, with the main
diagnosis of diabetes. There were compared the cases discharged from continuous
hospitalization and from day hospitalization, according to the personal characteristics of the
patients (sex, age of origin, age), diagnosis and diagnosis group, average length of
hospitalization, costs.
Inclusion criteria:
- discharged patients with the main diagnosis of diabetes
- age ≥18 years
- diagnostic code: E10.65, E10.71, E11.65, E11.71. E11.9
o E10.65 – Type 1 diabetes mellitus with poor control
o E10.71 – Type 1 diabetes mellitus with microvascular complications
o E11.65 – Type 2 diabetes mellitus with poor control
o E11.71 – Type 2 diabetes mellitus with microvascular complications
o E11.9 – Type 2 diabetes mellitus without complications
Exclusion criteria:
- age <18 years
- deaths
Results
During the study period, 207 cases were discharged from continuous hospitalization,
representing 198 patients, 9 patients having 2 hospitalizations (4.55%). 2415-day
hospitalizations were performed, 2041 patients were discharged in day hospitalization, 321
patients had 2-5-day hospitalizations (15.73%).
In both groups, women predominated (51.25% vs. 56.06%, p=0.196) (Table 1).
345
Patients from the urban area (55.17%) predominated in the day hospitalization, and those
from the rural areas (53.03%) (p=0.027) (Table 1) continued in the continuous hospitalization,
demonstrating the easier accessibility of the patients from the urban area to health services.
Table 1. Distribution of discharged patients by sex and living environment

Discharged patients Day hospitalisation Continuous hospitalisation
No. % No. %
Sex Women 1.046 51.25 111 56.06
Men 995 48.75 87 43.94
Living Urban 1.126 55.17 93 46.97
environment Rural 915 44.83 105 53.03
Total 2.041 100.00 198 100.00
The age limits for discharged patients in day hospitalization were between 18-81 years, and
in continuous hospitalization 20-87 years. The mean age was significantly lower in patients
with day hospitalization compared to those with continuous hospitalization (59.81 years vs.
63.39 years, p<0.001) (Table 2).
Table 2. Distribution of discharged patients by age

Limits Mean age
Day hospitalisation 18-81 years 59.81±10.24
Continuous hospitalisation 20-87 years 63.39±9.75
The distribution of cases discharged according to diagnosis is shown in table 3. In the day
hospitalization, the diagnosis of type 2 diabetes mellitus with poor control (32.96%) prevails,
and in the continuous hospitalization – type 1 diabetes mellitus with microvascular
complications (50.24%). We note that in the continuous hospitalization we have no cases of
type 2 diabetes without complications. In day hospitalization type 1 diabetes accounted for
46.63%, while in continuous hospitalization this percentage was 70.05% (p<0.001). Also, the
cases with diabetes mellitus with microvascular complications accounted for 46.42% in day
hospitalization and 67.15% in continuous hospitalization (p<0.001), the difference being
significant in type 1 diabetes mellitus (29.86% vs 50, 24%, p<0.001) and insignificant in type
2 diabetes (16.56% vs. 16.91% .p=0.87) (Table 3).
Table 3. Distribution of cases discharged cases according to diagnosis

Diagnostic Diagnostic Day hospitalisation Continuous
code hospitalisation
No. % No. %
DM type 1 with poor control E10.65 405 16.77 41 19.81
DM type 1 with microvascular E10.71
complications 721 29.86 104 50.24
DM type 2 with poor control E11.65 796 32.96 27 13.04
DM type 2 with microvascular E11.71
complications 400 16.56 35 16.91
DM type 2 without complications E11.9 93 3.85 0 0.00
Total hospitalisations 2415 100.00 207 100.00
For a more detailed analysis, for the continuous hospitalization, the diagnostic groups related
to the diagnostic codes E10.65, E10.71, E11.65, E11.71, E11.9 were taken into consideration.
Thus, we find as diagnostic groups: Diabetes mellitus with catastrophic/severe complications
– K3011, respectively Diabetes mellitus without catastrophic/severe complications – K3012.
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Of the 207 discharged patients, the diagnostic group K3011 (Diabetes with
catastrophic/severe complications) accounted for 85.02% (176 cases), and K3012 ((Diabetes
without catastrophic/severe complications) only 14.98% (31 cases) (Table 4).
Table 4. Distribution of cases of continuous hospitalization according to diagnosis, code and diagnostic group,
average length of hospitalization
Diagnostic Diagnostic Diagnostic No. % LOS
code group
K3011 28 13.53 3.68±1.81
E10.65
DM* type 1 with poor control K3012 13 6.28 3.15±2.02
DM type 1 with microvascular K3011 95 45.89 4.65±2.23
E10.71
complications K3012 9 4.35 4.78±2.12
K3011 21 10.14 3.90±1.77
E11.65
DM type 2 with poor control K3012 6 2.90 2.33±1.80
DM type 2 with microvascular K3011 32 15.46 3.75±2.11
E11.71
complications K3012 3 1.45 3.00±1.66
*DM= Diabetes mellitus
Most of the hospitalized cases are represented by the diagnostic group K3011, with the
diagnosis Diabetes type 1 with microvascular complications (45.89%) and Diabetes type 2 with
microvascular complications (15.46%).
Of the 145 discharges with type 1 diabetes, group K3011 represented 84.83%, and of the 62
discharges with type 2 diabetes, group K3011 represented 85.48% (p=0.932). In the diagnostic
group K3012, the cases of type 1 diabetes were 15.17%, and type 2 diabetes, 14.52%.
Regarding the average length of hospitalization, this was 4.13±2.07 days, significantly lower
in the cases with group K3011 compared with group K3012 (4.24±2.12 days vs. 3.45±1,78
days, p=0.052) (Table 5).
Table 5. Distribution of continuous hospitalization cases according to the diagnosis group, diagnosis
and average length of hospitalization
Diagnostic No. of DM type 1 % DM type 2 % LOS
group cases
K3011 145 123 84,83 22 85.48 4.24±2.12
K3012 62 53 15,17 9 14.52 3.45±1.78
The longest mean duration of hospitalization is found in the diagnosis of type 1 diabetes
mellitus with microvascular complications, in both diagnostic groups: K3011 – 4.65±2.23 days,
K3012 – 4.78±2.12 days. The rest of the diagnoses in the diagnostic group K3012 have an
average length of hospitalization from 2.33±1.80 days to 3.15±2.02.
Eliminating discharges with the longest duration of hospitalization: the diagnostic group
K3011, and the cases from K3012 – E10.71, resulted in 22 discharges from the continuous
hospitalization that could have been made in the day hospitalization (10.62%).
Given that the costs of continuous hospitalization are higher than in day hospitalization, the
costs of the two types of hospitalization were analysed, to determine what the costs would have
been in the 22 cases, if they had been treated in day hospitalization.
Based on the contract with the National Health Insurance House (NHIH), the hospital is
payed for the medical services reported and validated by the National School of Public Health
and Health Management. These tariffs are set out in the Framework Contract and the
Implementing Rules [3]. In the continuous hospitalization the fee settled (tariff by weighted
case) by the NHIH for each discharged case, is a reimbursement value of a discharged case
adjusted with its relative complexity in relation to the other cases [10, 11]. For the cases
discharged from continuous hospitalization, with the diagnosis of diabetes mellitus, the tariff
settled by NHIN to the hospital, represented in 2018 917 RON (191.84 EUR). For the day
347
hospitalization, the fee settled by the NHIH to the hospital differs depending on the diagnosis,
the patient model and the list of medical services. We find these rates in Table 6.
Table 6. The fees settled by the National Health Insurance House (NHIH), and the average costs supported by
the hospital, by types of diagnosis
Diagnostic Currency Day hospitalisation Continuous hospitalisation
NHIH Average Difference NHIH Average cost Difference
fee cost fee
DM type 1 with RON 274 152±32 122 917 1.257±142 -340
poor control EUR 57.32 31.80±6.7 25.52 191.84 262.98±29.71 -71.13
DM type 1 with RON 340 153±35 187 917 1.945±161 -1.028
microvascular
complications EUR 71.13 32.00±7.32 39.12 191.84 406.91±33.68 -215.07
DM type 2 with RON 310 152±33 158 917 1.422±137 -505
poor control EUR 64.85 31.80±6.9 33.06 191.84 297.50±28.66 -105.65
DM type 2 with RON 385 155±30 230 917 1.364±128 -447
microvascular
complications EUR 80.55 32.43±6.28 48.12 191.84 285.36±26.78 -93.52
DM type 2 RON 275 149±29 126 - - -
without EUR 31.17±6.07 -
complications 57.53 26.36 - -
Total RON 153±33 1.499±145 -582
EUR 32.00±6.9 313.6±30.34 -121.76
*1 EUR=4,7799 RON (27 December 2019)
The actual cost supported by the hospital for solving the case differs from the fee settled by
the NHIH.
In calculating the costs of continuous hospitalization, we only introduced continuous
hospitalization with the diagnostic group K3012 (diabetes mellitus without catastrophic/severe
complications).
We note the significantly higher costs in continuous hospitalization compared to day
hospitalisation (p<0.001). If the costs in the day hospitalization are below the rate set by the
NHIH, in the continuous hospitalization the costs are much higher, the payments being the same
regardless of the number of days of hospitalization.
For the 22 cases treated in continuous hospitalization, the hospital lost RON 1650 (EUR
345.20), and if these cases were treated in day hospitalization, it would have earned RON 3482
(EUR 728.47).
When the diagnosis cannot be established and the treatment cannot be performed and/or
monitored in day hospitalization (which lasts up to 12 hours/visit – day), continuous
hospitalization is indicated. For doing a continuous hospitalization will be taken into account
the severity of the signs and symptoms presented by the patient with diabetes, as well as the
medical predictability of a negative, unwanted evolution of the patient.
Conclusion
During the analysed period (6 months) the number of cases discharged in day hospitalization
is 10 times greater than the number of cases discharged in continuous hospitalization (207 cases
in continuous hospitalization and 2415 cases in day hospitalization).
In the continuous hospitalization we have no cases of type 2 diabetes without complications.
In day hospitalization type 1 diabetes accounted for 46.63%, while in continuous
hospitalization this percentage was 70.05% (p<0.001). Also, the cases with diabetes mellitus
with microvascular complications accounted for 46.42% in day hospitalization and 67.15% in
continuous hospitalization (p<0.001), the difference being significant in type 1 diabetes mellitus
348
(29.86% vs 50, 24%, p<0.001) and insignificant in type 2 diabetes (16.56% vs. 16.91%,
p=0.87). Most of the hospitalized cases are represented by the diagnostic group K3011, with
the diagnosis diabetes type 1 with microvascular complications (45.89%).
The mean duration of hospitalization was 4.13±2.07 days, slightly higher in the cases with
group K3011 compared with group K3012 (4.24±2.12 days vs. 3.45±1.78 days, p=0.052).
From the analysis, 22 cases discharged in continuous hospitalization could have been treated
in day hospitalization. For the 22 cases treated in continuous hospitalization, the hospital lost
RON 1650 (EUR 345.20), and if these cases were treated in day hospitalization, it would have
earned RON 3482 (EUR 728.47). Prevention of continuous hospitalizations considered
avoidable can be achieved by early diagnosis, approach, treatment and monitoring in day
hospitalization.
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rationalizing hospitals, Published in the Official Monitor no. 0223 of March 31, 2011.
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(2019), Improving performance of a pharmacy in a Romanian hospital through implementation of an
internal management control system, Science of the Total Environment, 675: pp. 51-61.
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Blood Pressure in the Young Adults from North-western Romania, Revista de chimie, vol. 70.
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D.C., Buștea C., Radavoi D.G. (2019) Prediction Models of Albumin Renal Excretion in Type 2 Diabetes
Mellitus Patients. Revista de chimie, vol 70 (11): pp. 3802-3807.
349
Impact of Diabetes Mellitus on Hospital Indicators
DAINA Cristian Marius1, POPA Amorin Remus1, VESA Cosmin Mihai1,

TIMAR Calin1, COTRĂU Petru1, DAINA Lucia Georgeta1
1Faculty of Medicine and Pharmacy University of Oradea (ROMANIA)
Emails: cristi_daina@yahoo.co.uk, popa_amorin@yahoo.com, v_cosmin_15@yahoo.com, calin_bh@yahoo.com,
petrucotrau@yahoo.com, lucidaina@gmail.com
Abstract
An important activity of the hospital management is the evaluation of the performance

indicators and the elaboration of measures regarding their improvement. Diabetes mellitus, by
the large number of people affected is a public health problem, and by the high costs that it
generates, increasing from year to year, it requires a correct evaluation of the hospital indicators.
The study evaluates the impact of diabetes on the evolution of patients with the same
pathology and the impact on the costs of hospitalization in an emergency clinical hospital.
Patients with a primary diagnosis of diabetes and those discharged from the Diabetes
Compartment were excluded from the study. Of the total number of discharged patients during
one year (2018), patients with secondary diagnosis of diabetes accounted for 17.53%.
Significantly higher values were recorded in patients having with diabetes mellitus as
comorbidity, at the following indicators: length of hospital stay (LOS)/hospitalization,
LOS/ICU section, LOS/total admissions, drug costs, total costs (930.39 vs. 786.64 EUR). A
relevant evaluation of the performance indicators of the hospital leads to a more efficient
management of the hospital resources and the implementation of measures that will increase
the efficiency and effectiveness of the medical act.
Keywords: diabetes mellitus, hospital, average length of hospitalisation, costs
Introduction
The hospital as a public institution carries out activities that correspond to the needs of the
territorially rounded population, satisfying the needs of the citizens by offering necessary
medical services that they cannot obtain through the market mechanism. These activities do not
produce revenues or produce minimal incomes insufficient to cover the needs of the hospital
[1]. The hospital management system (principles, rules, methods and decision-making
procedures) leads to the achievement of the objectives given the resource economy, as well as
the achievement of the performance [2]. For a unitary evaluation and correct comparison
through the Order of the Health Minister no. 1490/2008 [3] was approved the methodology of
calculation of the performance indicators of the hospital management. The most common
indicators used are: case complexity index (CCI), average length of hospitalization stay (LOS),
degree of use of beds, mortality, cost of hospitalization/day, bed and patient.
The large number of people diagnosed with diabetes determines the state of this condition
as a public health problem, in 2017, 424 million people had diabetes, and estimates show that
in 2045 the number of people with this disease will reach 628 million of cases [4, 5]. The
American Diabetes Association estimated a 26% increase in costs from 2012 to 2017, when
total costs of diabetes accounted for $327 billion [6, 7]. A study published in 2019 comprising
584 abstracts and 52 publications, on the costs of diabetes, estimates annual hospitalization
costs at about $ 290, ranging from about $10 to over $1000 [8]. Cost reduction but also
350
therapeutic success can be achieved through the patient’s multidisciplinary approach,

compliance and use of diagnostic and therapeutic guidelines [9-11].
The aim of the study is to analyse the hospital indicators in patients discharged from the
hospital by determining the impact of diabetes on the evolution of patients with the same
pathology, as well as the impact of this condition on the costs of hospitalization.
Methods
A retrospective study was carried out, based on the collection and statistical processing of
the data provided by the statistical services and the calculation office of the Oradea County
Clinical Hospital. In the study were included the discharged patients during the period
01.01.2018-31.12.2018.
Inclusion criteria:
- patients discharged with secondary diagnosis of diabetes
- patients with the same demographic characteristics and the same pathology (same main
diagnosis + same surgery)
- age ≥18 years
Exclusion criteria:
- patients discharged from the Diabetes Compartment or with a primary diagnosis of
diabetes
- patients <18 years of age
- patients without personal numeric code (CNP)
- discharge <24 hours (death, transfer, discharged on request)
- neoplasms
- accidents, injuries
- pregnant women/postpartum women
Discharged patients were divided into two groups:
- the first group was constituted by patients who, on discharge, had as secondary
diagnosis diabetes mellitus
- the second group was made up of patients without a diagnosis of diabetes.
During the study period, 22514 cases over 18 years with CNP, representing 20567 patients,
were discharged. Of the total discharged patients, 17.53% represented patients with secondary
diagnosis of diabetes (3605 patients).
The number of eligible patients was 11681, out of which 3605 patients with a secondary
diagnosis of diabetes (4557 discharges) and 8076 patients without a diagnosis of diabetes (9259
discharges).
The difference between the number of discharges and the number of patients is due to the
fact that we find patients with several discharges during the analysed period (2-8 discharges).
Results
The number of discharged women predominates in both groups (53.07% vs 53.47%,

p=0.699), which shows the greater addressability in women compared to men.
In the group with diabetes most patients came from the urban area (51.26% vs 51.98%,
p=0.472) (Table 1).
The age limits were between 18 and 92 years. The mean age was relatively equal in patients
with diabetes compared to those without diabetes (65.36 vs. 65.60, p=0.250) (Table 2).
In patients with secondary diagnosis of diabetes, 38.25% of the discharges were on surgical
sections, and in the patients without diabetes the percentage was 38.60% (p=0.691).
351
The hospitalization of patients with diabetes on the general surgery, ophthalmology,

dermatology, internal medicine predominates significantly statistically. Significantly lower
statistic we find the hospitalization of patients in the sections: obstetrics-gynaecology,
orthopaedics and cardiology (Table 3).
Table 1. Distribution of patients according to sex and environment of origin

Discharged patients Diabetes mellitus Without Diabetes mellitus
No. % No. %
Sex 1.913 53,07 1.913 53,07 56.06
1.692 46,93 1.692 46,93 43.94
Living 1.848 51,26 1.848 51,26 46.97
environment 1.757 48,74 1.757 48,74 53.03
Total 3.605 100.00 3.605 100.00
Table 2. Distribution of patients according to age

Age limits Average age
Patients with Diabetes mellitus 19-89 years 65.36±12.70
Patients without Diabetes mellitus 18-92 years 65.60±9.22
Table 3. Distribution of discharges by ward

With Diabetes Without Diabetes
Speciality Type of ward mellitus mellitus p
No. % No. %
General surgery 924 20.28 1.515 16.36 <0.001
Obstetrics and gynaecology 120 2.63 643 6.94 <0.001
Surgical Ophthalmology 166 3.64 227 2.45 <0.001
speciality Orthopaedics 272 5.97 723 7.81 <0.001
Urology 261 5.73 466 5.03 0.083
Total 1.743 38.25 3.574 38.60 0.691
Cardiology 1.152 25.28 2.629 28.39 <0.001
Dermatology 208 4.56 311 3.36 <0.001
Medical internal medicine 1.039 22.80 1.840 19.87 <0.001
speciality Nephrology 61 1.34 117 1.26 0.695
Neurology 354 7.77 788 8.51 0.138
Total 2.814 61.75 5.685 61.40
Total discharges 4.557 100.00 9.259 100.00
Multiple admissions
Of the patients with diabetes, almost 670 patients had between 2-8 hospitalizations in a year
(18.57%), and of the patients without diabetes, 914 patients had between 2-7 hospitalizations
(15.64%, p<0.001) (Table 4).
Table 4. Average length of hospitalization (LOS)

Limits LOS
Per discharge With Diabetes mellitus 0-89 days 6.07±4.46
Without Diabetes mellitus 0-84 days 5.46±4.01
Per total discharges With Diabetes mellitus 0-139 days 7.67±4.10
Without Diabetes mellitus 1-101 days 6.26±3.88
The mean LOS per hospitalization was significantly higher in patients with diabetes than
those without diabetes (6.07 vs. 4.46, p<0.001). Also, total LOS was significantly higher in
patients with diabetes than those without diabetes (7.67 vs. 6.26, p<0.001).
352
Surgery
Of the patients with diabetes, 1282 underwent surgery, and of those without diabetes, 2709
patients (35.56% vs 33.54%, p=0.034).
Intensive Care Days

The transfer to the sections of Anaesthesia and Intensive Care Unit (ICU) was required in
293 patients with diabetes and 655 patients without diabetes (8.13% vs. 8.11%, p=0.968).
The mean duration of hospitalization in the ICU ward was 7.31±4.46 days in patients with
diabetes and 4.83±3.12 days in patients without diabetes (p<0.001).
Death rate
Intra-hospital mortality was 6.41% among patients with diabetes mellitus and 5.78% among
patients without diabetes (p=0.185). The mean age of death was significantly lower in patients
with diabetes compared to patients without diabetes (71.58±8.04 years vs. 73.74±10.75 years,
p=0.007).
Costs (Table 5)
Table 5. Drug costs and total costs (expressed in RON and EUR)
With Diabetes Mellitus Without Diabetes mellitus
RON EUR RON EUR
Drugs 437.48±122.12 91.78±25.62 316.33±110.34 66.37±23.15
Without surgical
457.86±141.52 96.06±29.69 375.10±127.12 78.70±26.67
intervention
With surgical
385.42±112.41 80.87±23.59 225.04±132.22 47.22±27.74
intervention
Total costs 4434.41±1384.25 930.39±290.43 3749.23±1201.13 786.64±252.01
Without surgical
4443.76±1422.11 932.36±298.38 3954.25±1372.20 829.65±287.90
intervention
With surgical
4410.53±1379.22 925.38±289.38 3430.83±1194.33 719.83±250.59
intervention
*1 EUR = 4.77 RON (26.12.2019)
Regardless of the type of pathology that caused the hospitalization (medical or surgical), the
cost of the drugs was significantly higher in the patients with diabetes compared to the patients
without diabetes (p<0.001).
Given that LOS was higher in patients with diabetes, a significantly higher overall cost of
hospitalization was obtained in patients with diabetes compared to patients without diabetes
(p<0.001).
Secondary diagnosis of diabetes does not bring a significant additional amount to the solved
case. Through the Diagnosis Related Groups (DRG) system, patients can be classified
simultaneously by pathology and by the cost of care, which ensures the possibility of
associating the types of patients with the hospital expenses that are necessary [12]. Given the
high costs involved in treating a patient with a secondary diagnosis of diabetes, it is necessary
to re-evaluate the DRG coding.
Conclusion
Of the total number of discharged patients during one year (2018), patients with secondary
diagnosis of diabetes accounted for 17.53%.
Although the condition that caused the hospitalization did not represent diabetes, the
comparative evaluation (patients with diabetes vs. patients without diabetes) of some hospital
353
indicators, reflects significant differences for patients with diabetes: LOS/hospitalization,

LOS/ICU section, LOS/total admissions (p<0.001), multiple admissions per year, 2-8
admissions (18.57% vs. 15.64%), drug costs (91.78±25.62 vs. 66.37±23.15 EUR), total costs
(930.39±290.43 vs. 786.64±252.01 EUR). The mean age of death was significantly lower in
patients with diabetes compared to patients without diabetes (71.58+8.04 years vs. 73.74+10.75
years, p=0.007).
For the same pathology, a patient with the diagnosis of diabetes involves significantly higher
costs, costs that are not fully found in DRG funding.
In order to improve the indicators and reduce the costs, a correct monitoring and supervision
of the patients with diabetes mellitus is required. The evaluation of the hospital indicators is
important for the hospital management in order to allocate the necessary resources for the
treatment of these patients.
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1. Mullins, L. J., (2005) Management and Organisational Behaviour, Seventh Edition, Prentice Hall, Essex,
UK
2. Daina L.G., Sabău M., Daina C.M., Neamțu C., Buhaș C. L., Bungau C., Aleya L., Bungau S., Tit D. M.
(2019) Improving performance of a pharmacy in a Romanian hospital through implementation of an
internal management control system, Science of the Total Environment, 675: pp. 51-61.
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of calculation of the performance indicators of the hospital management. Text published in the Official
Gazette of Romania no. 628 of August 29, 2008.
4. Daina L., Popa A., Daina C., Vesa C., Bonta M., Venter A., Zaha C., (2018) Morbidity and mortality
through diabetes mellitus – Romania in a European context, Annals of the University of Oradea, Fascicle:
Ecotoxicology, Animal Husbandry and Food Science and Technology, Vol. XVII/A, pp. 281-291.
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6. https://www.diabetes.org/resources/statistics/cost-diabetes, accessed la 25 December 2019.
7. American Diabetes Association, (2018) Economic Costs of Diabetes in the U.S. in 2017, Diabetes Care,
May; 41(5): 917-928, https://doi.org/10.2337/dci18-0007.
8. Moucheraud C, Lenz C, Latkovic M, Wirtz VW. (2019) The costs of diabetes treatment in low-and
middle-income countries: a systematic review. BMJ Glob Health, 4(1): e001258 10.1136/bmjgh-2018-
001258.
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emergency hospital. Farmacia, 65(3), pp. 361-367.
10. Popa AR, Vesa CM, Uivarosan D. et al., (2019). Cross Sectional Study Regarding the Association
Between Sweetened Beverages Intake, Fast-food Products, Body Mass Index, Fasting Blood Glucose and
Blood Pressure in the Young Adults from North-western Romania, Revista de chimie, vol. 70.
11. Popa A.R, Judea-Pusta C.T., Vesa C.M., Bungău S., Buhaș C.L., Sava C., Dimulescu (Nica) I.A., Zaha
D.C., Buștea C., Radavoi D.G. (2019) Prediction Models of Albumin Renal Excretion in Type 2 Diabetes
Mellitus Patients. Revista de chimie, vol 70 (11): pp. 3802-3807.
12. https://www.drg.ro, accessed la 27 December 2019.
354
Major Differences Regarding Lipid Profile, Apolipoprotein-B and

Insulin Resistance between Type II Diabetes Patients with Altered
and Normal Glomerular Filtration Rate
GHERDAN Violeta Valentina1,2, POPA Amorin Remus1,2, JURCA Alexandru1,

POPA Loredana2, POPOVICIU Mihaela1,2, VESA Cosmin Mihai1,2,
FERICIAN Anca2, DAINA Cristian Marius1,2, ZAHA Dana Carmen1,2
1Faculty of Medicine and Pharmacy, University of Oradea (ROMANIA)
2Clinical County Emergency Hospital, Oradea (ROMANIA)
Email: v_cosmin_15@yahoo.com
Abstract
Knowing the factors that can influence glomerular filtration rate is of vital importance in
type 2 diabetes mellitus because of the possibility to influence those specific risk factors and
delay the progression towards chronic kidney disease. In our study we included 224 patients
with type 2 diabetes mellitus divided in two groups, one group included patients with mildly
reduced glomerular function (GFR between 60 and 90 mil/min/1.73m2) and one group included
patients with normal glomerular filtration rate (GFR>90mil/min/1.73m2). Patients with normal
glomerular function had significantly better cardio metabolic parameters including lower total
cholesterol, LDL-cholesterol, triglycerides, HbA1c, systolic and diastolic blood pressure and
higher HDL-cholesterol. In univariate analysis HbA1c, systolic blood pressure, LDL-
cholesterol, total cholesterol and triglycerides correlated negatively with good statistical
significance (p<0.05) with glomerular filtration rate while diastolic blood pressure and HDL-
cholesterol failed to obtain statistical significance (p>0.05). All these influencers are traditional
cardiovascular risk factors. However, when testing the influence of apolipoprotein B and insulin
resistance index (HOMA-IR) we observed that high values of apolipoprotein B and high
HOMA-IR correspond to low values of glomerular filtration rate.
Apolipoprotein B and HOMA-IR were strong influencers of glomerular function
maintaining their statistical significance even after adjusting for multiple cofactors.
Keywords: diabetes mellitus, chronic kidney disease, apolipoprotein B, insulin resistance
Introduction
Chronic kidney disease is a major complication of diabetes mellitus, being estimated that in
the USA half of the patients that need dialysis are patients with diabetic nephropathy [1] and its
prevalence has continued to increase in European countries also [2]. While hyperglycaemia is
itself a risk factor for chronic kidney disease, in patients with type 2 diabetes there is an
enormous aggregation of risk factors some of them non-modifiable (age, family component,
gender, ethnicity) some of them modifiable (hypertension, dyslipidaemia, smoking, obesity)
[1], some of them classic or highly investigated and some of newly discovered. The
manifestation of diabetic kidney disease is represented either by a glomerular filtration rate
(GFR) <60/ml/min/1.73m2 or albuminuria [3]. Many studies investigated the factors that
influence GFR reduction and progression towards chronic kidney disease, one such study
concluded that age, sex, body-mass index, triglycerides and LDL-cholesterol were factors that
influenced the GFR reduction in type 2 diabetes mellitus patients [3]. Among traditional risk
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factors poor glycaemic control, hypertension, decreased pre-existing renal function, presence
of microvascular complications have been well validated and there is a general agreement
regarding their strong influence on progression to diabetic kidney disease, although the role of
certain risk factors such as dyslipidaemia, obesity, smoking, vitamin D status is still
controversial [3]. The importance of studying the factors that lead to a decline in GFR is the
higher cardiovascular risk of patients with diabetes and chronic kidney disease compared to
patients with diabetes but without chronic kidney disease and the possibility of attenuation of
these risk factors in diabetes patients in order to prevent or to delay the evolution towards
chronic kidney disease [4]. Studies usually explore the role of these factors in diabetes patients
with moderately reduced kidney function expressed as GFR ≤60 ml/min/1.73m2 mainly
because it was thought that this cut-off value of GFR is associated with the excess burden of
cardiovascular disease. However recent data suggests that even patients with GFR between 60
and 90 ml/min/1.73m2 are exposed to an increased cardiovascular risk compared to patients
with normal GFR [5, 6].
Therefore, our study has the purpose to explore the prevalence of several risk factors for
chronic kidney disease in diabetes type 2 patients with normal GFR and with mildly reduced
GFR. The majority assessed factors were the traditional ones however we put much emphasis
on the relationship between lipid profile and GFR and an element not so often explored
apolipoprotein B, also the relationship between insulin resistance and GFR was explored.
The study was done between 15 March 2019 and 01 July 2019 including patients with type
2 diabetes mellitus from the Diabetes Clinic of the Clinical County Emergency Hospital of
Oradea that were evaluated during one-day hospitalization. Since the purpose of our study was
to evaluate the factors associated with a slight decrease in kidney function in these patients the
strategy was to determine the prevalence of these factors in a reference group of patients with
a GFR <90 ml/min/1.73m2 but GFR >60 ml/min/1.73m2 which represent patients with mildly
reduced kidney function and in a control group of type 2 diabetes patients with normal kidney
function, with a GFR >90 ml/min/1.73m2. It is important to state that the patients from the
reference group were not patients with chronic kidney disease; they were individuals with
mildly reduced GFR as the purpose of the study was to evaluate the risk factors associated with
an early decrease in glomerular function. Since the prevalence of normal kidney function is
reduced in type 2 diabetes patients, in the first group was included every second patient that
had an altered kidney function and in the second group was included every patient that had a
normal kidney function. The exclusion criteria were: patients with moderate to severe or severe
altered kidney function, GFR <60 ml/min/1.73m2, patients with known kidney disease such as
chronic glomerulonephritis or chronic pyelonephritis, patients with fasting glycaemia more than
300 mg/dL that could lead to a dehydration and falsely decreased GFR, patients with abnormal
urine test including leukocyturia, microscopic or macroscopic haematuria.
Every patient was evaluated clinically, body-mass index (BMI) was calculated and usual
biochemical tests were performed. Also, urine analysis was done for every patient. GFR was
calculated using the MDRD formula. The determination of apolipoprotein B and HOMA-IR
index were done at a private laboratory. Apolipoprotein B was determined using the
immunoturbidimetric method. HOMA-IR index was calculated with the help of the formula:
HOMA-IR = ((fasting insulin (mU/mL) x fasting glucose (mg/dL))/405).

All patients signed a written consent for the participation in the study. The study was
conducted according to the principles of the Helsinki Declaration and with the approbation of
the Ethics Commission of the Clinical County Emergency Hospital of Oradea.
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Results
The main parameters of the included patients are illustrated in Table 1.
Table 1. Clinical-biochemical characteristics of the included patients

GFR 60-90 GFR >90
Parameter ml/min/1.732 SD ml/min/1.732 SD p-value
(n=108) (n=116)
Age (years) 62.48 8.09 56.26 7.94 <0.01
Sex (%men) 66.66% - 63.79% - 0.58
GFR (ml/min/1.73m2) 77.75 8.4 98.61 5.88 <0.01
Creatinine (mg/dL) 0.95 0.14 0.76 0.12 <0.01
SBP (mmHg) 137.84 15.18 128.93 14.45 <0.01
DBP (mmHg) 76.07 9.48 73.82 8.64 0.06
Hypertension (%) 74.07 - 56.89 - <0.01
BMI (kg/m2) 32.17 5.9 30.98 4.89 0.04
HbA1c (%) 7.53 1.33 7.01 1.26 <0.01
Total Cholesterol (mg/dL) 197.32 38.87 174.02 35.11 <0.01
LDL-cholesterol (mg/dL) 113.94 30.56 99.79 26.53 <0.01
Triglycerides (mg/dL) 192.45 (133.5, 228.75) 155.62 (98.25, 194) <0.01
HDL-cholesterol (mg/dL) 43.32 13.07 47.07 13.1 0.03
Trig/HDL-ratio 5.15 3.47 3.83 2.56 <0.01
Apolipoprotein B (mg/dL) 88.96 19.66 81.92 19.3 <0.01
HOMA-IR 3.8 1.98 3.12 2.1 <0.01
Diabetes duration 7.4 4.6 6.8 4.2 0.3
In univariate analysis there is a statistically significant association (p<0.01) between the

values of apolipoprotein B and GFR in patients with type 2 diabetes (Fig. 1) and a statistically
significant association (p<0.01) between HOMA-IR index and GFR (Fig. 2).
Fig. 1. The association between apolipoprotein B and GFR in the included patients, both groups with normal
and low GFR
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Fig. 2. The association between HOMA-IR and GFR in the included patients, both groups with normal
and low GFR
We performed an analysis of factors that influence glomerular function by using multivariate

regression. Age, total cholesterol, HDL-cholesterol, apolipoprotein B were found to be
significant for changes of glomerular function in our diabetic patients, p<0.01.
Discussion
Our study demonstrates that the lipid profile has a major influence on the glomerular
function. Dyslipidaemia is considered a risk factor for chronic kidney disease progression.
Type 2 diabetes patients with normal glomerular function at baseline with high values of
total cholesterol had a statistically significant higher progression toward chronic kidney disease
compared with patients with normal total cholesterol [7]. High levels LDL-cholesterol and high
levels of triglycerides were confirmed as risk factors for albuminuria in type 2 diabetes mellitus
patients in large trials like the DCCT/EDIC study [8]. High triglycerides and high
triglycerides/HDL-cholesterol values were associated with higher creatinine and albuminuria
in a study that included over 10000 patients [9]. The presence of diabetic dyslipidaemia with
its’s characteristic high levels of total cholesterol, high levels of small particles of LDL-
cholesterol, hypertriglyceridemia and low HDL-cholesterol is a cause of atherosclerosis in the
kidney microcirculation, but also lipid glomerular infiltration leads to increased glomerular
inflammation and fibrosis [10]. The renal structures affected by lipid accumulation are the
mesangial cells and the glomerular podocytes because of inflammation and excess matrix
production resulting in glomerulosclerosis [11]. The target values defined by American
Diabetes Association (ADA) for lipid parameter are LDL-cholesterol <100mg/dl, triglycerides
<150 mg/dl, HDL-cholesterol <40mg/dl for men and <50mg/dl [12].
Therefore, screening for dyslipidaemia is mandatory in every type 2 diabetes mellitus patient
and should be included in the periodic evaluation of the status of every patient [13, 14].
Apolipoprotein B is an indicator of the total number of circulating atherogenic particles and
therefore apolipoprotein B level correlates with the intensity of cardiovascular disease [15]. In
our study was dosed the total level of apolipoprotein B; it is known that apolipoprotein B
circulates in 2 isoforms apolipoprotein B-48 and apolipoprotein B-100.
Previous research demonstrated that high apolipoprotein B-48 levels corresponded to lower
levels of GFR and increased proteinuria [16]. Plasma high levels of apolipoprotein B-100
correspond to elevated circulating levels of triglyceride-rich lipoproteins [17], indeed this was
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confirmed in our study and it its known that hypertriglyceridemia negatively influences the
local glomerular environment with a high cytokine secretion as a response to lipid loading [18].
Apolipoprotein B proved to be in our study a strong determinant of glomerular filtration rate,
with high values of apolipoprotein B corresponding to low values of GFR even after adjusting
for multiple factors.
Conclusion
In our study hypertension, poor glycaemic control and body-mass index were confirmed as
risk factors for mild glomerular function alteration. These factors are the classical, traditional
risk factors and their control is important, but studies prove that despite all effort’s hypertension
control, HbA1C<7% and normal weight are rarely achieved [19]. It must be stated that despite
the control of all risk factors some diabetes patients still progress towards chronic kidney
disease because of genetic factors [20, 21] environmental factors [22] or aging [23], in our study
age was a strong determinant of a reduced glomerular filtration rate and it kept the statistical
significance even in multivariate analysis.
Apolipoprotein B and HOMA-IR were strong influencers of glomerular function
maintaining their statistical significance even after adjusting for multiple cofactors.
Therefore, apolipoprotein B and HOMA-IR emerge as non-traditional risk factors for
glomerular reduced function in type 2 diabetes mellitus patients. This would allow for early
detection of a change in renal function and allows a different management of disease for slowing
progression to end stage renal disease.
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4. Ninomiya, T. Perkovic, V. De Galan, BE, et al., (2009). Albuminuria and Kidney Function
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disease in patients with type 2 diabetes. Kidney Int. 85(5): pp. 1192-1199.
6. Manea, M, Marcu, D, Motofei, I et al., (2019). Cardiovascular risk in patients with inflammatory bowel
diseases: a review. Rom Biotechnol Lett. 24(2): pp. 366-373.
7. Ravid, M, Brosh, D, Ravid-Safran, et al., (1998). Main risk factors for nephropathy in type 2 diabetes
mellitus are plasma cholesterol levels, mean blood pressure, and hyperglycaemia. Arch Intern Med.
158(9): pp. 998-1004.
8. De Boer, IH, Rue, TC, Cleary, PA, et al., (2011). Long-term renal outcomes of patients with type 1
diabetes mellitus and microalbuminuria: an analysis of the Diabetes Control and Complications
Trial/Epidemiology of Diabetes Interventions and Complications cohort. Arch Intern Med.171(5): pp.
412-420.
9. Aziz, K. (2017). Association of high serum triglycerides and triglycerides/HDL ratio with raised
HbA1c, creatinine, microalbuminuria and development of diabetic kidney disease and diabetic renal
failure. Mathematical and statistical regression models of 10,370 diabetic patients. Clin Nephrol Res.
1(1): pp. 17-25.
10. Abrass. CK (2004). Cellular lipid metabolism and the role of lipids in progressive renal disease. Am J
Nephrol. 24(1): pp. 46-53.
11. Moorhead, JF, Chan, MK, El-Nahas, M, Varghese, Z. (1982). Lipid nephrotoxicity in chronic
progressive glomerular and tubulo-interstitial disease. Lancet. 2: pp. 1309-1311.
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13. Daina, LG, Neamtu, C, Daina, CM. (2017). Evaluating the analgesic consumption in a clinical
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14. Silaghi CN, Fodor D, Gheorghe SR, Crăciun AM. (2019). Serum total matrix Gla protein: Reference
interval in healthy adults and variations in patients with vascular and osteoarticular diseases. Clin Chim
Acta. 490: pp. 128-134.
15. Marcovina, S, Packard, CJ. (2006). Measurement and meaning of apolipoprotein AI and apolipoprotein
B plasma levels. J Intern Med. 259: pp. 437-46.
16. Okubo, M, Hanada, H, Matsui, M, et al., (2014). Serum apolipoprotein B-48 concentration is associated
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The Prevalence of Cardiovascular Risk Factors Control among

Type 2 Diabetes Mellitus Patients – Epidemiological Study in a
Tertiary Care Hospital from North Western Romania
POPOVICIU Mihaela Simona1,2, POPA Loredana2, VESA Cosmin Mihai1,2,

MAGHIAR Octavian1, ZAHA Andreea Atena3, BILUȚĂ Adina2,
BUNGAU Alexa1, POPESCU Ioachim Mircea1,2, POPA Amorin Remus1,2
1 Faculty of Medicine and Pharmacy, University of Oradea (ROMANIA)
2 Clinical County Emergency Hospital, Oradea (ROMANIA)
3 “Iuliu Hatieganu” University of Medicine and Pharmacy, Cluj-Napoca (ROMANIA)
Email: v_cosmin_15@yahoo.com
Abstract
Control of all cardiovascular risk factors is the universal desiderate in type 2 diabetes
mellitus Our study is a real-world data study, performed in a tertiary care hospital in north
western Romania, that aimed to discover the percentage of patients with type 2 diabetes that
achieve cardiovascular risk factors control. Systematic sampling was performed in order to
ensure that the results are representative. Among the 167 evaluated patients, 42.51% had a value
of HbA1c<7%, while 57.49% had a value of HbA1c≥7%. Poor glucose control was associated
with statistically significant higher values of BMI, systolic blood pressure, fasting glycaemia,
total cholesterol, LDL-cholesterol, triglycerides, and C-reactive protein. The prevalence of
metabolic syndrome among patients with HbA1c<7% was 45.71% statistically significantly
lower than in patients with HbA1c≥7% were it was 61.86%. 34.73% of patients had
SBP<140mmHg and DBP<90mmHg while only 12.57% had SBP<130mmHg and
DBP<80mmHg. 34.73% had LDL-cholesterol lower than 100mg/dL while only 14.37% had
LDL-cholesterol lower than 70mg/dL. Patients with good glucose control had a better control
of blood pressure targets, statistically significant in the case of the target of SBP<130mmHg
and DBP<80mmHg, and LDL-cholesterol target. Overall a very small percent of patients,
8.98%, achieved SBP<130mmHg, DBP<80mmHg, LDL-cholesterol<100mg/dL and
HbA1c<7% and almost none achieved more demanding targets.
Keywords: glucose control, cardiovascular risk factors control
1. Introduction
Numerous studies support the idea that multifactorial interventions in patients with type 2
diabetes mellitus addressing glycaemic control, blood pressure control and lipid control are
necessary to reduce the incidence of major complications. Early trials such as UKPDS
demonstrated that a better glucose control with an average HbA1c of 7.8% in the intensive
groups vs. 8% in the conventional group resulted in a 36% reduction of all-cause mortality [1].
Another study that enrolled patients from the Steno-2 study demonstrated that aggressive
control of cardiovascular risk factors, HbA1c<6.5%, total cholesterol<175mg/dL and blood
pressure<130/80mmHG, for an initial period of 7.8 years resulted in a 20% less mortality of all
cause in patients with the above targets in a period of 13.3 years of fallow-up compared with
patients with less aggressive targets [2]. Most important is that the study’s design reconfirms
the concept of metabolic memory, first demonstrated by the DCCT/EDIC study, because after
361
the initial period of 7.8years of aggressive therapy in one group and conventional therapy in the
other, all patients were put on an aggressive target aimed therapy [2]. This evidence supports
the idea that not only the targets are important in the control of cardiovascular risk factors in
type 2 diabetes mellitus patients but most important is how fast the patients reach the targets.
However, promising these results may be data from epidemiological studies that evaluate
the degree of control of cardiovascular risk factors in populations of type 2 diabetes patients are
alarming.
Our study has the purpose to evaluate the degree of glucose, blood pressure and lipid control
among type 2 diabetes mellitus patients from north western Romanian evaluated in the Clinical
County Emergency Hospital of Oradea. Also, the study aims to explore the differences among
type 2 diabetes mellitus patients with good and poor glucose control.
We included in our study 167 patients with type 2 diabetes mellitus from the Clinical County
Emergency Hospital of Oradea. The inclusion criteria in the study were: patients between aged
18 years or above, patients with type 2 diabetes mellitus, patients that gave their acceptance for
the participation in the study. Exclusion criteria were: patients with type 1 diabetes mellitus,
patients suffering from neoplastic diseases, psychiatric diseases, endocrine diseases such as
hypothyroidism, Cushing disease and patients taking medications that can influence glycaemia,
blood pressure or lipid values such as corticosteroids. Patients that presented at the Clinical
County Emergency Hospital of Oradea for diabetes for routine medical evaluation and
medication prescription in the period 01 June-01 October 2019 were considered for the
inclusion in the study. In order that the results of the study to be epidemiologically relevant the
fallowing method for the inclusion in the study was performed.
The first two patients that presented for evaluation in all the days of the above specified time
interval were considered for inclusion in the study. A total of 185 patients were evaluated but
after applying the inclusion and exclusion criteria 167 patients remained in the study. The
research was conducted with the approbation of the Ethical Commission of the hospital. For
each patient we performed the anamnesis, physical exam, medical history evaluation and the
assessment of the therapeutic regimens that he fallowed. Blood pressure was measured and
body-mass index was calculated. Blood pressure was measured according to 2018 ESC/ESH
Guidelines for the management of arterial hypertension. Laboratory work-up was performed
including: CBC, urea, serum creatinine, HbA1c, total cholesterol, LDL-cholesterol, HDL-
cholesterol, triglycerides, C-reactive protein. Different targets were used for assessing
cardiovascular risk factors control. Glucose control was considered if HbA1c was <7%
according to ADA 2019 criteria and 2019 ESC Guidelines on diabetes, pre-diabetes, and
cardiovascular diseases developed in collaboration with the EASD criteria. For blood pressure
control it was determined the percentage of patients achieve the blood pressure target
<140/90mmHg (ADA 2019 criteria) but also it was determined the percentage of patients with
blood pressure <130/80mmHG (2019 ESC Guidelines). Concerning LDL-cholesterol control,
targets are individualised according to the 2 guidelines but since the nature of this study is an
epidemiological one, patients were divided in groups that have LDL-cholesterol<100 mg/dL
and then <70mg/dL.
Statistical analysis was done Biostat Software, a value of p<0.05 was considered statistically
significant. ANOVA test for independent means was performed to compare the parameters
between groups.
362
3. Results
Among the patients included in the study less than one half (42.51%) had an HbA1c<7%
(Fig. 1).
Fig. 1. Prevalence of glucose control among the included patients
Patients with poor glucose control had statistically significant higher body mass index,
systolic blood pressure, total cholesterol, LDL-cholesterol, triglycerides, TRIG/HDL-ratio and
C-reactive protein (Table 1).
Table 1. Clinic-biochemical parameters pf the included patients

HbA1c<7 Standard Deviation/ HbA≥7 Standard Deviation/
Parameter p
(n=71) (Q1, Q3) (n=96) (Q1, Q3)
Age (years) 64.57 10.28 61.05 10.48 0.12
Sex (%men) 54.29 - 54.64 - 0.96
Living environment (% urban) 65.71 - 72.16 - 0.37
Diabetes duration (years) 8.67 6.62 9.31 5.22 0.48
BMI (kg/m2) 30.23 5.39 31.9 6.37 0.04*
SBP (mmHg) 138.84 16.96 145.55 22.01 0.03*
DBP (mmHg) 83.71 13.1 85.74 11.57 0.19
HbA1c (%) 6.18 0.5 8.97 1.81 <0.01*
Fasting glycaemia (mg/dL) 125.68 20.38 210.72 82.36 <0.01*
Total Cholesterol (mg/dL) 181.34 43.39 204.41 55.27 <0.01*
LDL-cholesterol (mg/dL) 110.58 37.97 126.63 52.71 0.03*
Triglycerides (mg/dL) 134.65 (84, 167.75) 208.4 (115, 227) <0.01*
HDL-cholesterol (mg/dL) 46.53 16.62 42.6 11.39 0.07
TRIG/HDL ratio 3.31 2.7 5.41 4.25 <0.01*
GRF (ml/min/1.73m2) 72.93 24.17 67.74 22.27 0.15
C-reactive Protein (mg/dL) 0.81 (0.22, 0.99) 1.67 (0.19, 1.2) <0.01*
*Statistically significant
Microvascular complications, polyneuropathy and retinopathy, are statistically significant

more prevalent in patients with poor glucose control. Macrovascular complications are more
prevalent in patients with poor glucose (Table 2).
Table 2. Microvascular and macrovascular complications

HbA1c<7 HbA≥7
Parameter p
No (%) No (%)
Polyneuropathy 40(57.14) 72(74.23) 0.02*
Retinopathy 13(18.57) 34(35.05) 0.01*
CKD 21(30) 36(37.11) 0.34
Ischemic heart disease 40(57.14) 70(72.16) 0.04*
Heart failure 19(27.14) 36(37.11) 0.17
History of myocardial infarction 2(2.86) 10(10.31) 0.66
History of stroke 3(4.29) 8(8.25) 0.31
Peripheral artery disease 18(25.71) 24(24.74) 0.88
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Metabolic syndrome is statistically significant (p<0.05) more prevalent among patients with
poor glucose control.
Fig. 2. Prevalence of metabolic syndrome among patients with good and poor glucose control
Table 3 and Fig.3 illustrate how many patients achieve blood pressure control and how many
patients achieve lipid control. Overall 34.73% achieve blood pressure control according to ADA
general criteria while only 12.57% achieve blood pressure control according to 2019 ESC
Guidelines on diabetes. An LDL<100mg/dL was achieved by 34.73% of diabetes mellitus
patients and an LDL<70mg/dL was achieved by 14.37%. It can be observed that type 2 diabetes
mellitus patients with good glucose control achieve more frequently blood pressure and lipid
control when compared with patients with poor glucose control.
Table 3. Prevalence of blood pressure control and lipid control according to HbA1c groups
HbA1c<7% HbA≥7% All patients
p
No (%) No (%) %
SPP<140mmHG and DBP<90mmHg 35(36.08) 23(32.86) 34.73 0.66

SPB<130mmHg and DBP<80mmHg 14(20) 7(7.22) 12.57 0.01*
LDL<100mg/dL 30(42.86) 28(28.87) 34.73 0.06
LDL<70mg/dL 12(17.14) 12(12.37) 14.37 0.38
Fig. 3. Prevalence of blood pressure control and lipid control according to HbA1c groups
When analysing how many patients achieve the control of all cardiovascular risk factors, it
was revealed that a conservative approach, HbA1<7% and SBP<140mmHg and
DBP<90mmHg, LDL<100mg/dL, was achieved by 8.98% of patients while an intensive
approach, HbA1c<7%, SBP<130mmHg, DBP<80mmHg, LDL<70mg/dL was achieved by
4.79% of patients.
364
Table 4. Prevalence of control of all cardiovascular risk factors according to different criteria
Parameter No (%)
HbA1<7% and SBP<140mmHg and DBP<90mmHg, LDL<100mg/dL 15 (8.98)
HbA1c<7%, SBP<130mmHg, DBP<80mmHg, LDL<70mg/dL 8(4.79)
4. Discussion
In a study performed in USA evaluating a health record database among 31.340 patients with
type 1 diabetes mellitus only 20% of them had a HbA1c <7% [3]. Another study performed in
China on an extremely large population of 238 656 patients with type 2 diabetes mellitus
revealed that only 31.78% of patients achieved HbA1c<7% [4]. In our study 42.51% of patients
had an Hba1c <7%. Worldwide despite the advances made in diabetes therapy, the numerous
diabetes screening programmes, the understanding of diabetes importance and its associated
cardiovascular risk, the prevalence of uncontrolled diabetes varies between 45% and 75% [5].
When discussing the control of all three major cardiovascular risk factors, only 11.2% of
patients in a study performed in Japan reached blood pressure<140/90mmHg, HbA1c<7% and
LDL-cholesterol <100mg/dL [6]. The data from Bihor County obtained in our study are
comparable to the study performed in Japan, a country with a higher socio-economic
development, in our county 8.98% of diabetes mellitus patients have a HbA1<7%,
SBP<140mmHg and DBP<90mmHg, LDL<100mg/dL. It should be noted that these targets are
not the most demanding, an LDL<70mg/dL would be more appropriate for a population with
numerous cardiovascular risk factors. The importance of glucose control is visible even in our
study, where patients with glucose control had statistically significant higher prevalence of
diabetic neuropathy, diabetic polyneuropathy and ischemic heart disease. The finding that lack
of glycaemic control is associated with lack of hypertension control and blood lipids control in
our study is supported by similar findings, one study reported that patients with poor glucose
control had statistically significant higher values of systolic blood pressure and diastolic blood
pressure [7]. Lack of glycaemic control in diabetes mellitus is associated with increased
cardiovascular mortality, but also with high risk of infectious diseases [8] or malignancy [9,
10]. Why don’t patients with type 2 diabetes mellitus achieve control of cardiovascular risk
factors? The answer is complex but poor eating habits contribute to this situation, a study
realized in Japan confirmed that poor eating habits are associated with poor glycaemic control
and obesity [11]. Lack of treatment adherences could be another reason why type 2 diabetes
mellitus patient do not achieve glucose control, studies demonstrating that only one third of the
patients that receive antidiabetic medications have an adequate adherence [12]. This could
apply to antihypertensive therapy and lipid lowering therapy, it was demonstrated that
adherence to therapy for chronic diseases in developed countries is about 50% on average [13].
Development of innovative medications such as long acting GLP-1’s with weekly
administration and modern methods of drug administration such as single-dose prefilled pens
demonstrated in recent studies to be associated with a better adherence to therapy and a higher
degree of patient’s acceptance of the drug [14].
5. Conclusion
Data from a tertiary care hospital from Bihor county demonstrates that percentage of patients
with type 2 diabetes that achieve glucose control is 42.51% and even lower for blood pressure
and lipid control. Lack of glucose control is associated with lack of hypertension and blood
lipids control. The data from our county is in concordance to the worldwide situation.
365
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with type 2 diabetes (UKPDS 34). UK Prospective Diabetes Study (UKPDS) Group. Lancet. 1998
Sep 12; 352(9131): pp. 854-65. Erratum in: Lancet 1998 Nov 7; 352(9139): p. 1558.
2. Gaede P, Lund-Andersen H, Parving HH, Pedersen O. (2008). Effect of a multifactorial intervention
on mortality in type 2 diabetes. N Engl J Med. 358(6): pp. 580-91.
3. Pettus JH, Zhou FL, Shepherd L, Preblick R, Hunt PR, Paranjape S, Miller KM, Edelman SV. (2019).
Incidences of Severe Hypoglycaemia and Diabetic Ketoacidosis and Prevalence of Microvascular
Complications Stratified by Age and Glycaemic Control in U.S. Adult Patients with Type 1 Diabetes:
A Real-World Study. Diabetes Care. 42(12): pp. 2220-2227
4. Ji LN, Lu JM, Guo XH, et al., (2013). Glycaemic control among patients in China with type 2 diabetes
mellitus receiving oral drugs or injectables. BMC Public Health. 13: p. 602.
5. Al Mansari A, Obeid Y, Islam N, et al., (2018). GOAL study: clinical and non-clinical predictive
factors for achieving glycaemic control in people with type 2 diabetes in real clinical practice. BMJ
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6. Hu H, Hori A, Nishiura C, et al., (2016). Hba1c, Blood Pressure, and Lipid Control in People with
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Cardiovascular Risk Factors in Patients with Poorly Controlled Diabetes Mellitus. Med Arch.72(1):
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8. Zaha, DC, Bungau, S, Aleya, S, et al., (2019). What antibiotics for what pathogens? The sensitivity
spectrum of isolated strains in an intensive care unit, Science of The Total Environment, Volume 687,
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11. Gouda M, Matsukawa M, Iijima H. (2018). Associations between eating habits and glycaemic control
and obesity in Japanese workers with type 2 diabetes mellitus. Diabetes Metab Syndr Obes. 11: pp.
647-658.
12. García-Pérez LE, Alvarez M, Dilla T, Gil-Guillén V, Orozco-Beltrán D. (2013). Adherence to
therapies in patients with type 2 diabetes. Diabetes Ther. 4(2): pp. 175-194.
13. World Health Organization. Adherence to Long-Term Therapies: Evidence for Action. Geneva:
WHO; 2003.
14. Giorgino F, Penfornis A, Pechtner V, Gentilella R, Corcos A. (2018). Adherence to antihyperglycemic
medications and glucagon-like peptide 1-receptor agonists in type 2 diabetes: clinical consequences
and strategies for improvement. Patient Prefer Adherence.12: pp. 707-719.
366
Challenges in Treating Erectile Dysfunction in Diabetic Patients –

Review
MĂDAN Victor1,2, PAVALEAN Mihai1, PACU Ovidiu1, PAVALEAN Mihaela1,

BUCURICA Sandica1,2, DIMITRIU Mihai2,3, PACU Irina2,3
1 Department of Urology, “Dr Carol Davila” Central Military Emergency University Hospital, Bucharest (ROMANIA)
Emails: victmad@gmail.com, pavamisu@yahoo.com, ovidiupacu@live.com, bucuricasandy@yahoo.com,

drmihaidimitriu@yahoo.com, irinapacu@hotmail.com
Abstract
Erectile dysfunction (ED) represents the persistent inability to achieve or maintain penile
erection for satisfactory sexual intercourse. Diabetes mellitus (DM) is the most frequent
aetiology for ED, and in patients with DM, its prevalence is high. Multiple factors contribute to
this pathology; the significant role in diabetes-related ED have been considered vascular
abnormalities, while the impact of neuropathy has been underestimated. There are many
treatment options for ED, the most used being oral therapy with phosphodiesterase type-5
(PDE5) inhibitors. Recently, more treatments have been proposed and tested in clinical trials
for refractory cases with promising results.
Keywords: diabetes mellitus; erectile dysfunction; neuropathy; phosphodiesterase type-5 inhibitors;
low-intensity extracorporeal shockwave therapy
Background
Diabetes mellitus (DM) is one of the most prevalent and burden disorders in the whole world.
Accordingly, o the World Health Organization Global Report on Diabetes, the number of
patients with DM had grown from 108 million in 1980 to 422 million in 2014 [1].
Erectile dysfunction (ED) is one of the urologic complications of diabetes that affects men,
among lower urinary tract symptoms, low sexual desire, and orgasmic dysfunction. As people’s
lifestyle changes and the population ages, the incidence of diabetic mellitus erectile dysfunction
(DMED) continues to increase. Many improvements were made to reduce the severity of other
diabetes long term complications like retinopathy, nephropathy, and neuropathy, but urological
complications may still negatively affect the quality of life [2].
Erectile dysfunction (ED) aetiology is multifactorial, including endocrine, neurological,
vascular, systemic disease, local penile disorders, nutrition, psychogenic factors, and drug-
related. Current evidence suggests that up to 80% of cases have organic causes which are
subdivided into vasculogenic, neurogenic and hormonal etiologies [3]. Peripheral and
autonomic neuropathies, as well as a macrovascular disease, were significantly more frequent
among men with ED. [4, 5]
The physiology of erection is a complex interplay of psychogenic, hormonal and
noradrenergic, noncholinergic neurovascular mechanisms. Nitric oxide (NO) is released from
the endothelium of the corpora cavernosa and cavernous nerves in response to local physical or
central sexual stimulation. NO activates soluble guanylyl cyclase resulting in an increase in
cyclic guanosine monophosphate (cGMP) levels, which causes smooth muscle relaxation and
arteriolar dilation leading to penile erection. Thus, penile erection involves both autonomic
367
(sympathetic and parasympathetic) and somatic (motor and sensory) nerves supplying the shaft
and glans of the penis. The aetiology of ED in diabetes is considered to be due to both vascular
and neuronal dysfunction, and indeed PDE5 inhibitors promote NO release and mediate
increased penile blood flow. The relationship between ED and neuropathy is complex, but
studies have demonstrated evidence of large and small fibre neuropathy and autonomic
neuropathy in diabetic patients with ED [5].
Patients with diabetes mellitus (DM) have high rates of ED [6, 7]. Investigations in animal
models and diabetic patients reported that primarily responsible for ED are alterations in neural
and impaired penile vascular systems [8].
The necessity for a diagnostic evaluation for ED has been questioned after the availability
of effective oral therapies [9], as many patients could not have the arteriogenic component
involved, so it is necessary to assess the occurrence of penile neuropathy in diabetic patients.
Treatment includes oral drugs like PDE5 inhibitors, testosterone therapy, penile
intracavernosal injections (ICI), intraurethral medication (IU), vacuum erection devices, penile
implants and surgery.
Pharmacotherapy is the primary treatment for ED, including PDE5 inhibitors, androgen
therapy, and vasoactive agents [10]. Diabetic patients with ED represent a cluster of difficult-
to-treat ED population because the efficacy of oral therapies is lower if compared to non-
diabetic patients. In this systematic review, we aim to evaluate the effectiveness and safety of
PDE5 inhibitors for diabetic mellitus erectile dysfunction and to search the efficacy of other
treatments options for refractory cases.
Methodology
We searched PubMed, Cochrane Library, AMED, EMbase, WorldSciNet; Nature, Science

online and China Journal Full-text Database (CNKI). The search terms include erectile
dysfunction, diabetes mellitus, PDE5-inhibitors, diabetic mellitus erectile dysfunction,
impotence, treatment options for erectile dysfunction, clinical trials, including randomized
controlled trials of the control group (effective methods other than PDE5-inhibitors).
Results
Oral Drugs (PDE5 inhibitors) and neuropathy

Studies have shown that PDE5-inhibitors treatment of Diabetes Mellitus ED can improve
the International Index of Erectile Function-5 (IIEF-5) score and sexual success rate in a
considerable number of patients, especially in those with a normal nerve function to the penis.
Besides, meta-analyses have shown that PDE5-inhibitors can safely and effectively treat ED,
whether they are still safe and effective for diabetic mellitus erectile dysfunction with more
complex etiologies remains to be assessed [11].
The relationship between ED and neuropathy is complex, but studies have demonstrated
evidence of large and small fibre neuropathy and autonomic neuropathy in diabetic patients
with ED [12]. In a cohort of 341 patients with ED, the prevalence of neuropathy assessed using
nerve conduction studies and QST was 38% in those with diabetes and 10% in those without
diabetes. However, the incidence of neuropathy among those with neurogenic (21%) compared
with vasculogenic (23%) ED was comparable [13]. Wellmer et al., showed no difference in the
neurological examination or thermal sensory thresholds. Still, there was a difference in the
capsaicin-induced sensory axon reflex and sural nerve amplitude between diabetic patients with
and without ED [14]. Studies have also shown a link between measures of cardiac autonomic
neuropathy (CAN) and ED. Penile vaso-tactile and thermal thresholds assessing somatic small
fibre neuropathy are abnormal in diabetic patients with ED [14].
368
The management of ED in patients with type 2 diabetes is challenging with a non-responder

rate of over 50% for PDE5 inhibitors, which may reflect a more severe neurogenic component
for ED in these patients. In recent research, the assessment of nocturnal penile tumescence and
rigidity, which reveals predominantly neurogenic differences, had an area under the curve
(AUC) of 0.860 in differentiating sildenafil responders from non-responders.
Corneal confocal microscopy (CCM) has been used to identify an association between ED
and small fibre damage in subjects with obesity, type 1 diabetes and now, type 2 diabetes. The
potential role of CCM as an objective marker for neurogenic abnormalities that may predict the
response to therapy in ED warrants further study [15].
Clinical trials and new treatment options

Low-intensity extracorporeal shockwave therapy (LiESWT) it is one of the newest and most
promising treatment modalities for patients with ED [16]. The rationales behind this treatment
involve tissue regeneration and neoangiogenesis [17]. Shreds of evidence from in vitro
experiments showed that vascular stress induced by extracorporeal shockwaves stimulates the
release of angiogenic factors like as vascular endothelial growth factor, endothelial nitric oxide
synthase, and proliferating cell nuclear antigen, thus improving local blood supply and
emodynamics [18]. The controlled generated microtrauma can also rise the macrophage activity
and synthesis of cellular proteins and may potentially grow the recruitment and differentiation
of stem cells [19]. Several studies have been demonstrated the efficacy of extracorporeal
shockwaves in the field of andrology [18, 19]. Besides, treatment is well tolerated and can be
performed without anaesthesia in an outpatient setting. In addition, the different LiESWT setup
parameters and treatment protocols have been published, and the optimal approach strategy
remains to be defined. Preclinical evidence deriving from rat models with type 2 DM have
established a synergistic effect of LiESWT and PDE5Is and that the benefits provided by
LiESWT are independent of nitric oxide and cyclic guanosine monophosphate pathways [20].
In humans, synergistic and/or an additive interplay could be hypothesized, and other
investigations are needed to elucidate these aspects.
A recent retrospective study made by Verze et al., investigated the efficacy of the combined
therapy imply LiESWT performed with an electrohydraulic source and tadalafil 5 mg once daily
in patients with T2DM and ED and compared LiESWT protocols characterized by different
numbers of shockwaves administered during each session. Their results demonstrate that the
combined approach provides advantages in terms of both magnitudes of mean International
Index of Erectile Function (IIEF-5) score improvement and durability of results if compared to
tadalafil 5 mg once daily alone during 24 weeks follow-up [21].
Even though both tadalafil 5 mg once daily alone and the combined treatment involving
tadalafil 5 mg once daily and LiESWT, the procedure rises statistically significant improvement
in terms of mean IIEF-5 score. The scale of variation observed in patients receiving the latter
treatment modality was significantly higher at 12- and 24-month follow-up in the cluster of
patients who received 2400 shockwaves per session [21]. These results corroborate data from
a recently published meta-analysis performed by Man and Li showed a statistically significant
higher amelioration of erectile function in patients receiving LiESWT combined with PDE5
inhibitors concerning patients receiving LiESWT alone and that protocols adopting higher
number of shockwaves per treatment reported a considerable increase of erectile function
compared with protocols delivering fewer shockwaves [22].
The energy from the acoustic waves delivered by LiESWT has been hypothesized to spur
the cellular and molecular pathways, strengthening the expression of local growth factors,
improving endothelial function, angiogenesis, and regeneration of nerve fibres [23, 24].
369
Consequently, LiESWT has the potential to restore physiological erections to cure the
disease [23]. Based on the results of the study made of Verze et al., diabetic ED patients can
benefit from the combined procedure with LiESWT and daily tadalafil.
Conclusion
Erectile dysfunction has a high prevalence in diabetic patients that have treatment difficulties
compared with non-diabetic patients regarding the efficacy of oral therapies.
Neuropathy should be evaluated properly in this category of patients, especially if they are
non-responders to PDE5 inhibitors therapy. Recently, for this category of patients was found
and tested new treatment modalities, for example, the low-intensity extracorporeal shockwave
therapy (LiESWT) with promising results.
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M. (2001) Erectile dysfunction and lower androgenicity in type 1 diabetic patients. Diabetes and
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3. Miller, T. A. (2000) Diagnostic evaluation of erectile dysfunction. American Family Physician 61, pp.
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function: a controlled study. Diabetologia 36, pp. 745-751.
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role of medical comorbidities and lifestyle factors. The Urologic Clinics of North America 32, pp. 403-
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7. El Latif, M. A., Makhlouf, A. A., Moustafa, Y. M., Gouda, T. E., Niederberger, C. S. & Elhanbly, S. M.
(2006) Diagnostic value of nitric oxide, lipoprotein (a), and malondialdehyde levels in the peripheral
venous and cavernous blood of diabetics with erectile dysfunction. International Journal of Impotence
Research 18, pp. 544-549.
8. Lue, T. F. (2000) Erectile dysfunction. New England Journal of Medicine 342, pp. 1802-1813.
9. Hatzichristou, D., Hatzimouratidis, K., Bekas, M., Apostolidis, A., Tzortzis, V. & Yannakoyorgos, K.
(2002) Diagnostic steps in the evaluation of patients with erectile dysfunction. Journal of Urology 168,
pp. 615-620.
10. Gaman M, Dobrica Elena C, Pascu EG, et al., (2019).Cardio metabolic risk factors for atrial fibrillation
in type 2 diabetes mellitus: Focus on hypertension, metabolic syndrome and obesity. Journal of Mind and
Medical Sciences 6(1), pp. 157-161.
11. Porst H, Gacci M, Büttner H, et al., Tadalafil once daily in men with erectile dysfunction: an integrated
analysis of data obtained from 1913 patients from six randomized, double-blind, placebo controlled,
clinical studies. Eur Urol 2014; 65: pp. 455-64.
12. Pantea-Stoian A, Pituru SM, Hainarosie, R, et al., (2018). Testosterone therapy, new opportunities in
diabetes mellitus. Farmacia 66(1), pp. 1-7.
13. Vardi Y, Sprecher E, Kanter Y, Livne PM, Hemli JA, Yarnitsky D. Poly-neuropathy in impotence. Int J
Impot Res. 1996; 8(2): pp. 65-68.
14. Wellmer A, Sharief MK, Knowles CH, et al., Quantitative sensory and autonomic testing in male diabetic
patients with erectile dysfunction. BJU Int. 1999; 83(1): pp. 66-70.
15. Chen X, Graham J, Dabbah MA, et al., Small nerve fiber quantification in the diagnosis of diabetic
sensorimotor polyneuropathy: comparing corneal confocal microscopy with intraepidermal nerve fiber
density. Diabetes Care. 2015; 38(6): pp. 1138-1144.
16. Chung E, De Young L, Brock GB. Investigative models in erectile dysfunction: a state-of-theart review
of current animal models. J Sex Med 2011; 8: pp. 3291-305.
17. Chung E, Wang J. A state-of-art review of low intensity extracorporeal shock wave therapy and
lithotripter machines for the treatment of erectile dysfunction. Expert Rev Med Devices 2017; 14: pp.
929-34.
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18. Ruffo A, Capece M, Prezioso D, Romeo G, Illiano E, et al., Safety and efficacy of low intensity
shockwave (LISW) treatment in patients with erectile dysfunction. Int Braz J Urol 2015; 41: 9 pp. 67-74.
19. Palmieri A, Imbimbo C, Creta M, Verze P, Fusco F, et al., Tadalafil once daily and extracorporeal shock
wave therapy in the management of patients with Peyronie’s disease and erectile dysfunction: results
from a prospective randomized trial. Int J Androl 2012; 35: pp. 190-5.
20. Assaly-Kaddoum R, Giuliano F, Laurin M, Gorny D, Kergoat M, et al., Low intensity extracorporeal
shock wave therapy improves erectile function in a model of type II diabetes independently of NO/cGMP
pathway. J Urol 2016; 196: pp. 950-6.
21. Verze P, Capece M, Creta M, La Rocca R, Persico F, Spirito L, Cardi A, Mirone V. Efficacy and safety
of low-intensity shockwave therapy plus tadalafil 5 mg once daily in men with type 2 diabetes mellitus
and erectile dysfunction: a matched-pair comparison study. Asian J Androl 2019 [Epub ahead of print].
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P, et al., Low-intensity shockwave therapy for erectile dysfunction: Is the evidence strong enough? Nat
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24. Silea C, Cucu IA, Zarnescu O, et al., (2019).Influence of age on sperm parameters in men with suspected
infertility. Romanian Biotechnological Letters 24(1), pp. 82-90.
371
New Data about Urinary Bladder Complications of Diabetes

mellitus – A Review
MĂDAN Victor1,2, PAVALEAN Mihai1, PACU Ovidiu1, PAVALEAN Mihaela1,

BUCURICA Sandica1,2, DIMITRIU Mihai2,3, PACU Irina2,3
1 Department of Urology, “Dr. Carol Davila” Central Military Emergency University Hospital, Bucharest (ROMANIA)
Emails: victmad@gmail.com, pavamisu@yahoo.com, ovidiupacu@live.com, bucuricasandy@yahoo.com,

drmihaidimitriu@yahoo.com, irinapacu@hotmail.com
Abstract
Diabetes mellitus is a worldwide systemic disorder that causes multisystem damages and
increases mortality and morbidity. Diabetes commonly affects the bladder nerves, so frequent
complications of diabetes include erectile dysfunction, urinary incontinence overactive bladder
and diabetic bladder dysfunction. Another frequent complication is represented by lower
urinary tract infection. We aim to detail these diabetes bladder complications in this review
with new literature data.
Keywords: diabetes mellitus, overactive bladder, lower urinary tract infection, sexual dysfunction.
Introduction and bjectives
Diabetes mellitus (DM) is a disease characterized by severe complications such as

retinopathy, nephropathy and neuropathy.
Urological complications are widespread health problems, whose incidence increases with
age and diabetic duration [1]. Erectile dysfunction, lower urinary tract infections, sexual
dysfunction and urinary incontinence in the female are the most common urological
complications.
Diabetic neuropathy is categorized into diabetic peripheral neuropathy (DPN) and
autonomic neuropathy. Diabetic bladder dysfunction is a type of automatic neuropathy that
manifests as lower urinary tract involvement or as overactive bladder (OAB).
One of the most common complications of diabetes mellitus is bladder dysfunction, and we
can find this type of pathology in half of all diabetic patients. DM and its associated
complications have a significant impact on patients’ quality of life. The magnitude of the effects
of different DM complications on life quality is diverse.
Methods
In this systematic review, we searched PubMed, Elsevier and Oxford Journal database for
English-language articles using search terms like diabetes mellitus, urology diseases such as
bladder dysfunction, urinary tract infections and urinary incontinence.
We included studies reporting the prevalence of overactive bladder, bladder dysfunctions
and lower urinary tract infections in patients with diabetes mellitus.
We extracted data for each pathology and relations between diabetes mellitus and bladder
complications.
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Results
Overactive bladder
According to 2010 IUGA/ICS joint report, overactive bladder syndrome (OAB) is defined
as ‘urinary urgency, usually accompanied by frequent nocturia, with or without urgency urinary
incontinence (UUI), in the absence of urinary tract infection or other obvious pathological
conditions’ [2]. The causes of diabetic bladder dysfunction include systemic inflammation,
diabetic angiopathy and neuropathy, all of which lead to chronic ischaemia of the bladder and
central nervous system. The neuropathy is primary peripheral and autonomic, affecting sensory
afferent pathways, causing the insidious onset of impaired bladder sensation, and ultimately
decreasing the detrusor contractility and bladder dysfunction. This may explain the association
between diabetic peripheral neuropathy and OAB [4]. Therefore, early discovery of the risk
factors for OAB progression in diabetic patients will boost the clinical management of these
two diseases.
A 2017 cross-sectional study involving 1025 diabetic individuals showed that those with
DM combined with overactive bladder (OAB) had the worst physical and mental quality of life
compared with other DM complications [3].
In a recent study made in China by Dongjuan Xu and al. they searched the prevalence of
overactive bladder (OAB) and quality of life in patients with type 2 diabetes mellitus. A total
of 1025 patients with type 2 diabetes were surveyed. Patients were grouped into no OAB, dry
OAB, and wet OAB groups according to the presence of OAB and urge incontinence. The
prevalence of OAB among patients with type 2 diabetes was 13.9% (with dry OAB, 6.1%; with
wet OAB, 7.8%), 2-fold greater than that in the general Chinese population. OAB symptoms
caused significant deterioration of the physical and mental aspects of quality of life. Compared
with dry OAB, wet OAB further decreased the mental aspect of quality of life.
Moreover, the effect sizes of the impacts of dry and damp OAB on quality of life were more
extensive to those of diabetic neuropathy or retinopathy, diabetes duration, or urinary tract
infection history [5].
Another study made by Zhu Y et al., published in 2019 showed that age, the duration of
diabetes and symptomatic diabetic peripheral neuropathy were found to be independent risk
factors for the progression of OAB. So, they recommend these three parameters to be monitored
in clinical practice when evaluating diabetic patients with OAB [6].
Bladder dysfunction
This has been traditionally described as a triad of decreased bladder sensation, increased
bladder compliance and capacity, and impaired detrusor contractility. Currently, diabetic
bladder dysfunction (DBD) refers to a diverse group of symptoms that include storage problems
such as the overactive bladder (OAB) with urgency incontinence, and voiding problems such
as impaired bladder emptying, urinary retention, and overflow incontinence.
Previously, it was not well known whether bladder dysfunction occurs without somatic
neuropathy, or whether bladder dysfunction is related to postural hypotension in patients with
diabetes. The results of a study made by Ryuji Sakakibara et al., showed that 17% of the diabetic
subjects had an abnormal post-void residual (PVR) volume. Notably, most patients were
unaware of their urinary retention [7]; This is in accordance with the findings of previous
reports [8]. The PVR volume was associated with the duration of diabetes, which is also
following the results of earlier reports [8]. Similarly, bladder neuropathy was correlated with
somatic and peri-arterial neuropathies. These findings indicate that bladder, somatic and peri-
arterial neuropathies might share the same pathological process, which could include
hyperglycaemia-induced molecular changes (i.e., the intra-neuronal polyol cascade) or
ischemia of the vasa nervosum [9]. However, bladder-specific pathophysiology might include
373
over-distension injury (due to polyuria) and changes in the urothelium [10]. Besides, there are
different nerve receptors in the bladder and vessels (muscarinic M3 receptors and alpha-1A/D
receptors are abundant in the lower urinary tract, while alpha-1B receptors are abundant in the
arterial wall) [10]. This finding is clinically relevant, and the regular measurement of the post-
void residual volume seems necessary in diabetes as bladder neuropathy might occur
insidiously in such patients.
Lower urinary tract infections

Another complication in diabetes mellitus is the possibility to contract common infections,
compared with patients without diabetes. Urinary tract infections (UTI) and mucosal
candidiasis are quite common and frequently more severe in people with diabetes than in the
general population. The risk is tightly related to diabetes duration and was lower in men
compared to women. Several studies have documented an association between the degree of
glycaemic control and the incidence or severity of infectious complications. Still, the
relationship between DM, hyperglycaemia, immune function and infections is complex, and
many issues remain unresolved.
Hyperglycaemia has been speculated to increase the genital infection and UTI risk because
it is known to weaken the immune system and cause several infections [11].
A survey reported UTI to be the most common microbial infection worldwide. Globally, it
was estimated at about 150 million people each year affected by this infection. Complications
of UTI in diabetic patient increases with age, poor metabolic control, various impairments in
the immune system and incomplete bladder emptying due to autonomic neuropathy [11]. The
most contributed pathogen of UTIs is Escherichia coli, in diabetic and non-diabetic patients,
others are Klebsiella pneumonia, Staphylococcus saprophyticus, Proteus mirabilis,
Enterococcus faecalis, Group-B Streptococcus, Pseudomonas aeruginosa, Candida spp, and
Staphylococcus aureus [12-14].
A meta-analysis made in 2016 by Dandan Li M. et al., showed that a class of oral antidiabetic
drugs named sodium-glucose co-transporter 2 inhibitors (SGLT2) could increase the incidence
of urinary tract infections (UTIs) for patients treated with SGLT2 inhibitors compared with
placebo [15]. Glycosuria was considered to be a more likely risk factor because SGLT2
inhibitors showed their anti-hyperglycaemic effects by inhibiting glucose reabsorption from the
kidney, and thus resulting in high glycosuria [15, 16].
Urinary incontinence
One of the complications caused by diabetes is urinary incontinence (UI) which is defined
by the International Continence Society as any involuntary leakage of urine. It is commonly
found in the elderly population, especially in women [17]. UI has widespread human and social
implications causing discomfort, shame, and loss of self-confidence. It affects the quality of life
but also has significant cost implications. Clinically, three forms of urinary incontinence have
been identified: stress incontinence, urge incontinence and overflow incontinence. The
pathogenesis of urinary incontinence is associated with diseases that cause frailty, muscle
weakness or atrophy and neurodegeneration. Several studies have demonstrated that diabetes
mellitus contributes to urinary incontinence [18]. It is well known that incontinence is more
common in patients with diabetes, but precise mechanisms by which type 2 diabetes may
contribute to its development or severity are not well defined [19, 20]. A likely ethiology for
incontinence in diabetic patients is microvascular damage that leads to diabetic neuropathy [21].
This may increase involuntary bladder contractions and decrease bladder sensation [21].
With severe neuropathy, the function of the detrusor muscle may be affected, resulting in
increased bladder volume and over distention [22, 23].
374
A study conducted by Arati Mahishale et al., showed that a long duration of diabetes and
altered glycaemic control could increase the risk of UI among type 2 diabetes mellitus patients
[24]. Brown et al., reported an increased risk of female UI inpatient with microvascular
complications and categorized UI as a microvascular complication [25].
Consistently, Danforth et al., suggested that microvascular changes in bladder and pelvic
floor muscles increased the risk of urgency UI. Diuresis related to hyperglycaemia increases
the risk of urgency UI and triggers the first pathologic changes in the bladder [26].
Consistently, Yu et al. reported that voiding difficulty was more frequent in patients with
DM than in those without DM. DM impairs detrusor innervation and contractility, and impaired
detrusor functions cause voiding distress and increased residual urine in the bladder [27].
Previous studies have shown that UI did not become symptomatic until the advanced stages
of DM as was also demonstrated by our research [27].
Female UI causes significant physical restriction and has psychological and economic
consequences as well. Recent evidence has revealed a significant association between female
UI and quality of life [27].
Conclusion
In conclusion, this review showed that bladder complications in diabetic patients are more
common than expected, and the symptoms affect their quality of life, so they should be
monitored cautiously in clinical practice.
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on bladder uroepithelial cells. Am J Physiol Regul Integr Comp Physiol 304: pp. R84-R93, 2013.
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Sponsor(s): Assoc Renal Metab & Nutrit Studies; Romanian Soc Diabet Nutr & Metab Dis; Natl Inst
Diabet, Nutr & Metab Dis; AstraZeneca Diabet; MSD Diabet; Novo Nordisk; Sanofi; Lilly Diabet; Berlin
Chemie Menarini; Accu Chek; Boehringer Ingelheim; Pfizer; Fresenius Med Care; Amgen; Servier; Vifor
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376
Does Obesity Cause Type 2 Diabetes?
MICIC Dragan D.1, POLOVINA Snezana2,3, MICIC Dusan D.4

1 Department of Medical Sciences, Serbian Academy of Sciences and Arts, Belgrade, (SERBIA)
2 Clinic for Endocrinology, Diabetes and Diseases of Metabolism, Clinical Center of Serbia, Belgrade, (SERBIA)
3 Faculty of Pharmacy, Novi Sad, (SERBIA)
4 Clinic for Emergency Surgery, Clinical Center of Serbia, Medical Faculty of Belgrade, University of Belgrade, (SERBIA)
Abstract
Obesity is believed to be a promoter of type 2 diabetes mellitus (Malone JI et al., 2019).

Obesity and fat accumulation predominantly in visceral depots are important risk factors for
the development of type 2 diabetes, dyslipidaemia, fatty liver disease, chronic subclinical
inflammation, hypertension, and cardiovascular disease (Bluher M, 2014). Obesity and diabetes
are global health problems and two epidemics of these diseases are escalating parallel and their
burdens are expected to increase in coming years (Bhupathiraju et al., 2017). Over the last
decade there has been a rapid escalation in the prevalence of obesity, one that parallels the
equally rapid increase in type 2 diabetes (Colagiuri S, 2010). Prevalence of obesity nearly
doubled worldwide since 1980 (Leitner S et al., 2017). In 2008 about 1 billion people in the
world are thought to be overweight or obese and at least 300 million people among them are
thought to be obese (Ford ES et al., 2008). According WHO data in 2016. more than 1.9 billion
adults, 18 year and older were overweight. Of these over 650 million were obese. 41 million
children under the age of 5 were overweight or obese in 2016, while over 340 million children
and adolescents aged 5-19 were overweight or obese in 2016 (WHO, 2016).
Overweight and obese persons are at risk of a number of medical conditions which can lead
to further morbidity and mortality. In a meta-analysis, it was demonstrated that elevated body
mass index (BMI) and waist circumference (WC) were significantly associated with type 2
diabetes in men and women (Guh DP et al., 2009). In concordance with the WHO, overweight
and obesity account for 44% of the diabetes cases. WHO estimates that, globally, 422 million
adults aged over 18 years were living with diabetes in 2014? The number of people with
diabetes has substantially increased between 1980 and 2014, rising from 108 million to current
numbers that are around four times higher (WHO, 2016; Boles A. et al., 2017.). Being
overweight or obese is strongly linked to diabetes. The prevalence of obesity-related diabetes
is expected to double to 300 million by 2025 (Leitner D et al., 2017). Data from International
Diabetes Federation (IDF) indicate that in 2015 more than 415 million persons in the world
have diabetes. It is expected that this number will increase to 642 million by 2040. Among the
countries, China and India have the largest numbers of the subjects with diabetes (109.6 million
and 69.2 million, respectively) (Unnikrishnan R. et al., 2017). Regions with the highest
prevalence rates of diabetes are the Pacific Islands and the Middle East, while the regions with
the lowest prevalence are South and Central America and Africa (IDF, 2015).
The increase in prevalence of diabetes is linked to economic development and the subsequent
changes in lifestyle that promote an obesogenic environment (Unnikrishnan R. et al., 2017).
There is relationship between the degree of excess weight and risk for getting diabetes – the
risk is increasing threefold with a body mass index (BMI) of 25.0-29.9 kg/m2 and 20-fold with
a BMI over 30 kg/m2 (Field AE et al., 2001). The term diabesity was coined by Sims and
colleagues in the 1970s, to highlight the close relationship between diabetes and obesity (Sims
EA et al., 1973, Haslam D, 2010.). Professor Zimmet suggested that the “Diabesity” epidemic
377
is likely to be the biggest epidemic in human history (Zimmet P, 2017 Increase in obesity in
developed countries began in the 1980s and accelerated from 1992 to 2002, and then in 2006
starts to slow, while on the contrary, the increase in obesity is likely to continue in developing
countries, where it is about two-thirds of the current world’s obese population (Verma S et al.,
2017). Obesity is frequently preceding and the most important factor in the increase of type 2
diabetes (Toplak H et al., 2016). The driving force behind the current global type 2 diabetes
epidemic is insulin resistance among overweight and obese persons (Mutie PM et al., 2017).
The health impact of diabesity include long-term complications, reduction in health-related
functioning, reduction of quality in life and reduced overall life expectancy (Farag YMF et al.,
2011). Individuals of certain ethnicities are more prone to develop type 2 diabetes. Obesity-
driven reductions in insulin sensitivity appears to be behind the dramatic increase in diabetes
prevalence among the Pacific Island populations, as well as among the Pima Indians in the U.S.,
where high levels of obesity and insulin resistance are associated with one of the highest
prevalence of type 2 diabetes in the world (Zimmet P, 1979; Knowler WC et al., 1990).
However, recently it was postulated that there are two subtypes of type 2 diabetes: one
characterized by marked obesity, insulin resistance and relatively preserved beta cell function
(among the Pima Indians) and the other characterized by leaner body mass and more severe
beta cell dysfunction (among the Asian Indians) (Narayan KM, 2016). Obesity is associated
with an increased risk of developing insulin resistance and consequent type 2 diabetes. It was
postulated that insulin resistance of type 2 diabetes occurs primarily in the muscles of lean
individuals predisposed to diabetes before they become obese. These insulin resistances are the
cause of the excessive fat accumulation associated with type diabetes and participates in the
ethiology of the hypolipidemia and excess fat accumulation that are characteristic for type 2
diabetes mellitus (Malone JI et al., 2019). Adipose tissue of obese persons releases increased
amounts of non-esterified fatty acids, glycerol, hormones and different proinflammatory
cytokines that are involved in the development of insulin resistance (Kahn SE et al., 2006).
Metabolic disorders like obesity, insulin resistance and diabetes mellitus all possess
inflammatory components, and there are lot of data that indicate the fundamental role of
inflammation in the initiation, development and progression of these metabolic diseases (Shim
K et al., 2020). Later on, development of beta cells failure lead to the appearance of
hyperglycaemia and full-blown diabetes mellitus. Depending on the postulated pathogenesis,
different therapeutic strategies are recommended (insulin sensitizers for obese type 2 diabetic
and insulin secretagogues for lean type 2 diabetics (Unnikrishnan R. et al., 2017). Since the
majority of people with type 2 diabetes are overweight or obese, weight reduction is seen as the
key therapeutic goal in the prevention and the management of type 2 diabetes (Leitner DR et
al., 2017). In conclusion, current epidemiological evidence indicates that obesity and diabetes
are related multifactorial, complex diseases and a large proportion of the cases are preventable
by changing the life styles of the patients (Bhupathiraju SN et al., 2016).
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379
The Amount of Weight Loss after Metabolic Surgery Matters
POLOVINA Snezana1,2, MICIC Dragan D.3, MICIC Dusan D.4

1 Clinic for Endocrinology, Diabetes and Diseases of Metabolism, Clinical Center of Serbia, Belgrade (SERBIA)
2 Faculty of Pharmacy, Novi Sad, (SERBIA)
3 Department of Medical Sciences, Serbian Academy of Sciences and Arts, Belgrade, (SERBIA)
4 Clinic for Emergency Surgery, Clinical Center of Serbia, Medical Faculty of Belgrade, University of Belgrade, (SERBIA)
Abstract
Obesity is a chronic disease with excessive fat accumulation in the body and is a leading
public health problem beginning in the 20th and continuing in the 21st century. Obesity is
associated with cardiopulmonary function disorders and with increased mortality rate due to
cardiovascular disease, but also with increased total mortality (1, 2). Obesity lovering of the
oxidative capacity of skeletal muscles and lead to structural and functional worsening of the
cardiovascular and pulmonary system (3).
In general, bariatric/metabolic surgery causes certain improvement in metabolic and
cardiorespiratory function. Roux-en-Y gastric bypass (RYGB) represents one of two the most
effective options for surgical treatment of obesity (4). The second one is laparoscopic sleeve
gastrectomy (LSG).
There are different parameters which might improve after RYGB, including:
pharmacotherapy for hypertension, dyslipidaemia, obstructive sleep apnoea, type 2 diabetes,
peak oxygen uptake, ventilatory efficacy and breathing reserve.
Percentage of weight loss (%WL) and excessive weight loss (%EWL) are similar between
two procedures five years after operation, but patients after LSG have more weight regain
(30%) after five years than patients after RYGB (10%). The targets in surgical treatment of
obesity are comorbidities. The remission rate of comorbidities are similar in both procedures
after 5 years of surgery (5). One investigation shown that resolution of comorbidities are
associated with higher %EWL at first year after surgery, but there was no correlation between
early weight loss, during fist month after operation and effect on comorbidity resolution (6).
The authors from India compared LSG and RYGB in term of diabetes remission and they
concluded that amount of weight loss was similar in both procedures and one year after LSG
%EWL was 87%, after RYGB was 97%, and remain about 10% less after three years. That was
sufficient to provoke diabetes remission (7). The other study demonstrated mini gastric bypass
(MGB) and RYGB showed greatest %EWL at 5 years and thus they are recommended for mild
T2DM. LSG is an effective in %EWL in first 2 years after surgery and has high remission rate
in mild T2DM. Authors recommended MGB and RYGB for patients with moderate and LSG
for mild T2DM (8).
The other comorbidity in obesity, obstructive sleep apnoea (OSAS) resolve in about 75%
patients with EWL more than 60% after bariatric surgery. Predictive factors for the persistent
OSAS were also preoperative hypertension, older patients with ≥50years of age and AHI ≥30/hr
(9).
Poor cardiorespiratory function is a powerful predictor of mortality in patients with obesity.
Previous studies demonstrated that physical function improved within a couple of weeks after
bariatric surgery, but cardiopulmonary system requires certain conditions to provides structural
and functional improvement after surgical treatment of obesity.
380
Cardiopulmonary functional capacity provides diagnostic and prognostic information

regarding CPET (Cardio Pulmonary Ergospirometry Treadmill) parameters (10, 11).
One study investigated possible association of the amount of weight loss and improvement
of CPET parameters. Patients who lost more than 18% of their initial weight 6 months after the
surgery had significantly better CPET parameters results.
None of the patients continued to use statins after intervention. There was a significant
decrease in VO2 peak after 6 months in patients who had more than 18% body weight reduction.
Ventilatory efficiency and breathing reserve were significantly better in patients with greater
weight reduction after RYGB (2).
The respiratory exchange ratio (RER) showing an increase in some studies and no change in
one study(12). After RYGB, in most studies, RER was higher six months after greater surgical
weight loss, in one study authors suggest that an 18% reduction of initial weight after surgery
is associated with significant improvement in cardiorespiratory parameters.
After greater loss of kilograms, patients achieved of higher intensity of activity in
comparison with patients with smoker amount of weight loss. An old study demonstrated that
weight loss of just 8 kg after six months, is associated with a significant decrease in left
ventricular mass (13). Another, older study demonstrated that cardiac function in morbidly
obese individuals improved with an average weight loss of 55 kg. Earlier investigation have
shown that peak VO2 representing the highest achieved oxygen consumption during the
cardiovascular stress test is the most often analysed parameter for cardiorespiratory fitness
assessment (14). One new study demonstrated that VO2 kinetics was altered by LSG, but it
improved within the 16 months of follow-up. Authors speculated that LSG improved peak work
capacity along with a lower maximal absolute aerobic capacity. That might be due to impaired
peripheral oxidative muscle metabolism and a loss of muscle mass independently of %EWL
(11).
Study conducted on patients six months after RYGB, using treadmill shown that a weight
loss of 18% of initial kilograms was associated with significant differences between groups
regarding relative oxygen consumption, suggesting improvement in aerobic capacity. Authors
speculated that improved aerobic capacity may be due to better efficiency in performing
activities rather than real improvements in cardiorespiratory or muscle function. Although VO2
peak (mL/min) as represented of absolute aerobic capacity was unchanged after bariatric
surgery, increase of VO2 peak per body weight (mL/min/kg) confirmed better physical
functioning and mobility(15).
Some studies demonstrated that major postoperative increases in physical activity were seen
after sleeve gastrectomy, and these findings were associated with greater weight loss and
improved quality of life (16). Study on patients after RYGB surgery shown that about 64% of
patients with WL more than 18% six month after operation, become physically active.
Physical activity was defined by having average daily of 1 h by self-report. Regarding peak
O2 pulse after surgery, results are different, from no change to significant amelioration of peak
O2 pulse after RYGB. Same study has shown that weight reduction of less than 18% is not
associated with any statistically significant improvement of ventilatory function (2). Some
authors suggests that improvement of physical functioning in patients after bariatric surgery
might be a mechanical phenomenon of consequential weight loss, more than an absolute
improvement of cardiopulmonary and muscle function (16).
Decrease in minute ventilation at peak exercise after RYGB might be explained by
mechanical facilitation in filling the lungs’ alveoli after sufficient weight loss (17). RYGB is
more potent procedure in hypertension remission than LSG, probably due to effects of
adipokines on structural and functional changes in the cardiovascular system after rearranging
in gastrointestinal system after RYGB (18).
381
The 18% of weight loss of initial body weight is enough to reduces number of
antihypertensive drugs after surgery and should lead to an improvement in all ventilatory
parameters including FEV1, breathing reserve, ventilatory efficiency, minute ventilation at
peak exercise and PETCO2 after 6 months (2).
Conclusion
In comparison with conservative treatment, metabolic surgery results in more extensive body
weight loss and better control of comorbidities. Remission rate of comorbidities depends of
amount of weight loss. Therefore, metabolic comorbidities require more %EWL for remission,
although 18% of reduction of initial body weight may be a threshold value for significantly
better aerobic capacity and other cardiorespiratory outcomes after bariatric surgery.
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© Filodiritto Editore – Proceedings

6th International Conference on

(Bucharest, Romania, 5-7 March 2020)

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First Edition February 2020

© Copyright 2020 Filodiritto Publisher

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