PharmaNutrition 16 (2021) 100266

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PharmaNutrition
journal homepage: www.elsevier.com/locate/phanu

Kefir improves blood parameters and reduces cardiovascular risks in

patients with metabolic syndrome
Angela Camila da Silva Ghizi a, Mirian de Almeida Silva a, Flávia Souza de Andrade Moraes a,
Cristiane Lyrio da Silva a, Denise Coutinho Endringer a, Rodrigo Scherer a, Dominik Lenz a,
Ewelyne Miranda de Lima a, Girlandia Alexandre Brasil a, June Ferreira Maia a,
Nazaré Souza Bissoli b, Tadeu Uggere de Andrade a, *
a
Pharmaceutical Sciences Graduate Program, University Vila Velha, Vila Velha, ES, Brazil
b
Department of Physiology, Health Center, Federal University of Espirito Santo, Vitória, ES, Brazil

A R T I C L E I N F O A B S T R A C T

Keywords: Background: Metabolic syndrome (MS) occurs when different metabolic and hemodynamic components change
Probiotic simultaneously, being one of the factors related to high mortality rates in patients with cardiovascular diseases.
Blood pressure This study investigated the effects of the probiotic kefir on anthropometric and physiological parameters in
Oxidized LDL cholesterol
human subjects with MS.
Fasting glucose
Lipids
Methods: forty-eight patients diagnosed with MS were assigned into two groups in this randomized, double-blind,
Metabolic diseases placebo-controlled clinical trial. Kefir group (KG) and control group (CG) drank kefir beverage and curd (1.6 mL/
kg for men or 1.9 mL/kg for women) for 12 weeks, respectively. Blood pressure, anthropometric, and
biochemical parameters (fasting glycemia, glycated hemoglobin (HA1c), total cholesterol (TC), high density li­
poprotein (HDLc), triglycerides (Tg), oxidized LDL cholesterol (oxLDL), high-sensitivity C-reactive protein (hs-
CRP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinophosphokinase (CPK), γ-Glu­
tamyl Transferase (γ-GT), urea nitrogen, urea, and creatinine were evaluated before and after treatment. The risk
of cardiovascular events for the next ten years was calculated through the Framingham Score method.
Results: Kefir intake decreased blood pressure, fasting glycemia, LDLc, non-HDLc, Tg, and oxLDL, and increased
HDLc levels in women. Kefir also reduced the risk of cardiovascular events for the next ten years, although
anthropometric parameters remained unchanged.
Conclusions: Kefir intake improved blood pressure, fasting glucose, and lipid levels, reducing oxLDL and the risk
of developing cardiovascular events in the next ten years. These results suggest regular kefir intake may have
positive effects on MS treatment.

1. Introduction three of five events: accumulation of abdominal fat, identified by

Metabolic syndrome (MS) is related to high mortality rates in pa­
tients with cardiovascular diseases [1,2]. Cardiovascular diseases are
the main cause of death worldwide, accounting for 17.7 million deaths
in 2015 and representing 45 % of all deaths caused by
non-communicable diseases [3].
Metabolic syndrome occurs when different metabolic and hemody­
namic components change simultaneously [1,2,4]. According to the
main regulatory agencies, MS is defined by the occurrence of at least
increased abdominal circumference, elevated triglycerides levels,
reduced HDL cholesterol levels, increased blood pressure, and elevated
fasting glucose levels [5,6].
The parameters involved in the development of MS are influenced by
genetic factors, sedentary lifestyle, eating habits, and aging, among
others [5,7]. However, insulin resistance and obesity, with accumula­
tion of abdominal fat, seem to play a major role in the development of
the disease [5,7].
MS treatment can be complex, as it may involve various

* Corresponding author at: Department of Pharmaceutical Sciences, Universidade Vila Velha, Av. Comissário José Dantas de Melo, n◦ 21, Boa Vista, Vila Velha, ES,
CEP 29102–920. Brazil.
E-mail address: tadeu.andrade@uvv.br (T.U. de Andrade).

https://doi.org/10.1016/j.phanu.2021.100266
Received 16 December 2020; Received in revised form 24 April 2021; Accepted 26 April 2021
Available online 29 April 2021
2213-4344/© 2021 Elsevier B.V. All rights reserved.
A.C.S. Ghizi et al. PharmaNutrition 16 (2021) 100266

pharmacological interventions, lifestyle changes, and even bariatric Velha University - UVV/ES, who was also responsible for the identifi­
surgery in cases of severe obesity [8,9]. cation of bottled beverages intended for volunteers. Double-blind refers
Thus, studies that investigate elements potentially associated with to the blinding of patients, researchers, data collectors, and outcome
conventional treatment, contributing to the improvement of MS symp­ evaluators.
toms, can be quite promising. In this context, probiotics have been used Participants were recruited from a university teaching institution.
as a therapeutic alternative to alleviate cardiovascular and metabolic Follow-up lasted eleven weeks, what classified the trial as long-term.
diseases [10–12]. The presence of MS parameters was confirmed through anthropo­
Kefir, for instance, a probiotic beverage known in several countries, metric analysis, blood pressure measurement, and biochemical param­
has been drawing the attention of the scientific community because of its eters evaluation, following the NCEP-ATP-III criteria [5]. The
beneficial properties [13]. It comprises an aggregate of bacteria – pre­ participants were then randomized into two groups, namely control
dominantly lactic and acetic acid bacteria – and yeasts, enclosed in a (CG) and kefir (KG) groups. After the 11-week period all analyses per­
polysaccharide matrix known as kefiran [14–16]. Several bioactive formed before the randomization process were repeated.
peptides with antihypertensive, antimicrobial, immunomodulatory, Drugs for the reduction of serum cholesterol, arterial hypertension,
opioid, and anti-oxidative functions have been identified in kefir and any other medication of continuous use were maintained during the
[17–20]. study.
Previous studies have shown that treatment with kefir improves This study was submitted to and approved by the Ethics Committee
certain MS parameters in experimental animals [21,22]. For instance, in Research with Human Beings of the Vila Velha University – UVV
mice treated with kefir had reduced triglycerides levels, glycemia, (Protocol # 1.025.083). All volunteers signed the free and informed
fasting insulin, body fat, and oxidative stress markers, in addition to consent form, agreeing to participate in the research. The study was
improved levels of anti- and pro- inflammatory cytokines [21]. registered in ClinicalTrials.gov (ID: NCT03649828).
According to recent clinical studies, the use of probiotic foods can
alleviate the severity of symptoms and affect anthropometric and
2.2. Participants and selection criteria
biochemical results of MS patients [23–25]. Body mass index, total
cholesterol, and low-density lipoprotein levels were significantly
Samples were calculated to detect a 12 % reduction in high-sensitive
reduced in MS patients using Bifidobacterium lactis, when compared with
C-reactive protein (hs-CRP) levels – considering a standard deviation of
control individuals. Moreover, a decrease in tumor necrosis factor-α
15 %, a 2-tailed of 0.5 and power of 80 % – in 24 individuals per group
(TNF-α) and interleukin-6 (IL-6), both pro-inflammatory cytokines, was
(48 total). Considering possible sample loss during the protocol, 70
observed in these patients [23].
subjects were invited to participate in the study and 63 agreed to do so.
In a recent review article, Torres et al. [26] discussed evidence that
The criteria adopted for the inclusion of volunteers in the study were
the composition of the intestinal microbiota modulates adipose tissue,
to be over 18 years of age, to be physically inactive, and to present MS,
body weight, and the prevalence of a low-grade inflammatory state,
characterized by alteration of at least three of five criteria used as in­
suggesting probiotics may improve MS by modulating gut microbiome
dicators of the disease: abdominal circumference ≥ 102 cm for men and
[26].
≥ 88 cm for women, HDL cholesterol ≤ 40 mg/dL for men and ≤ 50 mg/
In addition, our group demonstrated that kefir reduces blood pres­
dL for women, fasting blood glucose ≥ 100 mg/dL, systolic blood
sure by improving baroreflex [27], endothelial function [28], and car­
pressure (SBP) ≥ 130 mmHg or diastolic blood pressure (DBP) ≥ 85
diac function (by improving contractility), also reducing sympathetic
mmHg, and triglycerides (Tg) ≥ 150 mg/dL [32,5].
activity [29] and inhibiting the activity of the angiotensin converting
Pregnant and lactating women, plus individuals who consumed other
enzyme (ACE) [22] in spontaneously hypertensive rats. Recently,
probiotics and prebiotics, were excluded from the study. Volunteers who
Amorim et al. [19] identified 35 peptides with ACE inhibitory activity in
used drugs for dyslipidemias that interfered with intestinal metabolism
kefir, reaffirming its potential in reducing blood pressure.
such as ezetimibe and anion exchange resin, hormones of any kind,
Uncontrolled blood pressure has been associated with major car­
drugs for weight loss, and antioxidant supplements such as vitamin C or
diovascular events in MS patients, whereas type 2 diabetes has been
ω-3 were also excluded.
associated with an increased risk of macrovascular events such as
This clinical trial was therefore conducted with 48 volunteers of both
myocardial infarction and stroke [30]. Therefore, it is important to
genders, split into two groups (CG and KG) with 24 subjects each. Fifteen
decrease the blood pressure of these patients to reduce mortality and
volunteers were excluded from the study before the beginning of treat­
other negative clinical outcomes [31].
ment: three did not show up for the initial assessment, 11 did not meet
Human and animal studies have shown the biological effects of
the criteria for MS and one was under hormone replacement, as detailed
probiotic microorganisms on MS parameters and other cardiovascular
in Fig. 1.
risks; however, the effect of kefir and its impact on cardiovascular events
of MS patients has not been reported.
Therefore, in this study we investigated whether a daily consumption
of kefir beverage has beneficial effects on metabolic parameters and
reduces the cardiovascular risk in human individuals with MS. To do so,
we evaluated determinant components of MS before and after kefir
treatment, and assessed the cardiovascular risk through ultra-sensitive
C-reactive protein and Framingham score measurements.

2. Material and methods

2.1. Study design

This double-blind, placebo-controlled, parallel-blind, randomized,

controlled trial was developed using the automated randomization
system Random Allocation Software (version 1.0, May of 2004).
Randomization codes were under the responsibility of a professional
supervisor from the Laboratory of Technique and Dietetics of the Vila Fig. 1. Flowchart of volunteers participating in the research.

2
A.C.S. Ghizi et al.
2.3. Preparation of fermented milk with kefir grains
The kefir beverage (test drink) was produced from kefir grains,
kindly provided by Prof. Dr. Célia Lúcia L. F. Ferreira, Department of
Food Technology, Federal University of Viçosa - UFV, Minas Gerais.
Kefir (test drink) and curd (placebo) drinks were prepared in the
Laboratory of Dietary Technique of the Nutrition and Gastronomy
Course of UVV - ES, using pasteurized whole milk with 3% fat (Fiore®,
Santa Teresa - ES / Brazil). Kefir grains were added to the milk (5% w/v)
in a sanitized glass vessel and fermented at room temperature (~ 25 ◦ C)
for approximately 24 h. After fermentation, the filtrate was separated
from the grains with a plastic sieve and submitted to a second fermen­
tation process. This process consisted of further 24 h under refrigeration
at 10 ◦ C, to promote yeast growth [28] and resume the release of pep­
tides, besides conferring specific flavor and aroma to the drink. Organic
strawberries and organic demerara sugar were added to the fermented
product. Homemade curd prepared from starter culture in 3% fat milk,
flavored and sweetened identically to kefir, was used as a control drink.
The products were packed in individual disposable packages, labeled,
and kept under refrigeration until consumption by the volunteers. For
diabetic volunteers, a proportional amount of artificial sucralose
sweetener was used according to the manufacturer’s indication.
2.4. Treatment
Individuals from the KG group received probiotic milk fermented
with kefir grains and those from the CG group received curd drinks.
The volume of probiotics to be taken by the two groups was calcu­
lated based on studies by Reagan-Shaw et al. [33] and Rosa et al. [34].
The mean of the minimum and maximum recommendations, equivalent
to 1.6 mL/kg of body weight/day for men and 1.9 mL/kg of body
weight/day for women, was adopted as consumption volume.
The products were prepared daily and distributed to the volunteers
in their workplaces during office hours, in the morning, Monday through
Friday. The routine was followed daily until the end of treatment.
2.5. Assessment of body composition
Anthropometric measurements were assessed at the UVV Nutrition
Clinic before the beginning and at the end of the study. Body mass and
stature were measured using a digital platform scale (RAMUZA DP-300)
and a digital vertical anthropometer (CHORDER HM 210D),
respectively.
Body mass index (BMI) (kg/m2) was calculated according to the
formula:
BMI (kg/m2) =(Weight(kg))/([Stature] 2 (m))
To classify BMI, the ranges proposed by the WHO [35] were adopted.
Waist circumference (WC) was measured with a millimeter and in­
elastic tape measure. The cut-off points for risk assessment associated
with metabolic complications of obesity were classified according to the
NCEP-ATP III [5], with WC ≥ 102 cm and ≥ 88 cm being considered risk
values for men and women, respectively. Body composition was
analyzed by electrical bioimpedance (MaltronBody Fat Analyzer BF
906), as described by Lukaski et al. [36], with the percentage of lean
mass and body fat having been evaluated.
During the experiment the selection of food was at the discretion of
each volunteer.
2.6. Evaluation of blood pressure and biochemical parameters
Blood pressure (SBP and DBP) was measured according to the pro­
tocol recommended by the American Heart Association Council [37],
through an automatic measuring instrument (HEM-705CPINT, Omron,
OmronHealth Care, INC., Illinois-EUA). The average between two
3
PharmaNutrition 16 (2021) 100266
measurements was considered for analysis.
For biochemical analysis, fasting blood samples were collected by
traditional venipuncture to evaluate the following biochemical param­
eters: fasting glycemia, glycated hemoglobin (HA1c), total cholesterol
(TC), high density lipoprotein (HDLc), triglycerides (Tg), high-
sensitivity C-reactive protein (hs-CRP), aspartate aminotransferase
(AST), alanine aminotransferase (ALT), creatinophosphokinase (CPK),
γ-glutamyl transferase (γ-GT), urea nitrogen, urea, and creatinine. The
samples were sent to the Tommasi Clinical Analysis Laboratory (Vila
Velha – ES) for further analysis. Low density lipoprotein (LDLc) and non-
HDL cholesterol levels were calculated from the results of the
biochemical analysis.
2.7. Cardiometabolic risk evaluation
The relative and absolute risk for the development of coronary heart
disease in the next decade of life was assessed using the Framinghan risk
score model for coronary heart disease. This score considers age, sex,
total cholesterol, HDL cholesterol, smoking, systolic blood pressure, and
presence of diabetes mellitus, providing a continuous measure of the
CVD risk in 10 years [38].
All data from anthropometry, dietary intake, blood biochemical
analysis, SBP and DBP, and Framinghan scores were collected before
and after the period of curd or kefir consumption.
2.8. Oxidized LDL levels
Oxidized LDL was determined on blood plasma samples through
ELISA kit (Elabscience, Wuhan, China, E-El-M0066) according to the
manufacturer’s instructions. Readings were performed on a microplate
reader (Filter Max F3/F5 Multi-Mode Microplate Readers) at 450 nm
and the results expressed as ng/mL.
2.9. Statistical analysis
The data were compiled into Microsoft Excel spreadsheets and
expressed as mean ± standard deviation (SD). The database was
analyzed using the statistical program SPSS 11.5 (Statistical Package
Social Science version 11.5). Data normality was tested with the Sha­
piro–Wilk test. Simple relative frequencies were performed for qualita­
tive variables within each group. Quantitative data were analyzed
through descriptive statistics and the differences determined using
paired (in the same group) and unpaired (between groups) Student’s t-
test with 95 % confidence interval. Differences between means were
considered statistically significant when p <0.05.
3. Results
3.1. Baseline characteristics of the population studied
Of the 48 volunteers who participated in this clinical study, 23 %
were men and 77 % women. Table 2 shows the demographic parameters
and anthropometric measurements of the participants before and after
administration of kefir or curd. Volunteers had high WC and BMI, as well
as a high percentage of fat to begin with. Kefir treatment did not induce
alterations in any anthropometric parameters analyzed here (Table 1).
3.2. Evaluation of blood pressure and biochemical parameters
Before the beginning of treatment, individuals from both groups (CG
and KG) had high SBP and DBP. Kefir administration decreased SBP and
DBP, with DBP falling below the level that characterizes arterial hy­
pertension (Table 2).
Biochemical parameters were evaluated before and after treatment.
Neither group had individuals with diabetes mellitus in the beginning of
the study. Kefir consumption decreased fasting glycemic levels even
A.C.S. Ghizi et al. PharmaNutrition 16 (2021) 100266

Table 1 were reduced by the treatment (Table 2).

Demographic parameters and anthropometric measurements of volunteers, for To assess liver function, the enzymes AST, ALT, and γ-GT were
both groups, before and after administration of curd (CG) or kefir (KG). analyzed. Renal function was assessed through the measurement of
CG KG urea, creatinine, and urea nitrogen levels. None of these parameters
Before After Before After
were altered before interventions, remaining unchanged after the use of
kefir or curd.
Age (years) 42 ± 14 44 ± 10
Gender (Male/ 6/18 5/19
Female) 3.3. Framingham score
BMI (kg/m2) 31.4 ± 4.98 31.4 ± 32 ± 4.37 32 ± 4.5
5.13 The data shown in Table 3 indicate that kefir treatment reduced the
Lean Mass (%) 49.4 ± 13.7 50.2 ± 56.5 ± 9.1 55.4 ± 8.9
risk of cardiovascular events for the next ten years, calculated through
14.1
Fat Mass (%) 36.8 ± 10 36.1 ± 39.2 ± 6.9 39.9 ± 7.1 the Framingham Score.
10.7
Women’s WC (cm) 94.6 ± 11.1 94.9 ± 94 ± 10.1 95 ± 9.8 3.4. Levels of oxidized LDL
10.1
Men’s WC (cm) 108.5 ± 10.3 ± 108.3 ± 108.3 ±
14.5 13.8 13.4 11.5 Fig. 2 shows oxidized LDL cholesterol levels in both control and kefir
groups before (CGB and KGB) and after treatment (CGA and KGA). Kefir
Values expressed as mean ± Standard Deviation (S.D.). BMI: body mass index;
treatment significantly reduced the levels of oxidized LDL cholesterol
WC: waist circumference. Gender (Male/Female) = number of men/women in
(KGB: 12.9 ± 1.6; KGA: 9.8 ± 1.3; p < 0.05).
each group.

4. Discussion
Table 2
Blood pressure values and blood biochemical parameters of volunteers, for both The main finding of the present study was that kefir treatment
groups, before and after administration of curd or kefir. reduced the cardiovascular risk, according to Framingham scores and
CG KG oxidized LDL – a marker of atherosclerosis – in individuals with meta­
bolic syndrome, while hs-CRP levels remained unchanged. In addition,
Before After Before After
blood pressure (SBP and DBP), triglyceride levels, and fasting glucose
SBP (mmHg) 140 ± 15 137 ± 11 139 ± 12 130 ±
were reduced after kefir consumption, with increased HDL-c levels
9*#§
DBP (mmHg) 93 ± 10 90 ± 6 90 ± 4 83 ± 5*#§
having also been detected in women following treatment. The effect of
Fasting glycemia (mg/ 87 ± 20 85 ± 18 95 ± 9 83 ± 8* chronic treatment with kefir was not affected by the participants’ diet
dL) and physical activity habits.
HbA1c (%) 5.5 ± 0.8 5.3 ± 0.6 5.5 ± 0.5 5.4 ± 0.5 This is the first randomized clinical trial showing that kefir con­
Total cholesterol (CT) 180 ± 48 178 ± 44 193 ± 27 189 ± 16
sumption induces clinical benefits in individuals with metabolic syn­
(mg/dL)
LDLc (mg/dL) 136 ± 36 131 ± 35 133 ± 16 123 ± drome. Bellikci-Koyu et al. [39] conducted an elegant, randomized trial
11*#§ with MS volunteers receiving kefir, designed to address changes in the
HDLc man (mg/dL) 40 ± 3 41 ± 3 38 ± 3 42 ± 5 relative abundance of microorganisms in gut microbiota. They used a
HDLc woman (mg/dL) 43 ± 4 43 ± 5 44 ± 5 48 ± 3*#§
commercial culture containing seven species of microorganisms to
Non-HDLc (mg/dL) 170 ± 42 159 ± 47 163 ± 23 147 ±
19*#§
ferment whole milk. However, our kefir beverage is completely
Triglycerides (Tg) (mg/ 191 ± 44 198 ± 54 196 ± 67 147 ± different, as it was obtained through the fermentation of cow milk by
dL) 44*#§ kefir grains. These grains are complex systems of symbiotic living bac­
hs-CRP (mg/L) 2.27 ± 1.7 2.4 ± 1.2 2.33 ± 1.1 2.16 ± teria and yeasts, comprising more than 30 species of bacteria and more
1.35
than 12 species of yeasts and fungi [revised by 13]. Therefore, consid­
CPK (U/L) 138.6 ± 135.3 ± 126.3 ± 124.6 ±
63.2 56.1 47.9 61 ering that the effects of kefir beverage are due not only to its probiotic
γ-GT (U/L) 26.9 ± 12 27.9 ± 27.3 ± 14 28.3 ± action, but also to the bioactive compounds synthesized during the
10.9 13.8 fermentation process, the culture used to ferment the milk is a key
Urea (mg/dL) 27.7 ± 9.1 27.1 ± 6.3 27.2 ± 8.4 27.6 ± 6.8 component for the biological action of this functional drink.
Urea N (mg/dL) 12.9 ± 4.4 12.7 ± 3 12.7 ± 4 12.9 ± 3.2
Creatinine (mg/dL) 0.75 ± 0.72 ± 0.79 ± 0.1 0.8 ± 0.16
This could explain, at least in part, why we found more clinical
0.18 0.17 benefits than the study by Bellikci-Koyu et al. [39]. We observed sig­
AST (U/L) 26.5 ± 6.5 26 ± 7.6 27.1 ± 26.9 ± 12 nificant improvements in a number of clinical parameters, such as blood
11.8 pressure, blood lipid profile, fasting glucose, and oxidized LDL, which
ALT (U/L) 26.3 ± 25.3 ± 28 ± 18.6 26.7 ±
reflected in the reduction of cardiovascular risk, evaluated by the Fra­
17.8 12.1 14.9
mingham score.
Values expressed as mean ± S.D. *p < 0.05 vs KG before; #p < 0.05 vs CG before; Even though our kefir treatment reduced the cardiovascular risk
§
p < 0.05 vc CG after. SBP, systolic blood pressure; DBP, diastolic blood pressure,
HbA1c: glycated hemoglobin, LDLc: low-density lipoprotein cholesterol, HDLc:
Table 3
high density lipoprotein cholesterol; Non-HDLc: Non high density lipoprotein
Risk of cardiovascular events in the next decade determined by the calculation of
cholesterol; hs-CRP: high-sensitivity C-reactive protein; CPK: Crea­
the Framingham Score, for both groups, before and after administration of curd
tinophosphokinase; γ-GT: γ-Glutamyl Transferase; AST: Aspartate Aminotrans­
or kefir.
ferase; ALT: Alanine Aminotransferase.
CG KG

further, although HbA1c remained unchanged after treatment. No dif­ Before After Before After
ferences were observed in total cholesterol levels and hs-CRP between Framingham score 6.75 ± 4.4 6.25 ± 4.6 6.67 ± 3.0 5.5 ± 2.7
# §

groups. We detected an increase in HDLc levels in women after kefir

Values are expressed as the mean ± S.D.
treatment, whereas in men such levels remained unchanged. Kefir also
*p < 0.05 vs KG before.
decreased serum triglyceride and non-HDL cholesterol levels, demon­ #
p < 0.05 vs CG before.
strating that atherogenic lipoproteins (LDL, VLDL, and their remnants) §
p < 0.05 vc CG after.

4
A.C.S. Ghizi et al.
Fig. 2. Effect of kefir treatment on oxLDL. CGB: control group before treat­
ment; CGA: control group after treatment; KGB: kefir group before treatment;
KGA: kefir group after treatment. Values are presented as mean + SD, for n = 24
subjects per group, after eleven weeks of treatment. *p < 0.05 vs KG before; #p
< 0.05 vs CG before; §p < 0.05 vc CG after.
according to the Framingham score, the level of hs-CRP – a marker of
cardiovascular risk – remained unaffected. That can be explained, at
least in part, by the volunteers having considerable adipose tissue
accumulation. A high level of hs-CRP is associated with high cardio­
vascular risk [40,41], as it contributes to endothelial lesion by mediating
the formation, destabilization, and rupture of atherosclerotic plaques
[42]. The peptides present in kefir can reduce adipose tissue, improving
obesity via inhibition of lipogenesis, modulation of oxidative damage,
and stimulation of lipid oxidation [43]. Nevertheless, here we found that
both BMI and WC remained elevated following kefir treatment. Ac­
cording to Pearson et al. [40], hs-CRP levels are proportional to body
weight. We must point out that in this study the volunteers did not
receive nutritional guidance for elimination of excess body weight. Be­
sides, although the follow-up of this clinical trial was classified as
long-term (eleven weeks of treatment), it was not sufficiently long for a
significant reduction in body weight to take place. It would be necessary
to reduce body mass by ~ 20 kg, considering a maximum BMI of 24.9
kg/m2 as eutrophic, and a reduction of WC by ~ 6 cm for men and ~ 7
cm for women, considering the composition of risk factors for MS ac­
cording to the WHO [34] and NCEP-ATP III [5].
According to the NCEP-ATP III [5], MS is characterized by the as­
sociation of at least three of five risk factors, namely hypertension,
increased WC, reduced HDL-cholesterol, increased triglycerides, and
fasting glucose. Among these factors, all but elevated glycemia were
observed in the participants from both groups (CG and KG), confirming
our sample was indeed composed by MS individuals. This is the first time
a study demonstrated biochemical, cardiovascular, and antioxidant
beneficial effects after the consumption of kefir beverage by humans.
The benefits of kefir metabolites on cardiovascular parameters were
previously reported by Brasil et al. [22], who demonstrated that the
treatment of spontaneously hypertensive rats (SHR) with the soluble
fraction of kefir reduced mean arterial pressure, heart rate, and cardiac
hypertrophy, while improving baroreflex.
Other studies using experimental animals have addressed the effects
of kefir in restoring vascular function and decreasing mean arterial
pressure levels in hypertensive rats [27,28]. In the study by Friques et al.
[28] using hypertensive rats, kefir attenuated endothelial dysfunction in
large vessels, decreased the production of intravascular ROS, restored
vascular architecture, and, consequently, reduced blood pressure levels.
The beneficial effects of kefir can result from the release of peptides,
lipids, and other substances that may improve health [19,44]. In a recent
study, Amorim and co-workers [19] identified peptides with ACE
inhibitory activity, having shown kefir’s antihypertensive capacity in a
two-kidney one-clip model of secondary hypertension that depends on
RAS activation. This effect can explain, at least in part, the positive
5
PharmaNutrition 16 (2021) 100266
cardiovascular effects of kefir.
In vivo and in vitro studies have shown that kefir has promising effects
on treatment or prevention of cardiovascular and metabolic diseases
[16,22,45,46]. According to evidence presented by He and Shi [47], the
modulation of the intestinal microbiota induced by the use of probiotic
foods contributes to improve changes in the metabolic and hemody­
namic parameters that characterize the development of MS. The mi­
croorganisms present in probiotic products are beneficial because they
colonize the intestine, acting as a barrier against pathogens that prevent
the absorption of nutrients, and also by producing antimicrobial agents
and short chain fatty acids.
The participants of this study had high BMI, in addition to the other
components that characterize MS, and were classified as having class I
obesity [34]. Obesity is a multifactorial disease that involves genetic
predisposition and environmental factors, such as physical inactivity
and the intake of foods with high energy density, being strongly asso­
ciated with the development of MS [48,49]. MS was in fact detected in
the volunteers of this study, being associated to the high percentage of
body fat, with accumulation of adipose tissue in the abdominal region.
Here we observed that kefir did not affect BMI, lean mass, fat mass, and
WC.
Our results corroborate previous ones reported by Rosa et al., [21],
who observed that administration of kefir for 10 weeks to rats with MS
did not influence body weight, despite having reduced triglycerides,
fasting glucose, and the products of lipid oxidation. In addition, a pre­
vious study conducted by Bellikci-Koyu et al. [39] in a group of people
with MS reported a decrease on proinflammatory cytokines (TNF-α and
IF-γ), fasting insulin, HOMA-IR, and blood pressure. However, they did
not observe changes on gut microbiome composition, indicating that
fermentation products, e.g., peptides, could be responsible for the
beneficial effects attributed to kefir.
We can infer that kefir attenuated the deleterious effects of MS
through the control of oxidative stress and inflammatory/anti-
inflammatory cytokines. Kim et al. [50], while assessing the effects of
Lactobacillus kefiri DH5 (L. kefiri) isolated from kefir in an animal model
of obesity induced by high-fat diet (HFD), observed lower triglycerides
and LDL-cholesterol plasma levels, corroborating our findings in
humans. L. kefiri administration also alleviates the gut microbiota dys­
biosis induced by HFD.
No renal, cardiac, or liver damage was observed in individuals who
received kefir. Mert et al. [51] already showed that kefir does not affect
liver enzymes, such as AST and ALT, and heart enzymes, such as CPK.
Also, infarcted animals treated with kefir had lower levels of these en­
zymes when compared with control infarcted animals [51], demon­
strating kefir can play a protective role.
However, the Framingham score that predicts an individual’s risk of
developing a major cardiovascular event in the next ten years, such as
myocardial infarction or cardiovascular death, was decreased in the KG
group. The Framingham score is related to variables such as gender, age,
SBP and DBP, LDLc and HDLc, diabetes mellitus, and tobacco use. In this
study, LDLc levels, SBP, and DBP were significantly improved by kefir
treatment, explaining the risk reduction observed according to the
parameter analyzed here.
Another important finding was that oxLDL levels were low in KG
individuals, what probably occurred due to antioxidant effects induced
by the use of kefir [18,52]. Oxidized LDL is considered a pro-atherogenic
factor when present in high levels, because of its ability to trigger a
complex inflammatory process, including proliferation of macrophages
and adhesion molecules [53]. LDL cholesterol retained in the sub­
endothelial layer is susceptible to reactive oxygen species (ROS). Lipo­
proteins undergo oxidation and act as triggers for the expression of
adhesion molecules and chemokine secretion from endothelial cells
[53].
Other studies have shown that antioxidant products have the po­
tential to be used in the treatment and prevention of cardiovascular
diseases, such as atherosclerosis [54,55]. A study by Liu et al. [52]
A.C.S. Ghizi et al.
evaluated the antimutagenic effect and the antioxidant activity of cow
and soy milk fermented with kefir, revealing that both products had high
free radical sequestering capacity and high reduction power, which
increased with longer fermentation periods. In addition, the aforemen­
tioned study showed that products fermented with kefir inhibited lipid
peroxidation and had high antimutagenic capacity [52]. These findings
indicate that probiotic foods based on kefir have antioxidant activity and
can act as protective agents against the oxidation of LDL cholesterol.
Taken together, our results show that the probiotic kefir improved
important parameters used as indicators of MS and reduced the risk of
developing cardiovascular disease. The use of this probiotic reduced
blood pressure (SBP and DBP), triglycerides, and non-HDL cholesterol
levels. In addition, women had an increase in HDL levels. The treatment
also reduced oxLDL levels, an important indicator of ischemic cardio­
vascular disease, and the risk for the development of cardiovascular
diseases assessed by the Framingham Risk Score method. These findings
indicate that the consumption of kefir may be a therapeutic option
against the development of metabolic and cardiovascular diseases.
However, further studies are needed to better elucidate the pathways
and effects of kefir use.
5. Limitations
In the present study the volunteers did not receive any diet or ex­
ercise guidance to avoid interference with the results. However,
considering that food behavior and exercise practice can influence the
composition of the microbiota, the lack of guidance brings a limitation
to our study: as we did not evaluate diet and exercise behavior during
the experiment, we cannot guarantee the two groups had the same
profile.
Declaration of Competing Interest
The authors declare no conflict of interest.
Acknowledgments
This work was supported by the Coordenação de Aperfeiçoamento de
Pessoal de nível Superior Brasil [CAPES; finance code: 001]. TUA re­
ceives a fellowship from Conselho Nacional de Desenvolvimento Cien­
tífico e Tecnológico [CNPq; grant number: 311925/2018-9] and from
Fundação de Amparo a Pesquisa do Estado do Espírito Santo [FAPES;
grant number: 522/2018]. NSB receives serach features from Fundação
de Amparo a Pesquisa do Estado do Espírito Santo [FAPES; grant num­
ber: 591/2018].
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