Gynecological Endocrinology

ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/igye20

Proteomic and bioinformatic analysis of human

endometrium from polycystic ovarian syndrome
with and without insulin resistance

Xin Yang, Wang Xiaoping, Ding Nan, Zhang Jian, Li Xiaofeng, Yuan Liwei,
Mengni Zhao & Fang Wang

To cite this article: Xin Yang, Wang Xiaoping, Ding Nan, Zhang Jian, Li Xiaofeng, Yuan
Liwei, Mengni Zhao & Fang Wang (2023): Proteomic and bioinformatic analysis of human
endometrium from polycystic ovarian syndrome with and without insulin resistance,
Gynecological Endocrinology, DOI: 10.1080/09513590.2023.2173948

To link to this article: https://doi.org/10.1080/09513590.2023.2173948

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Gynecological Endocrinology
https://doi.org/10.1080/09513590.2023.2173948

Research Article

Proteomic and bioinformatic analysis of human endometrium from polycystic

ovarian syndrome with and without insulin resistance
Xin Yang†, Wang Xiaoping†, Ding Nan, Zhang Jian, Li Xiaofeng, Yuan Liwei, Mengni Zhao and Fang Wang
Reproductive Medicine Center, Second Hospital of Lanzhou University, Lanzhou, China

ABSTRACT ARTICLE HISTORY

Objective: The aim of this study was to investigate the endometrial proteomic profiles of patients with Received 3 July 2022
polycystic ovary syndrome (PCOS) with and without insulin resistance (IR). Method of Study: We collected Revised 9 January 2023
40 endometrial samples, including PCOS-IR (n = 21), PCOS-non-IR (n = 12), and control (n = 7). Accepted 24 January 2023
Published online 8
Data-independent acquisition (DIA)-based proteomics method is used to identify the expressed proteins
February 2023
among the three groups. The correlation between pregnancy outcomes and identified proteins was
analyzed by Lasso regression. Results: A total of 5331 proteins were identified, while 275 proteins were KEYWORDS
differentially expressed in the PCOS vs. control group and 215 proteins were differentially expressed in Polycystic ovary syndrome;
the PCOS-IR vs. PCOS-non-IR group. Platelet degranulation, neutrophil degranulation, and very long-chain insulin resistance;
fatty acid catabolic processes have been found to play important roles in the endometrium of patients endometrium; pregnancy;
proteomics
with PCOS-IR. Lasso regression analysis found that ACTR1A, TSC22D2, CKB, ABRAXAS2, and TAGLN2 were
associated with miscarriage in patients with PCOS. ACTR1A and CKB were higher in the PCOS-IR group
and were positively correlated with HOMA-IR (p < .05). Conclusion: In this study, a panel of proteins was
found to be differently expressed in the endometrium. ACTR1A and CKB may be considered as PCOS-IR
candidate biomarkers.

Introduction Currently, it is widely believed that the important cause of endo-

Polycystic ovary syndrome (PCOS) is one of the most common
endocrine and metabolic disorders that affects between 8% and
13% among women of reproductive age [1]. The characteristics
of PCOS including hyperandrogenism, ovulatory dysfunction, and
polycystic ovaries, is often accompanied by insulin resistance
(IR) [2,3].
IR is one of the important metabolic signatures of PCOS,
appearing in multiple tissues in patients with PCOS, and endo-
metrial IR was previously strongly associated with fertility disor-
ders. Insulin stimulates ovarian theca cells to produce and secrete
androgens by reducing the sex hormone binding protein (SHBG)
in the liver [4]. IR decreases in nitric oxide (NO) and increases
in endothelin-1 (ET-1) in arterial endothelial cells, increases the
risk of cardiovascular metabolic diseases. IR increases the likeli-
hood of developing type II diabetes [5]. Insulin also inhibits
apoptosis and promotes cell proliferation, inducing endometrial
hyperplasia and promoting the growth of endometrial cancer cells,
increasing the long-term risk of endometrial cancer in patients
with PCOS [6]. However, the mechanism of long-term endometrial
cancer in patients with PCOS has not been fully elucidated.
Fertility is a major requirement for people with PCOS [7]. It
has been shown that even when age, BMI, and embryo quality
are controlled, patients with PCOS-IR have lower implantation
and clinical pregnancy rates than non-IR patients, suggesting that
local IR in the endometrium of patients with PCOS may be an
important factor in the decrease in endometrial receptivity [8].
metrial IR in patients with PCOS is the signal transduction dis-
order after the binding of insulin to its receptor, which is mainly
manifested by the inhibition of endometrial insulin receptor sub-
strate, abnormal phosphorylation of PI3K, inactivation of Akt,
expression, and translocation of GLUTs. It was found that down-
regulation of GLUT4 was associated with decreased FoxO1 func-
tion, an important biomarker for evaluating endometrial receptivity.
Therefore, it has been speculated that one of the causes of reduced
endometrial receptivity in patients with PCOS is endometrial local
IR. However, more evidence is needed to clarify the mechanism
by which local IR affects the expression of tolerance-related mol-
ecules [9]. Thus, the present study conducted a proteomics-based
approach to identify and select novel proteins associated in the
endometrial of patients with PCOS-IR.
Materials and methods
Study subjects and sample collection
A total of 33 patients with PCOS aged from 21 to 40 years and
meeting Rotterdam diagnostic criteria were recruited at Second
Hospital of Lanzhou University from September 2019 to
September 2020 and then divided into the PCOS-IR (n = 21)
and PCOS-non-IR (n = 12) groups according to their homeostasis
model assessment of insulin resistance (HOMA-IR). (HOMA-IR
was calculated using the equation: fasting plasma glucose
(mmol/L) × fasting insulin (mU/L)/22.5, and HOMA-IR ≥ 2.6

CONTACT Fang Wang ery_fwang@lzu.edu.cn Reproductive Medicine Center, Second Hospital of Lanzhou University, Lanzhou 730000, China.
†
Xin Yang and Xiaoping Wang contributed equally to this research.
Supplemental data for this article can be accessed online at https://doi.org/10.1080/09513590.2023.2173948.
© 2023 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group
This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits
unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
2 X. YANG ET AL.

was considered to be IR [10]). The Rotterdam criteria met the DAVID and the results were visualized with the software package
following 2–3 items: (1) Oligo- and/or anovulation; (2) Clinical ggplot2 in R. A p value < .05 was considered significant, and
and/or biochemical signs of hyperandrogenism; (3) Polycystic enriched GO terms or pathways were sorted by p values.
ovaries. At the same time, women were excluded from the study The tool Venn calculator can generate counts and detailed
if they had any of the following conditions: hypothyroidism, elements for each non-empty intersection for datasets with an
hyperprolactinemia, adrenal disease, hypertension, diabetes, hor- arbitrary number of groups. We plot the Venn diagram using
mone medication and the drugs affecting glucose metabolism PCOS vs. control, PCOS-IR vs. PCOS-non-IR differential pro-
in the last 3 months, pregnancy, and lactation. The control group teins, and then perform PCA with the ggprepel package R.
(n = 7) included women with successful pregnancies due to male Lasso regression analysis was performed to screen the key
factors and a routine ultrasound scan revealed normal ovarian proteins affecting the pregnancy outcomes in PCOS patients by
morphology and they had regular menstrual cycles. Demographic R (http://www.R-project.org) and Empower Stats (http://www.
characteristics and clinical biochemical parameters of the par- empowerstats.com, X&Y Solutions, Inc., Boston, MA). Key pro-
ticipants were collected. Demographic characteristics include age teins were found from raw data and differences in expression
and BMI recorded from outpatient medical records. Clinical levels between groups were analyzed.
biochemical indexes included basal testosterone (T), basal lutein-
izing hormone (LH), and basal follicle-stimulating hormone
(FSH), fasting blood glucose (FBG), fasting insulin (FINS), cho- Statistical analysis
lesterol (CHO), triglyceride (TG), high-density lipoprotein cho-
Normally distributed continuous data are presented as the
lesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C),
means ± SE and were compared using the Student’s t test, while
and thyroid stimulating hormone (TSH), which were measured
non-parametric data were presented as the median (interquartile
at 2 to 5 days of menstruation by the Laboratory of Lanzhou
range) and were compared using the Mann–Whitney U test,
University Second Hospital. We then followed up on the preg-
categorical variables using the chi-square test or Fisher exact
nancy outcomes after the patient’s hospital visit, including live
test when appropriate. A two-sided p value < .05 was considered
births and miscarriages prior to 24 weeks. We recorded the time
statistically significant for all tests.
of endometrial collection and the time of the last menstrual
period of pregnancy, in months, and calculated the interval
between the time of endometrial collection and the time of final
Results
pregnancy. Informal permission was signed by all participants
prior to sample collection. The study was approved by the local Subject clinical characteristics
ethics committee of Lanzhou University Second Hospital. All
subjects gave informed consent prior to participation (Ethical Clinical and biochemical parameters for all subjects used in the
Approval Number: 2017 A-057). proteomic analysis are presented in Table 1. BMI, LH, T, FBG,
The endometrial samples were obtained using a pipelle endo- FINS, HOMA-IR, TG, and the interval between the time of
metrial aspirator at the time of hysteroscopy when the endometrium endometrial collection and the time of final pregnancy were
was in the proliferative phase (histological examination ensured statistically different among the three groups, and HDL-C and
that samples were taken during the proliferative phase) and during live birth rates were much lower in the PCOS-IR group than
the month in which the blood samples were obtained, then rinsed in the PCOS-non-IR group and the control group (p < .05). There
the surface with phosphate-buffered saline (PBS) to clean the excess were no statistical differences in age, FSH, CHO, LDL-C, and
blood and stored at –80 °C for proteomic techniques used. TSH between the three groups (p > .05).
Sample preparation and fractionation, data dependent acqui-
sition (DDA) mass spectrometry assay, mass spectrometry data
analysis, and database search were completed by Shanghai Table 1. Baseline clinical and biochemical parameters in PCOS-IR, PCOS-non-IR,
and control groups.
Applied Protein Technology (Supplementary document 1 provides
detailed procedures). PCOS-non-IR PCOS-IR
Control (n = 7) (n = 12) (n = 21) p value
Age (year) 27.00 ± 2.94 26.83 ± 2.533 25.24 ± 3.45 .246
BMI (kg/m2) 21.39 ± 2.81 20.80 ± 2.42 26.10 ± 3.00 <.001
Bioinformatic analysis FSH (mIU/mL) 5.67 ± 0.83 6.78 ± 0.96 6.42 ± 1.82 .295
LH (mIU/mL) 5.34 ± 0.63 10.93 ± 5.37 10.45 ± 5.33 .045
The data were normalized using the Limma package of the R T (ng/dL) 24.69 ± 11.36 38.8 ± 10.60 46.93 ± 17.65 .006
language. Differentially expressed proteins (DEPs) between PCOS FBG (mmol/L) 4.54 ± 0.41 4.94 ± 0.55 5.10 ± 0.59 .001
and control, PCOS-IR and PCOS-non-IR were identified as that FINS (mIU/mL) 8.97 ± 3.90 8.76 ± 11.77 24.68 ± 11.77 <.001
HOMA-IR 1.80 ± 0.80 1.94 ± 0.52 5.90 ± 2.94 <.001
with |log2 fold change (log2 FC) |>1 as the cutoff value. Proteins CHO (mmol/L) 3.65 ± 0.81 3.86 ± 0.75 4.17 ± 0.74 .279
with p < .05 calculated by the Bayes test were included in the TG (mmol/L) 0.99 ± 0.43 1.03 ± 0.63 1.85 ± 0.95 .013
DEPs list. The DEPs analysis was performed in R 4.1.0. The HDL-C (mmol/L) 1.45 ± 0.43 1.47 ± 0.24 1.14 ± 0.25 .007
ggpubr package was used to map volcanoes to visually show the LDL-C (mmol/L) 2.22 ± 0.52 2.44 ± 0.56 2.82 ± 0.71 .110
TSH (μIU/mL) 3.44 ± 2.22 2.61 ± 1.03 3.12 ± 1.51 .501
expression of differential proteins between PCOS and control, IP (mouth) 5.29 ± 1.60 8.42 ± 4.38 9.05 ± 2.69 .033
PCOS-IR and PCOS-non-IR. Live birth rate 7/7 (100.00%) 10/12 (83.33%) 10/21 (47.62%) .014
Gene ontology (GO) is an important method and tool to BMI: body mass index; FSH: follicle-stimulating hormone; LH: luteinizing hor-
annotate genes and their products, which is beneficial to the mone; T: testosterone; FBG: fasting blood glucose; FINS: fasting insulin;
integration and utilization of biological data. The annotated, HOMA-IR: homeostasis model assessment of insulin resistance; CHO: choles-
visualized Database DAVID is an online data synthesis tool that terol; TG: triglyceride; HDL-C: high-density lipoprotein cholesterol; LDL-C:
low-density lipoprotein cholesterol; TSH: thyroid stimulating hormone; IP:
lays the foundation for successful high-throughput gene function interval between the time of endometrial collection and the time of final
analysis. The GO and KEGG of the DEPs between PCOS and pregnancy.
control, PCOS-IR and PCOS-non-IR, were enriched using p < .05 was considered statistically significant.

Figure 1. Sample expression correction box diagram and volcano diagram. A: Protein expression before normalization. B: Protein expression after sample normal-
ization. The red sample represents the PCOS-non-IR group, the green sample represents the PCOS-IR group and the blue sample represents the control group. C:
PCOS vs. Control. D: PCOS-IR vs. PCOS-non-IR. (Red dots represent up-regulated differential proteins, green dots represent down-regulated proteins, and no sig-
nificantly changed genes are marked as gray dots).
Data processing and screening of differential genes
The original dataset is normalized and presented as boxplots before
and after normalization (Figure 1A,B). All samples had 5331 proteins
identified, 275 proteins were differentially expressed in the PCOS
vs. control group and 215 proteins were differentially expressed in
the PCOS-IR group vs. PCOS-non-IR group, differences have been
indicated in the volcano diagram (Figure 1C,D).
GO and KEGG enrichment of DEPs in endometrial tissues
between different groups
All DEPs were uploaded to the DAVID online tool for analysis
of GO and KEGG pathway enrichment. Then, the R language
ggplot2 package is used for visual processing. The DEPs were
classified into three groups including biological process (BP),
cellular component (CC), and molecular function (MF).
In the PCOS vs. control group, DEPs of BP were enriched
in cellular protein metabolic processes, platelet degranulation,
post-translational protein modification, extracellular matrix orga-
nization, translational initiation, and cellular response to oxida-
tive stress (Figure 2A). CCs were enriched in extracellular
exosomes, blood microparticles, endoplasmic reticulum, extra-
cellular spaces, spectrin-associated cytoskeletons, and focal adhe-
sions (Figure 2B). The MF were enriched in RNA binding,
structural constituent of the cytoskeleton, protein binding, spec-
trin binding, extracellular matrix structural constituent, collagen
binding, and actin binding (Figure 2C). KEGG were enriched
in coronavirus disease – COVID-19, complement and coagulation
cascades, DNA replication, ferroelectricity, ECM-receptor inter-
action, and carbon metabolism (Figure 2D).
In the PCOS-IR vs. PCOS-non-IR group, the DEPs of BP were
enriched in platelet degranulation, neutrophil degranulation, viral
processes, very long-chain fatty acid catabolic processes, golgi to
plasma membrane transport, protein localization to the plasma
membrane, and intracellular protein transport (Figure 3A). CC
Gynecological Endocrinology 3
was enriched in the extracellular exosome, membrane, cytosol,
blood microparticles, nucleoplasm, focal adhesions, and perinu-
clear regions of the cytoplasm (Figure 3B). The MF were enriched
in protein binding, RNA binding, long-chain fatty acid transporter
activity, electron carrier activity, GTP binding, and ATPase activity
(Figure 3C). The KEGG were enriched for salmonella infection,
prion disease, oxidative phosphorylation, Parkinson’s disease, and
metabolic pathways (Figure 3D).
Identification of key proteins in endometrial tissues
between different groups
We visually displayed the intersection of differential proteins between
PCOS vs. control and PCOs-IR vs. PCOs-non-IR groups using Venn
diagrams and found 25 proteins with significant differences between
the groups (ACTR1A, ZC3H15, C1QC, NPIPB5, EMC4, TSC22D2,
DDX1, BRI3BP, CKB, PRDX2, CNN3, ABRAXAS2, CYBA, MCM7,
TMEM167A, GLT8D1, MUC5B, ATRX, TTPAL, HM13, KRT9,
TAGLN2, USP5, TUBA4A, and ORM1) (Figure 4A). We performed
PCA using 25 PCOS vs. control, PCOS-IR vs. PCOS-non-IR dif-
ferential proteins and found that the differential proteins were clearly
distinguished between PCOS-IR, PCOS-non-IR, and control (Figure
4B). Then using the LASSO regression model, we screened out five
DEPs, which ACTR1A, TSC22D2, CKB, ABRAXAS2, and TAGLN2
were associated with live birth of patients with PCOS (Figure 4C,D).
Then using the LASSO regression model, we screened five DEPs,
ACTR1A, TSC22D2, CKB, ABRAXAS22, and TAGLN2, for associ-
ation with live birth in patients with PCOS (Figure 4E).
The expression of key proteins between PCOS-IR, PCOS-
non-IR, and control groups
The expression levels of the key proteins were extracted from the
raw data, the protein expression was transformed by LOG2 so
that the data followed a normal distribution, and the differences
4 X. YANG ET AL.

Figure 2. PCOS vs. Control: GO and KEGG enrichment of differentially expressed proteins (DEPs). (A) biological process, (B) cellular component, (C) molecular
function, and (D) KEGG pathway analysis.

Figure 3. PCOS-IR vs. PCOS-non-IR: GO and KEGG enrichment of differentially expressed proteins (DEPs). (A) biological process, (B) cellular component, (C) molecular
function, and (D) KEGG pathway analysis.
 Gynecological Endocrinology 5

Figure 4. A: Venn diagrams to visually display the intersection of differential proteins between PCOS vs. control and PCOs-IR vs. PCOs-non-IR groups. B: PCA between
the PCOS-IR, PCOS-non-IR and control groups. The red sample represents the PCOS-non-IR group, the green sample represents the PCOS-IR group and the blue
sample represents the control group. C and D: The LASSO regression model for live birth of PCOS-IR patients. E: ROC of different factors on pregnancy outcome.

Figure 5. Analysis of key proteins expression: (A, B, E) ACTR1A and CKB were higher in PCOS-IR group, while TSC22D2 was lower in PCOS-IR group (p < .05). (C
and D) ABRAXAS2 was higher in all PCOS groups and TAGLN2 was low in all PCOS groups (p < .05), but there was no difference between the PCOS-IR and
PCOS-non-IR groups (p > .05). Pearson correlation analysis between key proteins and HOMA-IR: (F and G) ACTR1A and CKB was significantly positively correlated
with HOMA-IR. (H, I, and J) there was no significant correlation between TAGLN2, ABRAXAS2 and TSC22D2 with HOMA-IR.

between the groups were analyzed. We find that ACTR1A and
CKB are higher in the PCOS-IR group, while TSC22D2 is lower
in the PCOS-IR group (p < .05) (Figure 5A,B,E). ABRAXAS2 was
higher in all PCOS groups and TAGLN2 was lower in all PCOS
groups (p < .05), but there was no difference between the PCOS-IR
and PCOS-non-IR groups (p > .05) (Figure 5D,C). Pearson cor-
relation analysis shows that ACTR1A and CKB are significantly
positively correlated with HOMA-IR and negatively correlated
with HOMA-IR, while TAGLN2, TSC22D2, and ABRAXAS2 are
not correlated with HOMA-IR (Figure 5F–J).
6 X. YANG ET AL.
Discussion
We found that cellular protein metabolic process, platelet degran-
ulation, post-translational protein modification, extracellular
matrix organization, translational initiation, and cellular response
to oxidative stress were more active in the endometrium with
PCOS compared to controls. Platelet degranulation, neutrophil
degranulation, viral process, and very long-chain fatty acid cat-
abolic process was more active in the PCOS-IR group compared
to the PCOS-non-IR group.
Platelet degranulation, which is crucial for hemostasis and
may be involved in inflammatory responses, was found to be
activated in PCOS-IR in this study [11]. New evidence suggests
that patients with PCOS have abnormalities in coagulation and
fibrinolytic pathways that may explain the increased cardiovas-
cular risk of PCOS, but the effects of PCOS on coagulation
and fibrinolysis remain largely unexplored [12]. Limited studies
found that platelet count and platelet volume (MPV) of PCOS
were higher than that of age- and BMI-matched healthy control,
and a positive correlation between MPV and testosterone level
was observed [13–15]. Activation of platelets can lead to degran-
ulation and the release of cytokines into plasma that promote
the development of a pro-inflammatory state. Platelets also
interact with monocytes and neutrophils to further enhance
inflammation [16] According to known data, PCOS has chronic
low-grade inflammation throughout the body, which is mani-
fested in increased C-reactive protein (CRP), interleukin 18
(IL-18), and tumor necrosis factor (TNF-α). This chronic
inflammatory state is aggravated by obesity and hyperinsulin-
emia [17]. It has shown that excessive uterine inflammation of
PCOS may lead to the occurrence of secondary abortion, hyper-
insulinemia and insulin resistance will aggravate uterine inflam-
mation, and the molecular regulation of uterine inflammation
may provide a new therapeutic target for preventing PCOS
abortion [18]. Our study found enhanced platelet degranulation
and reduced platelet degranulation in patients with PCOS with
IR endometrial tissue, which is expected to improve the inflam-
matory status of the uterus in patients with PCOS and reduce
the abortion rate.
Very long-chain fatty acids as precursors of lipid mediators
are intricately regulated to maintain lipid homeostasis [19].
Patients with PCOS exhibit abnormal levels of lipid metabolism,
including phosphatidylcholine, FFAs, and PUFA metabolites.
Insulin resistance and increased androgen production are
important manifestations of PCOS and may adversely affect the
lipid profile of patients with PCOS, particularly bioactive lipid
metabolites derived from PUFAs [20]. The study also found
that increased BMI may be associated with very-long-chain fatty
acids [21]. Women with PCOS had lipid-induced increases in
plasma inflammatory cytokines, which in turn manifested in
paracrine and endocrine effects [22]. Lipid-induced production
of reactive oxygen species (ROS) within skeletal muscle pro-
motes mitochondrial dysfunction and the development of IR
[23]. In PCOS, mitochondrial dysfunction and abnormal glucose
metabolism increase ROS production, induce the release of
inflammatory factors, and regulate insulin resistance [24]. In
addition, OS can cause an imbalance in the uterine function,
which in turn can lead to reduced endometrial receptivity and
implant failure in the implantation window [25], is one of the
main causes of infertility in patients with PCOS. Very long-chain
fatty acid also accumulate during necroptosis [26]. In PCOS, a
variety of hormonal and metabolic stimuli can trigger different
types of regulated cell death under physiological and patholog-
ical conditions, including ferroptosis, apoptosis, and necroptosis.
Studies have found that insulin-exposed PCOS pregnant rats
exhibited decreased apoptosis in the uterus and increased necro-
ptosis in the placenta [27]. Our study found that for the endo-
metrium of PCOS-IR, the accumulation of very-long-chain fatty
acids has occurred before pregnancy, which may indicate that
the endometrium of PCOS-IR has already occurred necroptosis
before pregnancy, and if this process can be intervened before
pregnancy, it may improve the live birth rate of PCOS-IR.
In this study, ACTR1A and CKB were higher in the PCOS-IR
group, while TSC22D2 was lower in the PCOS-IR group. An
RNA interference study revealed the important role of ACTR1A
in the induction of pro-inflammatory cytokines, and identified
ACTR1A, part of the dynactin complex, as a novel regulator of
TLR2-mediated immune signaling [28]. Studies have shown that
TLR2, and TLR4-mediated activation of interferon regulatory
factor-7 (IRF-7) and NF-κB signaling, cytokine production, and
endometrial inflammation in patients with PCOS compared to
patients with non-PCOS [29]. Therefore, ACTR1A may be a
discovery in the regulation of PCOS-IR endometrial inflamma-
tory immunity. CKB is overexpressed in hyperplastic endometrial
tissue and several types of cancers such as endometrial cancer,
ovarian cancer, breast cancer, colon cancer, and colorectal cancer
[30–32]. A label-free Proteomics study comparing the endome-
trial proliferative and receptive phases found that the upregula-
tion of creatine kinase B-type (CKB) in the receptive phase may
be related to endometrium receptivity [30]. In our study, endo-
metrium during the PCOS proliferation period was used, and
the increased CKB level of PCOS-IR endometrium was found
during the proliferation period, suggesting that endometrium
development was not synchronous and the receptivity of endo-
metrium had changed, which was not conducive to pregnancy.
Finally, there are some limitations to this study. First, the
selected sample is in the proliferative endometrium, and the
patient is not pregnant in the current month. The factors that
influence a patient’s pregnancy are complex and diverse, and this
study can only reflect some of the factors that influence pregnancy
outcomes. Second, lifestyle and medication interventions were
given according to the patient’s condition after the patient visited
the hospital and completed the corresponding checkup, but they
were not considered due to heterogeneity in interventions. Third,
the sample size is relatively small. In future experiments, we will
expand the sample size and conduct an in-depth study.
Conclusion
This study explored the endometrial proteome in PCOS-IR,
PCOS-non-IR, and controls. A panel of proteins associated with
platelet degranulation, neutrophil degranulation, viral process,
and very long-chain fatty acid catabolic process were found in
the endometrium of PCOS-IR patients. Moreover, ACTR1A and
CKB may be considered as candidate biomarkers of PCOS-IR.
The results provide us with a better understanding of the endo-
metrium abnormalities in PCOS-IR.
Acknowledgment
The authors thank Shanghai Applied Protein Technology for supporting this
methodology.
Disclosure statement
The authors declare no potential conflicts of interest.

Ethics approval
The study was approved by the ethics committee of Lanzhou University
Second Hospital (Ethical Approval Number: 2017 A-057). All subjects gave
informed consent before participating.
Availability of data and materials
The Trial data used in this manuscript have been deposited to the Proteome
Xchange Consortium (http://proteomecentral.proteomexchange.org) via the
iProX partner repository with the dataset identifier PXD032383.
Funding
This work was granted by the Science Foundation of Lanzhou University
(Grant No. 054000229) and Cuiying Scientific and Technological Innovation
Program of Lanzhou University Second Hospital.
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