Polycystic Ovary Syndrome Tissues Research Articles

Circ_0043314 Modulates Proliferation and Apoptosis of Ovarian Granulosa Cells in Polycystic Ovarian Syndrome via the MicroRNA-146b-3p/Apelin 13 Axis.

Polycystic ovary syndrome (PCOS) is a common endocrine and metabolic disorder in women. At present, the pathogenesis has not been clarified, and the clinical application of drugs and lifestyle intervention may not prevent disease progression. This study aimed to investigate how circ_0043314 regulates ovarian granulosa cell biological functions to provide a theoretical basis for the treatment of patients with PCOS. MicroRNA (miR)-146b-3p/Apelin 13 axis was used to investigate the mechanism by which circ_0043314 regulated ovarian granulosa cell proliferation and apoptosis in PCOS via miR-146b-3p/Apelin 13 axis. Participants/Materials, Methods: Ovarian tissues (cortical tissues) from 35 PCOS patients and 35 normal controls, as well as HEK293T and human ovarian granulosa cell line (KGN, COV434), were included in this study. We examined the expression levels of circ_0043314, miR-146b-3p, and Apelin 13 in PCOS tissues. Ovarian granulosa cells were transfected with corresponding plasmids to clarify the influence of circ_0043314, miR-146b-3p, or Apelin 13 on proliferation and apoptosis of ovarian granulosa cells through MTT and flow cytometry assays. Moreover, the relationships among circ_0043314, miR-146b-3p, and Apelin 13 were analyzed through dual-luciferase and RNA immunoprecipitation assays. Circ_0043314 and Apelin 13 were highly expressed and miR-146b-3p was lowly expressed in ovarian tissues of PCOS compared with non-PCOS controls. Downregulation of circ_0043314 or upregulation of miR-146b-3p hindered ovarian granulosa cell proliferation and advanced its apoptosis. Downregulation of miR-146b-3p reversed the impacts of downregulation of circ_0043314, and overexpression of Apelin 13 counteracted the influences of upregulation of miR-146b-3p in ovarian granulosa cells. Mechanically, circ_0043314 could bind to miR-146b-3p, and miR-146b-3p directly targeted and modulated Apelin 13 expression. This study was limited by the lack of animal experiments. Our data demonstrated that circ_0043314 enhances ovarian granulosa cell proliferation and suppresses its apoptosis via miR-146b-3p/Apelin 13 axis.

PGD: Shared gene linking polycystic ovary syndrome and endometrial cancer, influencing proliferation and migration through glycometabolism.

The relationship among polycystic ovary syndrome (PCOS), endometrial cancer (EC), and glycometabolism remains unclear. We explored shared genes between PCOS and EC, using bioinformatics to unveil their pathogenic connection and influence on EC prognosis. Gene Expression Omnibus datasets GSE226146 (PCOS) and GSE196033 (EC) were used. A protein-protein interaction (PPI) network was constructed to identify the central genes. Candidate markers were screened using dataset GSE54250. Differences in marker expression were confirmed in mouse PCOS and human EC tissues using RT-PCR and immunohistochemistry. The effect of PGD on EC proliferation and migration was explored using Ki-67 and Transwell assays. PGD's impact on the glycometabolic pathway within carbon metabolism was assessed by quantifying glucose content and lactic acid production. R software identified 31 common genes in GSE226146 and GSE196033. Gene Ontology functional classification revealed enrichment in the "purine nucleoside triphosphate metabolism process," with key Kyoto Encyclopedia of Genes and Genomes pathways related to "carbon metabolism." The PPI network identified 15 hub genes. HK2, NDUFS8, PHGDH, PGD, and SMAD3 were confirmed as candidate markers. The RT-PCR analysis validated distinct HK2 and PGD expression patterns in mouse PCOS ovarian tissue and human EC tissue, as well as in normal and EC cells. Transfection experiments with Ishikawa cells further confirmed PGD's influence on cell proliferation and migration. Suppression of PGD expression impeded glycometabolism within the carbon metabolism of EC cells, suggesting PGD as a significant PCOS risk factor impacting EC proliferation and migration through modulation of single carbon metabolism. These findings highlight PGD's pivotal role in EC onset and prognosis.

Immune dysfunction mediated by the competitive endogenous RNA network in fetal side placental tissue of polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS), a common endocrine and metabolic disorder affecting women in their reproductive years. Emerging evidence suggests that the maternal-fetal immune system is crucial for proper pregnancy. However, whether immune function is altered at the end of pregnancy in PCOS women and the underlying molecular mechanisms is currently unexplored. Herein, the basic maternal immune system was investigated (n = 136 in the control group; n = 103 in the PCOS group), and whole-transcriptome sequencing was carried out to quantify the mRNAs, miRNAs, and lncRNAs expression levels in fetal side placental tissue of women with PCOS. GO, KEGG, and GSEA analysis were employed for functional enrichment analysis. The process of identifying hub genes was conducted utilizing the protein-protein interaction network. CIBERSORT and Connectivity Map were deployed to determine immune cell infiltration and predict potential drugs, respectively. A network of mRNA-miRNA-lncRNA was constructed and then validated by qRT-PCR. First, red blood cell count, neutrophil count, lymphocyte count, hypersensitive C-reactive protein, and procalcitonin were significantly elevated, while placental growth factor was hindered in PCOS women. We identified 308 DEmRNAs, 77 DEmiRNAs, and 332 DElncRNAs in PCOS samples. Functional enrichment analysis revealed that there were significant changes observed in terms of the immune system, especially the chemokine pathway. Eight genes, including FOS, JUN, EGR1, CXCL10, CXCR1, CXCR2, CXCL11, and CXCL8, were considered as hub genes. Furthermore, the degree of infiltration of neutrophils was dramatically decreased in PCOS tissues. In total, 57 ceRNA events were finally obtained, and immune-related ceRNA networks were validated. Some potential drug candidates, such as enalapril and RS-100329, could have a function in PCOS therapy. This study represents the inaugural attempt to evaluate the immune system at the end of pregnancy and placental ceRNA networks in PCOS, indicating alterations in the chemokine pathway, which may impact fetal and placental growth, and provides new therapy targets.

Screening of potential biomarkers for polycystic ovary syndrome and identification of expression and immune characteristics.

Polycystic ovary syndrome (PCOS) seriously affects the fertility and health of women of childbearing age. We look forward to finding potential biomarkers for PCOS that can aid clinical diagnosis. We acquired PCOS and normal granulosa cell (GC) expression profiles from the Gene Expression Omnibus (GEO) database. After data preprocessing, differentially expressed genes (DEGs) were screened by limma package, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis and Gene Set Enrichment Analysis (GSEA) were performed. Recursive feature elimination (RFE) algorithm and the least absolute shrinkage and selection operator (LASSO) Cox regression analysis were used to acquire feature genes as potential biomarkers. Time-dependent receiver operator characteristic curve (ROC curve) and Confusion matrix were used to verify the classification performance of biomarkers. Then, the expression characteristics of biomarkers in PCOS and normal cells were analyzed, and the insulin resistance (IR) score of samples was computed by ssGSEA. Immune characterization of biomarkers was evaluated using MCP counter and single sample gene set enrichment analysis (ssGSEA). Finally, the correlation between biomarkers and the scores of each pathway was assessed. We acquired 93 DEGs, and the enrichment results indicated that most of DEGs in PCOS group were significantly enriched in immune-related biological pathways. Further screening results indicated that JDP2 and HMOX1 were potential biomarkers. The area under ROC curve (AUC) value and Confusion matrix of the two biomarkers were ideal when separated and combined. In the combination, the training set AUC = 0.929 and the test set AUC = 0.917 indicated good diagnostic performance of the two biomarkers. Both biomarkers were highly expressed in the PCOS group, and both biomarkers, which should be suppressed in the preovulation phase, were elevated in PCOS tissues. The IR score of PCOS group was higher, and the expression of JDP2 and HMOX1 showed a significant positive correlation with IR score. Most immune cell scores and immune infiltration results were significantly higher in PCOS. Comprehensive analysis indicated that the two biomarkers had strong correlation with immune-related pathways. We acquired two potential biomarkers, JDP2 and HMOX1. We found that they were highly expressed in the PCOS and had a strong positive correlation with immune-related pathways.

The ameliorative effect of CangFu Daotan Decoction on polycystic ovary syndrome of rodent model is associated with m6A methylation and Wnt/β-catenin pathway

Objective: This study investigates the effects of CangFu Daotan Decoction (CDD) on m6A methylation and Wnt/β-catenin pathway in rats with polycystic ovary syndrome (PCOS).Methods: The PCOS rat model was established by letrozole gavage. The rats were fed high-fat chow, and their body weight and blood glucose were recorded. The expressions of follicle-stimulating hormone(FSH), luteinizing hormone(LH), and testosterone(T) were quantified by ELISA. Chemical components in CDD were analyzed using UPLC-Q/TOF-MS. Based on network pharmacology methods, related targets of CDD on PCOS were screened. An enrichment analysis according to Tokyo Encyclopedia of Genes and Genomes (KEGG) was conducted to predict the potential signaling pathway of CDD in PCOS. The expressions of Wnt-1, β-Catenin, GSK-3β, C-MYC, Beclin1, LC3II, Bax, and PCNA were detected by western blotting. The expressions of Mettl3, Mettl14, Fto, Alkbh5, Ythdf1, and Ythdf2 were monitored by RT-PCR. The expressions of Mettl3, Fto, and Ythdf1 were detected by western blotting.Results: Letrozole and a high-fat diet induced ovarian dysfunction in rats, which was attenuated by CDD. CDD decreased blood glucose, LH, and T concentrations and increased FSH expression in PCOS. After removing duplicates, a total of 71 compounds were identified by UHPLC-Q/TOF-MS, among which terpenoids and flavonoids account for the main proportion. The clustering analysis showed that the active site of CDD might be in the Wnt/β-catenin pathway. CDD decreased the expressions of Wnt-1, β-Catenin, GSK-3β, C-MYC, Beclin1, LC3II, and Bax and increased PCNA expression in the ovarian tissue of PCOS rats. CDD decreased the m6A gene expressions of Mettl3, Mettl14, Fto, Alkbh5, Ythdf1, and Ythdf2 in peripheral blood and ovarian tissue of PCOS rats. CDD reduced the m6A proteins expressions of Mettl3, Fto, and Ythdf1 in the ovarian tissue of PCOS rats.Conclusion: CDD can regulate m6A modification and inhibit the Wnt/β-catenin signaling pathway in PCOS rats, thereby reducing body weight, lowering blood glucose levels, improving sex hormone disorders, and decreasing autophagy and apoptosis in ovarian tissue to promote the recovery of ovarian morphology.

The overexpression of neurokinin B-neurokinin 3 receptor system exerts direct effects on the ovary under PCOS-like conditions to interfere with mitochondrial function.

The increased hypothalamic neurokinin B (NKB) level may contribute to the hyperactive LH pulse secretion in Polycystic ovary syndrome (PCOS). However, the expression and role of the neurokinin B-neurokinin 3 receptor (NKB-NK3R) system in the local ovarian tissue of PCOS have not been clarified. We constructed in vivo and in vitro models to elucidate the mechanism of the NKB-NK3R pathway in reproductive endocrine disorders of PCOS. The granulosa cell line-KGN cells were set in palmitic acid (PA) and dihydrotestosterone (DHT) to simulate the PCOS-like conditions. And we used the high-fat/high-glucose diet to build a PCOS-like mice model and neurokinin 3 receptor antagonist (NK3Ra) was administered to half of the mice. The expression of the NKB-NK3R system, mitochondrial functions, hormone levels, and inflammatory state was evaluated. The PCOS-like stimulations induced the NKB-NK3R system and MAPK-ERK pathway overexpression in KGN cells, in an approximate dose and time-dependent manner. The NKB-NK3R system overactivated the MAPK-ERK pathway to increase NNT overexpression, disturb NADH/NADPH pools, aggravate the oxidation state, and decrease ATP production. With overexpression of the NKB-NK3R system in the local ovarian tissue, ovulatory dysfunction, progesterone deficiency, and pro-inflammatory states were apparent in PCOS-like mice. Antagonizing the receptor, NK3R, reversed the adverse reproductive endocrine phenotypes via improving mitochondrial dysfunction. In addition to the central regulation, local ovarian overexpression of the NKB-NK3R system participated in the adverse reproductive endocrine phenotypes, supporting the therapeutic implications of NK3Ra for PCOS.

Withdrawal: "Long Non-coding RNA NEAT1 Drives the Development of Polycystic Ovary Syndrome via Sponging Multiple MicroRNAs" by Xia Sang and Yuzhen Zhang.

A line of evidences have validated that multiple microRNAs (including miR-16, miR-483 and miR-324-3p) were abnormally expressed in granulosa cells, which may be closely related with the pathogenesis of polycystic ovary syndrome (PCOS). Long-chain non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) has been proved to participate in the progression of various human diseases via regulating microRNAs, but its function in PCOS are not yet clear. In this work, RT-PCR was performed to detect NEAT1 expression in PCOS tissues, and human ovarian granulosa cell line KGN were taken as the model to construct the cell line with high expression and low expression of NEAT1. CCK-8 and flow cytometry were carried out to monitor cells proliferation and apoptosis, respectively. Besides, we performed luciferase reporter gene assay and RNA binding protein immunoprecipitation (RIP) assay to verify the interaction between NEAT1, miR-16, miR-483 and miR-324-3p. We demonstrated that NEAT1 was highly expressed in PCOS tissues and cells. Over-expressed NEAT1 promoted the proliferation and inhibited the apoptosis. Moreover, NEAT1 was negatively correlated with the expression levels of miR-16, miR-483 and miR-324-3p. Dual luciferase analysis and RIP assay confirmed that NEAT1 can specifically bind to miR-16, miR-483 and miR-324-3p, by which NEAT1 can reduce their expression. In conclusion, NEAT1 can promote the proliferation of ovarian granulosa cells and arrest apoptosis via impeding expressions of miR-16, miR-483 and miR-324-3p. Our research would shed new light on the molecular mechanism of ovarian granulosa cell disorder. This article is protected by copyright. All rights reserved.

Cryptotanshinone Protects against PCOS-Induced Damage of Ovarian Tissue via Regulating Oxidative Stress, Mitochondrial Membrane Potential, Inflammation, and Apoptosis via Regulating Ferroptosis.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of childbearing age. Cryptotanshinone (CRY) has been shown to be effective in reversing reproductive disorders, but whether it can be used in the treatment of polycystic ovary syndrome remains unclear. We aimed to explore whether the mechanism of cryptotanshinone (CRY) in the treatment of polycystic ovary syndrome (PCOS) can be driven via regulating ferroptosis. A rat model of PCOS was established by daily injection of human chorionic gonadotropin and insulin for 22 days. An in vitro model of ischemia-reperfusion (IR) of granulosa cells was established. The in vitro and rat models of PCOS were subjected to different treatments including ferroptosis activators and inhibitors, CRY, and MAPK inhibitor. Oxidative stress was evaluated by measuring the activities of SOD, MDA, and GSH-PX. Total body weight and ovarian weight, as well as the levels of LH and the LH to FSH ratio, significantly increased in rats with PCOS, compared with controls. The expression of Bax was increased in PCOS tissues while PGC1α, NFR1, GPX4, catalase p-ERK, and Bcl-2 were all downregulated. Ferroptosis activator, erastin, had effects similar to those of PCOS while the contrary was found with CRY and ferroptosis inhibitor treatment groups. In vitro, CRY inhibited oxidative stress, MMP, and NF-κB and activated MAPK/ERK signaling by regulating ferroptosis. Overall, this study indicated that CRY protects against PCOS-induced damage of the ovarian tissue, via regulating oxidative stress, MMP, inflammation, and apoptosis via regulating ferroptosis.

Aberrant activation of the Hedgehog signaling pathway in granulosa cells from patients with polycystic ovary syndrome

ABSTRACT The molecular mechanism that triggers polycystic ovary syndrome (PCOS) is mysterious. Abnormal development of ovarian granulosa cells (GCs) is one of the causes of PCOS. Herein, our study was carried out using RNA-seq to detect the different gene expression levels in ovarian GCs between three patients with PCOS and four normal controls. To verify the RNA-seq data, GCs from 22 patients with PCOS and 21 controls with normal ovulation were collected to perform the RT-PCR analysis. Hedgehog signaling pathway (Hh) members, Ihh and Ptch2 were abnormally highly expressed in the PCOS tissue (PT). The qPCR also indicated that the expression levels of Hh signaling pathway downstream members, Ptch1, Gli1, and Gli2 in the PT were significantly higher than those in the normal tissue (NT). Besides, the expression of TNF-α mRNA in PCOS patients was higher than that in the control group. Through the chromatin immunoprecipitation assay (ChIP), we found that the Gli1-IP-DNA enriched from the granular cells of PCOS patients was higher than that of the control group. Finally, the Hh signaling pathway inhibitor, cyclopamine, can decrease the apoptosis of PCOS ovarian granulosa cells. These results suggest that abnormal activation of Hh signaling pathway, especially Ihh signal, may have a profound influence on PCOS.

MicroRNA-664a-3p inhibits the proliferation of ovarian granulosa cells in polycystic ovary syndrome and promotes apoptosis by targeting BCL2A1.

BackgroundTo investigate whether micro ribonucleic acid-664a-3p (miR-664a-3p) targeting BCL2A1 affects the proliferation and apoptosis of ovarian granulosa cells.MethodsReal-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-664a-3p in granulosa cells, granular tumor cell lines (KGN), and normal ovarian epithelial cell lines (IOSE80) in the polycystic ovary syndrome (PCOS) group and the control group. After overexpressing miR-664a-3p or inhibiting its expression in KGN cells, qRT-PCR and Western blotting were used to detect the messenger RNA (mRNA) and protein levels of related genes. At the same time, a cell counting kit-8 (CCK-8) and flow cytometer were used to detect cell proliferation and apoptosis. The TargetScan website was used to predict the potential binding sites of miR-664a-3p and B-cell lymphoma 2-related protein A1 (BCL2A1), which was further verified by qRT-PCR, Western blotting, and the luciferase reporter gene method.ResultsThe expression of miR-664a-3p was significantly decreased in both PCOS tissues and KGN cells (both P<0.05), and the overexpression of miR-664a-3p inhibited the proliferation of KGN cells and induced their apoptosis. Moreover, our results confirmed that miR-664a-3p directly targets BCL2A1 (P<0.05), and the inhibitory effect of miR-664a-3p on KGN cells was reversed by BCL2A1 overexpression (both P<0.05). The up-regulation of BCL2A1 promotes cell proliferation and reduces cell apoptosis by the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway (both P<0.05).ConclusionsThe up-regulation of miR-664a-3p inhibits the proliferation of KGN cells and increases apoptosis by down-regulating the expression of BCL2A1 and blocking the MAPK/ERK signaling pathway.

Expression of PGC-1ɑ in Ovarian Tissue of PCOS Rat Model

To investigate the expression of peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α) in the ovarian tissue of polycystic ovary syndrome (PCOS) rat model and obese PCOS rat model, and the possible mechanism of PCOS. Thirty SD rats were randomly divided into control group, PCOS rat model group and obese PCOS rat model group. DHEA dissolved in 0.2 mL soybean oil was injected daily into the rats of two PCOS groups for 21 d. Rats in obese PCOS model group were added with high-fat diet based on DHEA modeling, and each group had 10 rats. Body mass were measured before and on the 22 nd day after modeling. The serum testosterone (T) levels were measured by abdominal aortic blood, and the ovarian tissues of rats were taken for histological changes were observed by HE staining, immunohistochemical staining and Western blot to detect PGC-1ɑ protein expression. The body mass of rats in each group increased after modeling, and the body mass of rats in PCOS group and obese PCOS group increased significantly ( P<0.05). The serum T concentration of two PCOS model groups was higher than that of control group ( P<0.01). The serum T concentration in obese PCOS model group was higher than that in the PCOS group ( P<0.05). The results of HE staining of rat ovarian tissue showed that there were follicles and a small amount of corpus luteum at different developmental stages in the control group, and the granulosa cells were arranged in 4-6 layers. The number of immature small follicles in the two PCOS groups was significantly increased. The granulosa cells were arranged in 1-3 layers, relatively looser, and some follicles were atresia. In the obese PCOS group, the diameter of ovarian atretic follicles increased, the number of granulocyte layers decreased, and oocytes disappeared more obviously. Immunohistochemical staining showed that the PGC-1ɑ protein was mainly expressed in the cumulus and granulosa cells of ovarian tissue in the control group. The mean gray level of PGC-1ɑ protein expression in PCOS group (0.53±0.06) and obese PCOS group (0.36±0.03) was lower than that of the control group (0.75±0.03), with the statistical difference ( P<0.05). PGC-1ɑ expression in the obese PCOS group was lower than that in the PCOS group ( P<0.01). The results of Western blot were consistent with those of immunohistochemical staining. PGC-1ɑ is associated with damage of ovarian granulosa cells in high-fat environment. The decrease of PGC-1ɑ expression in granulosa cells of ovarian follicles may be an important cause of PCOS development.

Proteome analysis of endometrial tissue from patients with PCOS reveals proteins predicted to impact the disease.

Polycystic ovary syndrome (PCOS) is a complex disease that causes an ovulatory infertility in approximately 10% of reproductive-age women. We searched for candidate proteins that might contribute to endometrial receptivity defects in PCOS patients, and result in adverse reproductive outcomes. Shotgun proteomics approach was used to investigate the proteome profile of the endometrium at the luteal phase in PCOS patients compared to healthy fertile individuals. Biological process and pathway analyses were conducted to categorize the proteins with differential expressions. Confirmation was performed for a number of proteins via immunoblotting in new samples. 150 proteins with higher abundance, and 46 proteins with lower abundance were identified in the endometrial tissue from PCOS patients compared to healthy fertile individuals. The proteins with higher abundance were enriched in protein degradation, cell cycle, and signaling cascades. Proteins with lower abundance in PCOS patients were enriched in extracellular matrix (ECM) composition and function, as well as the salvage pathway of purine biosynthesis. Metabolism was the most affected biological process with over 100 up-regulated, and approximately 30 down-regulated proteins. Our results indicate significant imbalances in metabolism, proteasome, cell cycle, ECM related proteins, and signaling cascades in endometrial tissue of PCOS, which may contribute to poor reproductive outcomes in these patients. We postulate that the endometria in PCOS patients may not be well-differentiated and synchronized for implantation. Possible roles of the above-mentioned pathways that underlie implantation failure in PCOS will be discussed. Our findings need to be confirmed in larger populations.

Altered vitamin D3 metabolism in the ovary and periovarian adipose tissue of rats with letrozole-induced PCOS

Vitamin D3 (VD3) plays an important role in the ovary and its deficiency is associated with ovarian pathologies, including polycystic ovary syndrome (PCOS). However, there is no data related to VD3 metabolism in the ovary during PCOS. Herein, we investigated differences in the expression of VD3 receptor (VDR) and key VD3 metabolic enzymes, 1α-hydroxylase (CYP27B1) and 24-hydroxylase (CYP24A1), in the ovary and periovarian adipose tissue (POAT) of control (proestrus and diestrus) and PCOS induced by letrozole rats. Vdr, Cyp27b1 and Cyp24a1 mRNA expression was determined, their protein abundance was examined and immunolocalized. Furthermore, VD3 metabolite concentrations in plasma (25OHD) and tissues (ovary and POAT; 1,25(OH)2D3), and plasma calcium level were determined. 25OHD concentration decreased markedly in letrozole-treated rats in comparison with controls, whereas calcium concentration did not vary among the examined groups. The amount of 1,25(OH)2D3 decreased in both ovary and POAT of PCOS rats. In the ovary, we found decreased Cyp27b1 and increased Vdr mRNA expression in letrozole-treated and diestrus control group. Corresponding protein abundances were down-regulated and up-regulated, respectively but only following letrozole treatment. In POAT, only Cyp27b1 transcript level and CYP27B1 protein abundance were decreased in letrozole-treated rats. VDR was immunolocalized in healthy and cystic follicles, while CYP27B1 and CYP24A1 were found exclusively in healthy ones. Concluding, our results provide the first evidence of disrupted VD3 metabolism in the ovary and POAT of PCOS rats. The reduced 1,25(OH)2D3 concentration in those tissues suggests their contribution to VD3 deficiency observed in PCOS and might implicate in PCOS pathogenesis.

Lipidomic Biomarkers in Polycystic Ovary Syndrome and Endometrial Cancer.

Women with polycystic ovary syndrome (PCOS) are more likely to develop endometrial cancer (EC). The molecular mechanisms which increase the risk of EC in PCOS are unclear. Derangements in lipid metabolism are associated with EC, but there have been no studies, investigating if this might increase the risk of EC in PCOS. This was a cross-sectional study of 102 women in three groups of 34 (PCOS, EC and controls) at Nottingham University Hospital, UK. All participants had clinical assessments, followed by obtaining plasma and endometrial tissue samples. Lipidomic analyses were performed using liquid chromatography (LC) coupled with high resolution mass spectrometry (HRMS) and the obtained lipid datasets were screened using standard software and databases. Using multivariate data analysis, there were no common markers found for EC and PCOS. However, on univariate analyses, both PCOS and EC endometrial tissue samples showed a significant decrease in monoacylglycerol 24:0 and capric acid compared to controls. Further studies are required to validate these findings and investigate the potential role of monoacylglycerol 24:0 and capric acid in the link between PCOS with EC.

CircPUM1 promotes polycystic ovary syndrome progression by sponging to miR-760.

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders among reproductive-age women. The circRNA-miRNA axis functions in various diseases progression have been partially revealed in the past two decades. However, little is known about the role of the circRNA-miRNA axis in PCOS progression. MicroRNA miR-760, which is characterized by tissue-specific, has been studied in several cancers. Firstly, we found that miR-760 expression was decreased in PCOS tissues insulin treated GCs, KGN and SVOG cells. Secondly, The CCK-8 and apoptosis experiment results suggested that downregulated miR-760 promoted cell proliferation ability and suppressed apoptosis activity in KGN and SVOG cells. Then, the bioinformatic analysis result indicated that circPUM1 was a potential sponge to miR-760. By performing AGO2-RIP, RNA pull-down, Luciferase reporter, and qRT-PCR experiments, we demonstrated that circPUM1 acted as a molecular sponge to miR-760, and decreased miR-760 expression. Moreover, it was found that the promotive effect of circPUM1 was mediated by regulating miR-760. Collectively, our findings suggest that circPUM1 promotes PCOS progression through sponging to miR-760. We may provide a promising therapeutic target for PCOS.

MicroRNA-486-5p inhibits ovarian granulosa cell proliferation and participates in the development of PCOS via targeting MST4.

To explore whether microRNA-486-5p affected the proliferation of ovarian granulosa cells by targeting MST4 (silk/threonine protein kinase 4), thereby promoting the development of polycystic ovary syndrome (PCOS). The level of microRNA-486-5p in PCOS tissues and adjacent normal tissues was detected by quantitative real-time polymerase chain reaction (qRT-PCR). After microRNA-486-5p up-regulation in KNG cells, the mRNA and protein level of related genes was examined using qRT-PCR and western blot assay, respectively. Meanwhile, cell proliferation and cell cycle were analyzed by cell counting kit-8 (CCK-8) assay and flow cytometry. After insulin treatment of KNG cells, expressions of microRNA-486-5p and MST4, cell proliferation as well as cell cycle, were detected by qRT-PCR, CCK-8 and flow cytometry, respectively. Furthermore, cell proliferation and cycle situation were examined after simultaneous up-regulation of MST4 and microRNA-486-5p in vitro. MicroRNA-486-5p expression in PCOS tissues was significantly lower than that of normal tissues. In KNG cells, up-regulation of microRNA-486-5p significantly inhibited cell proliferation and cell cycle. The levels of cycle-associated proteins including CDK2 and CCNB1 decreased significantly. The results of dual-luciferase reporter gene assay showed that microRNA-486-5p could bind to MST4. After up-regulating microRNA-486-5p, both the mRNA and protein levels of MST4 decreased remarkably. MST4 expression was found significantly elevated in PCOS tissues as well. After overexpression of MST4, cell proliferation was enhanced, cell cycle was promoted, and expressions of cycle-related proteins increased. After treatment with different concentrations of insulin in KNG cells, the expression level of microRNA-486-5p decreased in a concentration-dependent manner. However, opposite results were observed in MST4 level. Meanwhile, the proliferation ability and cell cycle of insulin-treated cells were significantly enhanced. In addition, the inhibitory effect of microRNA-486-5p on cell proliferation and cell cycle could be partially reversed by simultaneous up-regulation of MST4 and microRNA-486-5p. MicroRNA-486-5p can bind to MST4 in a targeted manner and inhibit the proliferation of ovarian granulosa cells, thereby inhibiting the development of PCOS.

High level of visfatin and the activation of Akt and ERK1/2 signaling pathways are associated with endometrium malignant transformation in polycystic ovary syndrome

This study aimed to assess the clinicopathological significance of serum levels and endometrium tissue expression of visfatin in polycystic ovary syndrome (PCOS) patients. A total of 80 PCOS patients and 80 matching controls were included in this study. We analyzed the relationship between the expression of visfatin in endometrium and clinicopathological characteristics in PCOS patients. The correlation between the expression of visfatin and p-Akt, Akt, p-ERK1/2, and ERK1/2 in PCOS tissues was evaluated as well. Visfatin expression in PCOS endometrial tissues were significantly higher than those in controls (p = .001). The expression of phosphorylation of Akt and ERK1/2 were significantly higher in PCOS endometrium tissues compared to controls (p < .05). Moreover, a high expression of tissue visfatin in PCOS tissues was positively correlated with the expression of p-Akt (p = .015), and p-ERK1/2 (p = .013). Western blotting revealed that protein expression of visfatin in PCOS patients with endometrial hyperplasia and cancer was higher than that in patients with normal endometrium tissues, and the difference was statistically significant (p = .027). The expression of p-Akt (p = .018) and p-ERK1/2 (p = .035) in PCOS patients with endometrial hyperplasia and cancer was significantly higher than that in patients with normal endometrium tissues. Visfatin may be a potential biomarker for endometrial malignant transformation in PCOS patients.

MiR-222 promotes the progression of polycystic ovary syndrome by targeting p27 Kip1

Polycystic ovary syndrome (PCOS) is one of the most complex and common reproductive and endocrinologic disorders in the child-bearing age of women. Recently, miR-222 were reported to be associated with the etiology of PCOS. However, the function of miR-222 during the pathogenesis of PCOS remains unclear. In the present study, we aimed to investigate the role of miR-222 in PCOS. Firstly, miR-222 expression was examined by quantitative real-time PCR (qRT-PCR) in PCOS. The effects of miR-222 on proliferation, apoptosis and cell cycle in KGN cells were analyzed by CCK-8 assay and flow cytometry analysis, respectively. In addition, bioinformatics analysis was used to predict the target genes of miR-222, and dual-luciferase reporter assay was applied to verified the interaction between miR-222 and p27 Kip1 in KGN cells. Moreover, the expressions of p27 Kip1 in KGN cells treated with miR-222 mimics or miR-222 inhibitor were evaluated by qRT-PCR and western blot assays. The results showed that the expression of miR-222 was remarkably upregulated in PCOS tissues compared with corresponding normal tissues. In the gain-of-function and loss-of-function assays, we revealed that miR-222 mimics significantly promoted cell proliferation, while miR-222 inhibitor induced cell apoptosis and cell cycle arrested. Furthermore, p27 Kip1 was identified as a target gene of miR-222, and could be negatively regulated by miR-222 mimics in KGN cells. In conclusion, our findings suggested that miR-222 may promote the progression of PCOS by targeting p27 Kip1.

Autophagy is activated in the ovarian tissue of polycystic ovary syndrome.

The importance of autophagy in polycystic ovary syndrome (PCOS)-related metabolic disorders is increasingly being recognized, but few studies have investigated the role of autophagy in PCOS. Here, transmission electron microscopy demonstrated that autophagy was enhanced in the ovarian tissue from both humans and rats with PCOS. Consistent with this, ovarian granulosa cells from PCOS rats showed increases in the autophagy marker protein light chain 3B (LC3B), whereas levels of the autophagy substrate SQSTM1/p62 were decreased. In addition, the ratio of LC3-II/LC3-I was markedly elevated in human PCOS ovarian tissue compared with normal ovarian tissue. Real-time PCR arrays indicated that 7 and 34 autophagy-related genes were down- and up-regulated in human PCOS , Signal-Net, and regression analysis suggested that there are a wide range of interactions among these 41 genes, and a potential network based on EGFR, ERBB2, FOXO1, MAPK1, NFKB1, IGF1,TP53 and MAPK9 may be responsible for autophagy activation in PCOS. Systematic functional analysis of 41 differential autophagy-related genes indicated that these genes are highly involved in specific cellular processes such as response to stress and stimulus, and are linked to four significant pathways, including the insulin, ERBB, mTOR signaling pathways and protein processing in the endoplasmic reticulum. This study provides evidence for a potential role of autophagy disorders in PCOS in which autophagy may be an important molecular event in the pathogenesis of PCOS.

MicroRNA‑320a inhibition decreases insulin‑induced KGN cell proliferation and apoptosis by targeting PCGF1.

MicroRNAs (miRNAs) are widely involved in regulation of cellular processes of polycystic ovary syndrome (PCOS). However, the function of miR‑320a in PCOS remains unclear. The present study aimed to explore the effect of miR‑320a on PCOS cell proliferation and apoptosis following treatment with insulin, and to clarify the underlying mechanism. PCOS tissues and corresponding normal tissues were collected from 16 female patients with PCOS. KGN cells were pre‑treated with insulin, and KGN cells were transfected with ASO‑miR‑320a, miR‑320a mimics and polycomb group ring finger 1 (PCGF1) overexpression plasmids. Expressions of miR‑320a and PCGF1 were detected using the reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). Dual‑Luciferase reporter assays were performed to investigate the target gene of miR‑320a. MTS, colony formation and flow cytometry assays were performed to determine cell viability, colony formation, and apoptosis, respectively. Furthermore, mRNA and protein expression levels of B‑cell lymphoma 2 apoptosis regulator (Bcl‑2) and Bcl‑2 associated protein X apoptosis regulator (Bax) were examined using RT‑qPCR and western blotting. The results demonstrated that miR‑320a expression was significantly increased in PCOS tissues compared with normal tissues. Moreover, miR‑320a was upregulated in insulin‑induced cells in a dose‑dependent manner. Inhibition of miR‑320a suppressed insulin‑induced cell viability and colony formation, and promoted apoptosis. Luciferase reporter assays demonstrated that PCGF1 was a target of miR‑320a. Additionally, PCGF1 overexpression inhibited cell viability and colony formation and promoted apoptosis. Additionally, the mRNA and protein levels of Bcl‑2 were inhibited by miR‑320a suppression and PCGF1 overexpression, while Bax expression was promoted by them in insulin‑induced cells. The results of the present study demonstrated that miR‑320a inhibition decreased insulin‑induced KGN cell proliferation and apoptosis by targeting PCGF1. These data indicated that miR‑320a may serve as a potential diagnostic biomarker for PCOS.