Polycystic Ovary Syndrome Ovaries Research Articles

Ovarian ferroptosis induced by androgen is involved in pathogenesis of PCOS.

Does ovarian ferroptosis play an active role in the development of polycystic ovary syndrome (PCOS)? Increased ovarian ferroptosis was present in PCOS ovaries and the inhibition of ferroptosis with ferrostatin-1 (Fer-1) ameliorated polycystic ovary morphology and anovulation. Programmed cell death plays a fundamental role in ovarian follicle development. However, the types and mechanisms of cell death involved in the ovary are yet to be elucidated. Ferroptosis is a recently discovered iron-dependent programmed cell death. Impaired iron metabolism and cell death have been observed in women with PCOS, the main cause of anovulatory infertility. Additionally, previous studies reported that an abnormal expression of noncoding RNA may promote ferroptosis in immortalized ovarian granulosa cell lines. However, little is known about whether ovarian ferroptosis is increased in PCOS, and there is insufficient direct evidence for a role of ferroptosis in PCOS, and the underlying mechanism. Moreover, the effect of the inhibition of ferroptosis with Fer-1 in PCOS remains unclear. Ferroptosis was evaluated in human granulosa cells (hGCs) from non-PCOS (n = 6-16) and PCOS (n = 7-18) patients. The experimental study was completed in vitro using primary hGCs from women undergoing IVF. Improvements in PCOS indicators following ferroptosis inhibition with Fer-1 were investigated in a dehydroepiandrosterone (DHEA)-induced PCOS rat model (n = 8 per group). Ovarian ferroptosis was evaluated in the following ways: by detecting iron concentrations via ELISA and fluorescent probes; measuring malondialdehyde (MDA) concentrations via ELISA; assessing ferroptosis-related protein abundance with western blotting; observing mitochondrial morphology with transmission electron microscopy; and determining cell viability. Primary hGCs were collected from women undergoing IVF. They were treated with dihydrotestosterone (DHT) for 24 h. The effect of DHT on ferroptosis was examined in the presence or absence of small interfering RNA-mediated knockdown of the putative receptor coregulator for signaling molecules. The role of ovarian ferroptosis in PCOS progression was explored in vivo in rats. The DHEA-induced PCOS rat model was treated with the ferroptosis inhibitor, Fer-1, and the oocytes and metaphase II oocytes were counted after ovarian stimulation. Additionally, rats were treated with the ferroptosis inducer, RSL3, to further explore the effect of ferroptosis. The concentrations of testosterone, FSH, and LH were assessed. Increased ferroptosis was detected in the ovaries of patients with PCOS and in rats with DHEA-induced PCOS. Increased concentrations of Fe2+ (P < 0.05) and MDA (P < 0.05), and upregulated nuclear receptor coactivator 4 protein levels, and downregulated ferritin heavy chain 1 (FTH1) and glutathione peroxidase 4 (GPX4) proteins were observed in the hGCs in patients with PCOS and ovaries of PCOS rats (P < 0.05 versus control). DHT was shown to induce ferroptosis via activation of NOCA4-dependent ferritinophagy. The inhibition of ferroptosis with Fer-1 in rats ameliorated a cluster of PCOS traits including impaired glucose tolerance, irregular estrous cycles, reproductive hormone dysfunction, hyperandrogenism, polycystic ovaries, anovulation, and oocyte quality (P < 0.05). Treating rats with RSL3 resulted in polycystic ovaries and hyperandrogenism (P < 0.05). N/A. Although ovarian-targeted ferroptosis inhibition may be a more targeted treatment for PCOS, the underlying mechanisms in the cycle between ferroptosis and hyperandrogenism require further exploration. Additionally, since PCOS shows high heterogeneity, it is important to investigate whether ferroptosis increases are present in all patients with PCOS. Androgen-induced ovarian ferroptosis appears to play a role in the pathogenesis of PCOS, which potentially makes it a promising treatment target in PCOS. This study was supported by the National Key R&D Program of China (2023YFC2705500, 2023YFC2705505, 2019YFA0802604), National Natural Science Foundation of China (No. 82130046, 82320108009, 82101708, 82101747, and 82001517), Shanghai leading talent program, Innovative research team of high-level local universities in Shanghai (No. SHSMU-ZLCX20210201, No. SSMU-ZLCX20180401), Shanghai Jiaotong University School of Medicine, Affiliated Renji Hospital Clinical Research Innovation Cultivation Fund Program (RJPY-DZX-003) and Shanghai Municipal Education Commission-Gaofeng Clinical Medicine Grant Support (No. 20161413), Shanghai's Top Priority Research Center Construction Project (2023ZZ02002), and Three-Year Action Plan for Strengthening the Construction of the Public Health System in Shanghai (GWVI-11.1-36). The authors report no competing interests.

Dynamic changes in AI-based analysis of endometrial cellular composition: Analysis of PCOS and RIF endometrium

BackgroundThe human endometrium undergoes a monthly cycle of tissue growth and degeneration. During the mid-secretory phase, the endometrium establishes an optimal niche for embryo implantation by regulating cellular composition (e.g., epithelial and stromal cells) and differentiation. Impaired endometrial development observed in conditions such as polycystic ovary syndrome (PCOS) and recurrent implantation failure (RIF) contributes to infertility. Surprisingly, despite the importance of the endometrial lining properly developing prior to pregnancy, precise measures of endometrial cellular composition in these two infertility-associated conditions are entirely lacking. Additionally, current methods for measuring the epithelial and stromal area have limitations, including intra- and inter-observer variability and efficiency. MethodsWe utilized a deep-learning artificial intelligence (AI) model, created on a cloud-based platform and developed in our previous study. The AI model underwent training to segment both areas populated by epithelial and stromal endometrial cells. During the training step, a total of 28.36 mm2 areas were annotated, comprising 2.56 mm2 of epithelium and 24.87 mm2 of stroma. Two experienced pathologists validated the performance of the AI model. 73 endometrial samples from healthy control women were included in the sample set to establish cycle phase-dependent dynamics of the endometrial epithelial-to-stroma ratio from the proliferative (PE) to secretory (SE) phases. In addition, 91 samples from PCOS cases, accounting for the presence or absence of ovulation and representing all menstrual cycle phases, and 29 samples from RIF patients on day 5 after progesterone administration in the hormone replacement treatment cycle were also included and analyzed in terms of cellular composition. ResultsOur AI model exhibited reliable and reproducible performance in delineating epithelial and stromal compartments, achieving an accuracy of 92.40% and 99.23%, respectively. Moreover, the performance of the AI model was comparable to the pathologists’ assessment, with F1 scores exceeding 82% for the epithelium and >96% for the stroma. Next, we compared the endometrial epithelial-to-stromal ratio during the menstrual cycle in women with PCOS and in relation to endometrial receptivity status in RIF patients. The ovulatory PCOS endometrium exhibited epithelial cell proportions similar to those of control and healthy women’s samples in every cycle phase, from the PE to the late SE, correlating with progesterone levels (control SE, r2 = 0.64, FDR < 0.001; PCOS SE, r2 = 0.52, FDR < 0.001). The mid-SE endometrium showed the highest epithelial percentage compared to both the early and late SE endometrium in both healthy women and PCOS patients. Anovulatory PCOS cases showed epithelial cellular fractions comparable to those of PCOS cases in the PE (Anovulatory, 14.54%; PCOS PE, 15.56%, p = 1.00). We did not observe significant differences in the epithelial-to-stroma ratio in the hormone-induced endometrium in RIF patients with different receptivity statuses. ConclusionThe AI model rapidly and accurately identifies endometrial histology features by calculating areas occupied by epithelial and stromal cells. The AI model demonstrates changes in epithelial cellular proportions according to the menstrual cycle phase and reveals no changes in epithelial cellular proportions based on PCOS and RIF conditions. In conclusion, the AI model can potentially improve endometrial histology assessment by accelerating the analysis of the cellular composition of the tissue and by ensuring maximal objectivity for research and clinical purposes.

Metabolomics approach for the etiopathogenesis of polycystic ovarian syndrome

Polycystic ovary syndrome (PCOS) is a complex and multifaceted disorder that has been extensively studied from various angles, including genomics, metabolomics, and other multiomics approaches. This narrative review seeks to provide an updated understanding of PCOS, with a particular focus on metabolomics. PCOS encompasses four primary phenotypes: Classic PCOS with or without polycystic ovary morphology, ovulatory PCOS, and non-hyperandrogenic PCOS. Each phenotype exhibits distinct clinical and biochemical characteristics. Classic PCOS, characterized by hyperinsulinemia, insulin resistance (IR), and metabolic complications, has garnered significant attention in recent studies. In the pursuit of unraveling PCOS’s pathophysiology, researchers have embraced a multiomics approach, exploring the genome, epigenome, transcriptome, exosomes, proteome, and metabolome. The genomic landscape of PCOS includes genes related to androgen synthesis, regulation, insulin receptors, and growth factors, among others. Epigenetic mechanisms have also been investigated, revealing the roles of non-coding RNA, histone acetylation, and deacetylation. Transcriptomic biomarkers and mitochondrial RNAs targets, such as miR-9 and miR-32, have been explored in granulosa cells of PCOS patients. Metabolomics studies have identified various biomarkers and metabolic pathways that differentiate PCOS from healthy individuals. A diverse range of alterations in carbohydrate and lipid metabolism has been observed, shedding light on the underpinnings of PCOS-related metabolic dysfunction. These studies have uncovered numerous biomolecules with potential relevance to PCOS, including kininogen 1, cytoketatin 9, and many others. In addition, the intricate relationships between IR, inflammation, and metabolic factors have been elucidated. Recent research extends beyond high-end research centers, even using urine samples to identify metabolomic markers for PCOS. This approach has led to the discovery of distinct biomarker panels related to metabolic pathways that differentiate PCOS from healthy subjects. In summary, PCOS is a complex and heterogeneous condition, with multiomics studies offering promising avenues for a deeper understanding of its pathophysiology. These studies may ultimately provide valuable insights that can revolutionize the diagnosis and treatment of PCOS, ending a centurylong scientific expedition with a crescendo of discoveries.

#43 : Expression of miRNA-21-3p in Polycystic Ovary Syndrome and its Related Functions

Background and Aims: PCOS is an endocrine-related disease related to abnormal folliculogenesis and is a leading cause of infertility worldwide. Inhibition of GCs proliferation and increased GCs apoptosis have been identified as the major factors in aberrant follicle maturation. Recently, the roles of miRNAs were identified in polycystic ovary syndrome (PCOS). For instance, miR-21-3p was identified as an important miRNA regulating autophagy of BGCs. Besides, studies showed that AMSC-EXOs can rescue the polycystic phenotype and metabolic dysfunction in PCOS ovaries by transferring miR-21-5p to the livers of rats with PCOS, finally improve reproduction. However, there is no research clarified the association between miRNA-21-3p and GCs function to date. Method: In the present study, letrozole was used to establish the PCOS rat model. Separate the ovary and then miRNA-21-3p expression will be analyzed by RT-PCR. Primary rat ovarian GCs were isolated and incubated and transfected with mimic or inhibitor of miRNA-21-3p, CCK8 kit was used to detect cell viability, PR-PCR was used to detect cell proliferation and apoptosis related gene expression. Finally, flowcytometry was used to test apoptosis rate of Gcs. Results: The expression of miR-21-3p in the ovaries of PCOS rats was significantly increased compared with that in control rats. Primary rat ovarian GCs were isolated and incubated successfully, and then CCK-8 experiment showed that GCs viability was significantly decreased when miR-21-3p was overexpressed. Additionally, the miR-21-3p mimic significantly decreased the expression of PCNA, Bcl-2, cell cycle-related genes (CDK2, CCNE1) and increased the expression of Bax and caspase3 in primary rat ovarian GCs. By contrast, transfected primary rat ovarian GCs using the miR-21-3p inhibitor significantly upregulated the expression of PCNA, Bcl-2, cell cycle-regulated genes (CDK2, CCNE1) and reduced the expression of Bax and caspase3. Finally, flow cytometry analysis demonstrated that overexpression of miR-21-3p significantly increased the apoptosis rate of GCs. By contrast, the suppression of miR-21-3p resulted in a significant decrease in the apoptosis rate of GCs. Conclusion: In PCOS rat model, miRNA-21-3p was proved significantly increased compared with that in control rats. Which is participant in the development of PCOS by regulating the function of granular cells.

Down-regulation of Lon protease 1 lysine crotonylation aggravates mitochondrial dysfunction in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a prevalent reproductive endocrine disorder, with metabolic abnormalities and ovulation disorders. The post-translational modifications (PTMs) are functionally relevant and strengthen the link between metabolism and cellular functions. Lysine crotonylation is a newly identified PTM, the function of which in PCOS has not yet been reported. To explore the molecular mechanisms of crotonylation involved in the abnormalities of metabolic homeostasis and oocyte maturation in PCOS, by using liquid chromatography-tandem mass spectrometry analysis, we constructed a comprehensive map of crotonylation modifications in ovarian tissue of PCOS-like mouse model established by dehydroepiandrosterone induction. The crotonylation levels of proteins involved in metabolic processes were significantly decreased in PCOS ovaries compared to control samples. Further investigation showed that decrotonylation of Lon protease 1 (LONP1) at lysine 390 was associated with mitochondrial dysfunction in PCOS. Moreover, LONP1 crotonylation levels in PCOS were correlated with ovarian tissue oxidative stress levels, androgen levels, and oocyte development. Consistently, down-regulation of LONP1 and LONP1 crotonylation levels were also observed in the blood samples of PCOS patients. Collectively, our study revealed a mechanism by which the decrotonylation of LONP1 may attenuate its activity and alter follicular microenvironment to affect oocyte maturation in PCOS.

Bromodomain-containing protein 4 activates androgen receptor transcription and promotes ovarian fibrosis in PCOS

Polycystic ovary syndrome (PCOS) is an endocrine disorder and the main cause of anovulatory infertility, in which persistent activation of androgen receptor (AR) due to aberrant acetylation modifications of transcription is a potential trigger; however, the precise mechanisms of AR activation are poorly understood. In this study, AR activation in dehydroepiandrosterone- and letrozole-induced rat PCOS ovaries coincided with a marked increase of a chromatin acetylation "reader" BRD4. Further bioinformatic analysis showed that the AR promoter contained highly conserved binding motifs of BRD4 and HIF-1α. BRD4 and HIF-1α inducibly bound to the histone 3/4 acetylation-modified AR promoter, while administration of a BRD4-selective inhibitor JQ1 reduced the binding and AR transcription and improved the adverse expression of the core fibrotic mediators in PCOS ovaries and DHT-treated granulosa cells. Our data indicate that BRD4 upregulation and the resultant AR transcriptional activation constitute an important regulatory pathway that promotes ovarian fibrosis in PCOS.

Protocatechuic acid alleviates polycystic ovary syndrome symptoms in mice by PI3K signaling in granulosa cells to relieve ROS pressure and apoptosis

Background Polycystic ovary syndrome (PCOS) is a complicated gynecological endocrine disease that occurs in women of childbearing age. Protocatechuic acid is a phenol-rich compound derived from herbs and owns vital functions in numerous diseases. Howbeit, protocatechuic acid’s impact on PCOS is unknown. Methods A combination of in vivo and in vitro models was examined in this study. C57BL/6 mice were injected subcutaneously daily with dehydroepiandrosterone to establish a PCOS mouse model, and protocatechuic acid was intraperitoneally injected into PCOS mice. Granulosa cells of PCOS ovaries were also isolated. The function of protocatechuic acid was appraised using enzyme-linked immunosorbent assay, hematoxylin–eosin staining, reactive oxygen species (ROS) and LC3 levels analysis, flow cytometry, quantitative real-time PCR, and western blot. Meanwhile, the mechanism of protocatechuic acid was assessed with a series of molecular experiments. Results Protocatechuic acid owned no apparent toxic effect on mice. Functionally, protocatechuic acid owned a function of mitigating PCOS in vivo. Meanwhile, protocatechuic acid repressed ROS, autophagy, and apoptosis of PCOS ovarian granulosa cells in vitro. Mechanistically, rescue assays elucidated that the protective function of protocatechuic acid against PCOS was interrelated to the activation of the PI3K/AKT/mTOR axis. Conclusion Protocatechuic acid alleviated PCOS symptoms in mice through PI3K signaling in granulosa cells to reduce ROS levels and apoptosis.

P-625 Artificial intelligence-enabled analysis of endometrial CD138 positive plasma cells in infertility-associated conditions; Polycystic ovary syndrome (PCOS) and recurrent implantation failure (RIF)

Abstract Study question Can artificial intelligence (AI) algorithm identify CD138+ cells in the endometrial stroma, and is the expression altered in PCOS and RIF? Summary answer CD138+ cell detection by AI shows high accuracy. The cell count in controls was comparable to that in PCOS cases but differed in RIF patients. What is known already Immunohistochemical (IHC) staining for CD138+ plasma cells has been applied to detect endometrial inflammation; however, due to a lack of cost-effective and reliable tools, there is no golden standard for CD138+ analysis. PCOS is a common endocrine disorder characterized by irregular menstrual cycles and hyperandrogenism. The women are also affected by systemic low-grade inflammation, and the PCOS endometrium shows dysregulated immune profile. RIF is one of the main causes of a low IVF success rate, and chronic endometrial inflammation has been detected in women with RIF. To date, AI technology has been applied in the diagnostics of various clinical conditions. Study design, size, duration In this study, AI analysis was done for CD138+ cells in a total of 193 endometrial biopsy samples: 73 were obtained from healthy and fertile controls, 91 from women with PCOS (ovulatory samples; n = 78, anovulatory samples; n = 13, Rotterdam criteria), and 29 samples from RIF patients on day five after the initiation of P4 administration. Participants/materials, setting, methods The biopsy samples were collected by suction curette (Pipelle), and endometrial receptivity was assessed by gene expression profiling (beREADY test). The tissues were stained with CD138 antibody, and the AI algorithm, AITAH, was trained with 150 whole slide images (WSIs) to segment epithelium and stroma and segregate CD138- and CD138+ cells in the stroma. AITAH was validated by three external pathologists. The cell counts were compared according to cycle phases, ovulatory status, and endometrial receptivity. Main results and the role of chance The performance of AITHA was excellent, as the final training error for CD138+ cells was 3.23%, and the decisions for the cells between pathologists and AITAH were in complete agreement. The CD138+ cell percentages were higher in the proliferative phase (PE) endometrial tissue than in the ones obtained in the secretory phase (PCOS, p &amp;lt;0.001 or control, p &amp;lt;0.001), regardless of PCOS diagnosis. Similar CD138+ cell percentages were observed between PE endometrium and anovulatory endometrium in PCOS cases (p &amp;lt;0.001). The count of CD138+ plasma cells was negatively correlated with the endometrial thickness (PCOS, p =0.001 or control, p =0.009) and P4 (PCOS, p =0.001 or control, p =0.003), and positive correlations between the cell counts and gonadotropin levels were observed in ovulatory PCOS samples (FSH, p =0.001 and LH, p =0.031). More CD138+ cells were seen in RIF patients compared to the controls at the receptive stage (p = 0.046); however, there was no difference between RIF samples of three different receptivity statuses (p =0.426). Limitations, reasons for caution Further validation for AITAH using confirmed chronic endometritis cases as positive controls is required. And further analysis using control samples obtained during the hormone replacement treatment is required. The small number of anovulatory PCOS subjects may explain the absence of correlations between the CD138+ plasma cell counts and clinical characteristics. Wider implications of the findings AI-enabled analysis tools could improve histological examination of endometrial immune cells by analyzing high amounts of whole tissue slides within a short time with a high pixel resolution, thus overcoming many current impediments in manual microscopic evaluation. Trial registration number NA

P-641 Clinical significance of differential expression of miRNA642-a/miRNA5739/miRNA3648 in the follicular microenvironment in patients with polycystic ovary syndrome

Abstract Study question What is the relationship between differential expression of miRNA in the follicular microenvironment,follicular fluid hormone and oocyte quality in polycystic ovary syndrome (PCOS) patients. Summary answer In PCOS patients, differential expression of miRNA in the follicular microenvironment is associated with abnormal hormone levels in the follicular fluid and oocyte quality. What is known already PCOS has abnormalities in local ovarian morphology and function, and studies have shown that the expression of local miRNA in the ovaries will be accompanied by dynamic changes in follicular recruitment, oocyte growth, follicular microenvironment changes, etc., which may affect the quality of oocytes. Study design, size, duration 19 patients with PCOS (PCOS group) and 17 patients with infertility due to male factors alone (control group) aged 22-35 years was included in the study. The main results included differential expression of miRNA642-a/miRNA5739/miRNA3648, MII egg rate, high-quality embryo rate, testosterone and insulin levels in follicular fluid. Participants/materials, setting, methods From April 2017 to January 2019, 19 PCOS patients (PCOS group) and 17 patients (control group) who underwent Intracytoplasmic sperm injection-embryo transfer at the Reproductive Center of the Affiliated Hospital of Shandong University of Traditional Chinese Medicine were collected. RT-PCR was used to detect the differential expression levels of miRNA642-a/miRNA5739/miRNA3648 in follicular fluid and granule cells in PCOS group and control group, and Chemiluminescence was used to detect follicular fluid T and insulin levels. Main results and the role of chance The levels of testosterone and insulin in follicular fluid in PCOS group were significantly higher than those in the control group, and the difference was statistically significant (P &amp;lt; 0.05). The MII egg rate and optimal embryo rate in the PCOS group were both lower than those in the control group, and the difference was significant (P &amp;lt; 0.05). The expression level of miR-642a-3p in follicular fluid and granule cells was consistent, and the PCOS group was lower than that in the control group. The difference was statistically significant (P &amp;lt; 0.05). The expression levels of miRNA-5739 and miRNA-3648 in follicular fluid were higher than those in the control group, but the expression levels in granule cells were lower than those in the control group, and the difference was significant (P &amp;lt; 0.05). Correlation analysis showed that the expression levels of miRNA-3648 and miRNA-5739 in follicular fluid were closely related to the normal fertilization rate of PCOS patients, and the expression levels of miRNA-5739 were closely related to the optimal embryo rate of PCOS patients. The expression levels of miRNA-642a-3p, miRNA-5739 and miRNA-3648 in granule cells were closely related to the MII egg rate and 2PN cleavage rate of PCOS patients. Limitations, reasons for caution MiRNAs participate in follicle development and maturation and hormone secretion through multiple signaling pathways. We only examined miRNA-642a-3p, miRNA-5739, miRNA-3648 in follicular fluid and granule cells, and did not detect the exosomal miRNAs. Dynamic changes in miRNA in the follicular microenvironment are not fully revealed. Wider implications of the findings Our results further demonstrate that the abnormal expression of miRNA in follicular fluid and granule cells in women with polycystic ovary syndrome (PCOS) is closely related to the local abnormal hormone secretion and follicle development of PCOS ovaries. Trial registration number not applicable

Determining the age group-based cut-off values of serum anti-Mullerian hormone concentrations to diagnose polycystic ovary syndrome

Objective To determine the cut-off values of the serum anti-Müllerian hormone (AMH) concentration for different age groups (21–25, 26–30, 31–35 years) to diagnose polycystic ovary syndrome (PCOS). Methods In total, 187 women aged 21–35 years were included in this descriptive study. Patients diagnosed with PCOS according to the Rotterdam Criteria formed the PCOS group (n = 93), whereas those without symptoms related to PCOS formed the control group (n = 94). Follicular phase serum hormone concentrations were evaluated during the endocrinological assessment of patients with PCOS. Serum levels of estradiol, follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin, total testosterone, dehydroepiandrosterone sulfate, sex hormone-binding globulin, androstenedione, and AMH were measured. The free androgen index and LH/FSH ratio were calculated. Cut-off values of serum AMH concentrations for the age groups were measured using receiver operating characteristic curve analysis. Results The prevalence rates of frank, ovulatory, normoandrogenic, and non-polycystic ovary PCOS were 69.9, 10.8, 10.8, and 8.6%, respectively. Serum AMH concentrations >5.56 ng/mL were associated with PCOS in the 21–25-year-old group. The cut-off value was 4.01 ng/mL in the 26–30-year-old group, whereas it was 3.42 ng/mL in the oldest age group. The correlation between the antral follicle count (AFC) and serum AMH level was strong for each age group. Conclusions The serum AMH concentration is a valuable parameter for assessing patients with symptoms indicative of PCOS. We recommend measuring serum AMH levels to support the diagnosis or to use them instead of the AFC for the Rotterdam criteria.

Long-Lasting Effects of Spironolactone after its Withdrawal in Patients with Hyperandrogenic Skin Disorders.

Hyperandrogenic skin disorders, such as hirsutism, acne and alopecia, affect approximately 10-20% of women of reproductive age, reducing quality of life and causing psychological impairment. Spironolactone is a commonly used antiandrogen, especially in women who are not sexually active or have contraindications to hormonal contraceptives. The aim of this study was to evaluate the effects of spironolactone, especially after its withdrawal, in patients with hyperandrogenic skin disorders. Retrospective analysis of 63 women with hyperandrogenic skin symptoms due to polycystic ovary syndrome (PCOS), treated with spironolactone for at least 6 months as first-line treatment. After a mean time of treatment of 25.7 months, all patients reported a significant improvement in hyperandrogenic skin disorders; only 5 patients were dissatisfied and required the addition of an oral contraceptive. The therapy was well tolerated and the most frequent side-effect was intermestrual bleeding in 68.2% of cases, affecting mainly classic PCOS phenotype. Thirthyeight patients showed prolonged effects 33.7 months after spironolactone withdrawal, whereas 20 relapsed 17.5 months after discontinuation. No significant difference in clinical and biochemical parameters was observed between these two groups both at baseline and after spironolactone treatment. Ovulatory PCOS patients were treated for a shorter time and reported earlier relapse than classic PCOS patients. Spironolactone is an effective and safe treatment for hyperandrogenic skin disorders, showing long-lasting effects even several months after its discontinuation.

ALG2 inhibits the epithelial-to-mesenchymal transition and stemness of ovarian granulosa cells through the Wnt/β-catenin signaling pathway in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is an endocrine disorder that affects fertility in women of reproductive age, and a leading cause of anovulatory infertility. Ovarian granulosa cells are a major functional cell type in the ovary that undergo epithelial-to-mesenchymal transition (EMT) to initiate ovulation. Protein glycosylation, catalyzed by specific glycosyltransferases, has been implicated in reproductive events, such as embryo implantation, endometrial receptivity, and decidualization, etc. However, the relationship between glycosylation and EMT-mediated ovulation in PCOS is not well understood. To clarify the role of cobalt chloride (CoCl2) and α-1,3/1,6-mannosyltransferase (ALG2) in PCOS, transwell assay, Real-time PCR, Western blot, immunofluorescence, and sphere formation assay were applied to assess cell migration, invasion, EMT, and stemness of ovarian granulosa cells. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the serum level of ALG2 in PCOS patients. We found that CoCl2 promoted the migration, invasion, EMT, and stemness of ovarian granulosa cells by downregulating the expression of ALG2. Upregulation of ALG2 rescued the effects of CoCl2 partially, and inhibited the EMT and stemness of ovarian granulosa cells by inactivating the Wnt/β-catenin signaling pathway. Moreover, the serum level of ALG2 was increased in patients with PCOS. Elevated ALG2, in combination with testosterone, represented better diagnostic value for PCOS as a multimarker than ALG2 or testosterone alone as a single marker. Thus, ALG2, downregulated by CoCl2, was increased in PCOS patients which inhibited the EMT and stemness of ovarian granulosa cells by deactivating the Wnt/β-catenin signaling pathway. The combination of ALG2 and testosterone might thus act as a novel but promising biomarker for PCOS detection.

Therapeutic potentials of the caffeine in polycystic ovary syndrome in a rat model: Via modulation of proinflammatory cytokines and antioxidant activity.

Recent studies have shown that polycystic ovary syndrome (PCOS) affects about 6% of women worldwide. It is associated with reproductive and metabolic dysfunction. Caffeine is naturally found in tea, cocoa, and coffee. It has been shown that caffeine can change hormonal profiles, stimulate ovulation, and enhance fertility. Therefore, in this study, the effects of caffeine on rats with PCOS were investigated. For this purpose, 40 female rats were divided into five groups: (1) control group (without any intervention), (2) sham group (administration of olive oil as a caffeine solvent), (3) PCOS group (injection of 2 mg of estradiol valerate for each rat), (4) caffeine group (administration of 37.5 mg/kg caffeine for each rat), and (5) PCOS + caffeine group. After 21 days of treatment, the ovaries of rats were removed and prepared for further evaluations, including hematoxylin and eosin staining, TUNEL assay, real-time PCR, and biochemical analysis. Administration of caffeine in PCOS mice considerably reduced both the volume of the ovary (P < 0.05) and follicular clusters (P < 0.01). However, superoxide dismutase and glutathione peroxidase were dramatically active in the PCOS + caffeine group compared to others (P < 0.05). Besides, caffeine treatment in PCOS mice led to Bax reduction and increased Bcl-2 expression. On the other hand, the expression of IL-6 and TNF-α in PCOS + caffeine group was high compared to other groups. We found that caffeine can reduce apoptosis and inflammation in PCOS ovaries and enhance the unpleasant symptoms of PCOS.

Resveratrol improves follicular development of PCOS rats via regulating glycolysis pathway and targeting SIRT1

Polycystic ovary syndrome (PCOS) is a disease characterized by metabolic disorders. This study aimed to examine the effects of resveratrol treatment on ovulation in the PCOS rat model. Quantitative real-time PCR and immunohistochemistry were used to determine the mRNA and protein expression levels. TNUEL assay was used to evaluate cell apoptosis in ovary. The metabolites were evaluated by liquid chromatography with tandem mass spectrometry. Resveratrol alleviated disrupted estrous cycle and improved granular cell layers, and reversed the decreased proliferation and increased cell apoptosis of granulosa cells in the ovarian tissues of PCOS rats. Resveratrol restored the changes in the mRNA expression levels in the rate-limiting genes of glycolysis in the PCOS ovary. The expression of lactate dehydrogenase A (LDH-A), pyruvate kinase isozyme M2 (PKM2), and sirtuin 1 (SIRT1) was significantly downregulated in ovarian tissues of the PCOS rats; while the resveratrol treatment significantly increased the expression of LDH-A, PKM2, and SIRT1 in the ovarian tissues of PCOS rats. Collectively, the protective effects of resveratrol in the PCOS rats may be associated with the regulation of glycolysis-related mediators including PKM2, LDH-A, and SIRT1. Resveratrol may represent a good candidate in alleviating the development of PCOS.

The ADMA–DDAH1 axis in ovarian apoptosis of polycystic ovary syndrome

Dimethylarginine dimethylaminohydrolase 1 (DDAH1) mainly degrades asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor. Emerging evidence suggested that plasma ADMA is accumulated in patients with polycystic ovary syndrome (PCOS). However, ADMA–DDAH1 involvement in PCOS pathogenesis is unclear. Here, we used dehydroepiandrosterone (DHEA)-induced PCOS rats and the ovarian granulosa cell line KGN to investigate the effect of the ADMA–DDAH1 pathway on ovarian apoptosis. Moreover, we also quantified the ADMA levels and redox status in human serum specimens, Sprague Dawley rats and KGN cells to investigate the effect of ADMA–DDAH1 on redox status and ovarian apoptosis in PCOS. We enrolled 19 women with PCOS and 17 healthy women (controls) in this study. The women with PCOS had increased serum ADMA levels and decreased glutathione peroxidase (GSH-PX) compared with the controls. In Sprague Dawley rats, 21-day DHEA treatment established PCOS and the rat contained higher ADMA levels in serum and lower DDAH1 expression in ovaries. Moreover, the PCOS rat serum and ovaries exhibited increased levels of the oxidative stress marker malondialdehyde (MDA). ADMA treatment of the KGN cells induced reactive oxygen species accumulation and led to apoptosis. Contrastingly, overexpressing DDAH1 in the KGN cells significantly decreased ADMA levels, enhanced cell viability, and inhibited oxidative stress, while the effect was inverse in DDAH1 knockdown cells. Overall, our results demonstrated that PCOS involves elevated ADMA levels and redox imbalance. The ADMA–DDAH1 pathway exerted a marked effect on oxidative stress and ovarian apoptosis in PCOS. Our findings suggested that strategies for increasing DDAH1 activity in ovarian cells may provide a novel approach for ameliorating PCOS.

An extended machine learning technique for polycystic ovary syndrome detection using ovary ultrasound image

Polycystic ovary syndrome (PCOS) is the most prevalent endocrinological abnormality and one of the primary causes of anovulatory infertility in women globally. The detection of multiple cysts using ovary ultrasonograpgy (USG) scans is one of the most reliable approach for making an accurate diagnosis of PCOS and creating an appropriate treatment plan to heal the patients with this syndrome. Instead of depending on error-prone manual identification, an intelligent computer-aided cyst detection system can be a viable approach. Therefore, in this research, an extended machine learning classification technique for PCOS prediction has been proposed, trained and tested over 594 ovary USG images; where the Convolutional Neural Network (CNN) incorporating different state-of-the-art techniques and transfer learning has been employed for feature extraction from the images; and then stacking ensemble machine learning technique using conventional models as base learners and bagging or boosting ensemble model as meta-learner have been used on that reduced feature set to classify between PCOS and non-PCOS ovaries. The proposed technique significantly enhances the accuracy while also reducing training execution time comparing with the other existing ML based techniques. Again, following the proposed extended technique, the best performing results are obtained by incorporating the “VGGNet16” pre-trained model with CNN architecture as feature extractor and then stacking ensemble model with the meta-learner being “XGBoost” model as image classifier with an accuracy of 99.89% for classification.

Mesenchymal stem cells-derived exosomes as a promising new approach for the treatment of infertility caused by polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a multifactorial metabolic and most common endocrine disorder that its prevalence, depending on different methods of evaluating PCOS traits, varies from 4% to 21%. Chronic low-grade inflammation and irregular apoptosis of granulosa cells play a crucial role in the pathogenesis of PCOS infertility. Mesenchymal stem cells (MSCs)-derived exosomes and extracellular vesicles (EVs) are lipid bilayer complexes that act as a means of intercellular transferring of proteins, lipids, DNA and different types of RNAs. It seems that this nanoparticles have therapeutic effects on the PCOS ovary such as regulating immunity response, anti-inflammatory (local and systemic) and suppress of granulosa cells (GCs) apoptosis. Although there are few studies demonstrating the effects of exosomes on PCOS and their exact mechanisms is still unknown, in the present study we reviewed the available studies of the functions of MSC-derived exosome, EVs and secretome on apoptosis of granulosa cells and inflammation in the ovary. Therefore, the novel cell-free therapeutic approaches for PCOS were suggested in this study.

HDAC5 inhibits ovarian angiogenesis in dehydroepiandrosterone-induced mouse model of polycystic ovary syndrome.

Abnormal ovarian angiogenesis is a common feature of polycystic ovary syndrome (PCOS), a typical endocrine disorder affecting women of reproductive age. Histone deacetylase 5 (HDAC5) has been documented as a suppressor of angiogenesis. The aim of this study was to explore the effect of HDAC5 on ovarian angiogenesis in a PCOS mouse model. PCOS was induced in female C57BL/6 mice by 20-day administration of dehydroepiandrosterone (DHEA). HDAC5 was over-expressed in PCOS mice by corresponding adenovirus injection. In total, 120 mice were used in this study. Western-blotting, real-time PCR, hematoxylin and eosin staining, enzyme-linked immunosorbent assay (ELISA), immunohistochemical staining, flow cytometry, and co-immunoprecipitation were respectively used to evaluate the effect of HDAC5 on PCOS mice. PCOS ovaries showed a compensatory increase in HDAC5 expression, while HDAC5 over-expression alleviated abnormalities in ovarian morphology and serum hormone levels after PCOS modeling. HDAC5 inhibited ovarian angiogenesis in PCOS mice by regulating angiogenesis-related factors, such as VEGFA, platelet-derived growth factors B and D (PDGFB/D), and angiopoietins 1 and 2 (ANGPT1/2) and CD31. HDAC5 over-expression decreased levels of reactive oxygen species (ROS) and malondialdehyde, while promoting activities of catalase and superoxide dismutase in ovaries of PCOS mice, suggesting its suppressive effects on oxidative stress, an inducer of uncontrolled angiogenesis. Moreover, HDAC5 suppressed activation of angiogenesis-related HIF-1α/VEGFA/VEGFR2 signaling in PCOS ovaries partly via inhibiting VEGFR2 acetylation. This study reveals the protective role of HDAC5 in PCOS by inhibiting ovarian angiogenesis and provides a molecular candidate for PCOS therapy in the future.

Adipose mesenchymal stem cell–derived exosomal microRNAs ameliorate polycystic ovary syndrome by protecting against metabolic disturbances

Polycystic ovary syndrome (PCOS) is the most common endocrine and metabolic disorder in women of childbearing age. Adipose mesenchymal stem cells (AMSCs) secrete cytokines involved in the regulation of metabolism and immunity. However, it remains unclear whether exosomes secreted by AMSCs (AMSC-EXOs) can rescue the polycystic phenotype and metabolic dysfunction in PCOS ovaries. Here, we show that AMSC-EXOs can protect against metabolic disturbances, ameliorate ovarian polycystic, and improve fertility in a rat model of PCOS. AMSC-EXOs inhibited the expression of B-cell translocation gene 2 by transferring miR-21-5p to the livers of rats with PCOS, thus activating the IRS1/AKT pathway and increasing hepatic metabolism. The role of AMSC-EXOs in transferring miRNAs to the liver to improve metabolic dysfunction in PCOS and reproduction by rescuing a non-coding RNA pathway was also discovered. This study provides a theoretical basis for the use of rat adipose stem cells and their secreted exosomes to treat PCOS.

Roles of HIF-1α/BNIP3 mediated mitophagy in mitochondrial dysfunction of letrozole-induced PCOS rats.

Mitochondrial dysfunction plays a crucial role in the pathological physiology of polycystic ovary syndrome (PCOS). Mitochondrial quality control system is vital to maintaining mitochondrial function, includes mitochondrial biosynthesis, dynamics and mitophagy. While mitophagy as a specific autophagy, plays an important role in the mitochondrial quality control system and is mediated by some signaling pathways to eliminate the excessive production of reactive oxygen species (ROS), such as hypoxia-inducible factor (HIF)-1α/B-cell lymphoma-2 adenovirus E1B 19kDa interacting protein 3 (BNIP3). Our previous studies have found that excessive production of ROS and the decreased expression of HIF-1α in the ovaries of PCOS rats. Thus, we hypothesized that excessive ROS leads to mitochondrial dysfunction, attenuates HIF-1α/BNIP3-mediated mitophagy in the ovaries of PCOS rats, and further reduces the mitophagic defense. Firstly, the oxidative stress status was detected and found excessive ROS damages ovarian tissue in PCOS rats. Secondly, the marker proteins of mitochondrial biosynthesis/dynamics and amount were examined and found that their expression levels were abnormal, which showed that the abnormal mitochondrial quality control system leads to accumulate the excess or damaged mitochondria in PCOS ovaries. Finally, we detected the HIF-1α/BNIP3 pathway and found HIF-1α-mediated mitophagy is impaired in the ovaries of PCOS rats. Together, these results clearly demonstrated excessive ROS causes mitochondrial dysfunction via the abnormal mitochondrial quality control system, and attenuates HIF-1α/BNIP3-mediated mitophagic defense in the granulosa cells of PCOS rats, which will provide a new direction for further understanding the role of HIF-1α in the molecular mechanism of mitochondrial dysfunction in PCOS ovaries.