Anti-Müllerian Hormone: A Molecular Key to Unlocking Polycystic Ovary Syndrome?

Anti-Müllerian hormone (AMH) is an important component within androgen receptor (AR)-regulated pathways governing the hyperandrogenic origin of polycystic ovary syndrome (PCOS). In women with PCOS, granulosa cell AMH overexpression in developing ovarian follicles contributes to elevated circulating AMH levels beginning at birth and continuing in adolescent daughters of PCOS women. A 6 to 7% incidence among PCOS women of gene variants coding for AMH or its receptor, AMHR2, suggests genetic contributions to AMH-related pathogenesis. Discrete gestational AMH administration to pregnant mice induces hypergonadotropic hyperandrogenic, PCOS-like female offspring with high circulating AMH levels that persist over three generations, suggesting epigenetic contributions to PCOS through developmental programming. Moreover, adult-onset, selective hyperactivation of hypothalamic neurons expressing gonadotropin-releasing hormone (GnRH) induces hypergonadotropic hyperandrogenism and PCOS-like traits in female mice. Both gestational and adult AMH inductions of PCOS-like traits are prevented by GnRH antagonist coadministration, implicating luteinizing hormone-dependent ovarian theca cell testosterone (T) action, mediated through the AR in AMH-induced pathogenesis. Interestingly, gestational or peripubertal exogenous T or dihydrotestosterone induction of PCOS-like traits in female mice, rats, sheep, and monkeys fails to elicit ovarian AMH hypersecretion; thus, AMH excess per se may lead to a distinct pathogenic contribution to hyperandrogenic PCOS origins.

Role of Branched-Chain Amino Acids in Metabolic Changes of Polycystic Ovary Syndrome.

Importance: Polycystic ovary syndrome (PCOS) is a common endocrine syndrome with multiple causes and polymorphic clinical manifestations, which is one of the important causes of menstrual disorders in women of childbearing age. It has been found that branched-chain amino acids (BCAAs), a class of essential amino acids that cannot be synthesized by the human body, play a significant role in the metabolic changes of PCOS, which may be involved in the pathogenesis of PCOS. Objective: The purpose of this review is to summarize the relevance between BCAAs and metabolic abnormalities in PCOS and to explore their possible mechanisms. Evidence Acquisition: The evidence is mainly obtained by reviewing the literature on PubMed related to PCOS, BCAAs, and related metabolic abnormalities and conducting summary analysis. Results: The metabolism of BCAAs can affect the homeostasis of glucose metabolism, possibly by disrupting the balance of gut microbiota, activating mTORC1 targets, producing mitochondrial toxic metabolites, and increasing the expression of proinflammatory genes. The correlation between obesity and BCAAs in PCOS patients may be related to the gene expression of BCAA metabolism-related enzymes in adipose tissue. The association between BCAA metabolic changes and nonalcoholic fatty liver disease in PCOS patients has not been fully clarified, which may be related to the lipid accumulation caused by BCAAs. At present, it is believed that hyperandrogenism in patients with PCOS is not related to BCAAs. However, through the study of changes in BCAA metabolism in prostate cancer caused by hyperandrogenism, we speculate that the relationship between BCAAs and hyperandrogenism may be mediated by mTORC1 and amino acid transporters. Conclusions and Relevance: Review of prior articles reveals that BCAAs may be related to insulin resistance, obesity, nonalcoholic fatty liver, and hyperandrogenism in PCOS patients, and its mechanisms are complex, diverse, and interrelated. This review also discussed the mechanism of BCAAs and these metabolic disorders in non-PCOS patients, which may provide some help for future research.

Artemisinins ameliorate polycystic ovarian syndrome by mediating LONP1-CYP11A1 interaction.

Polycystic ovary syndrome (PCOS), a prevalent reproductive disorder in women of reproductive age, features androgen excess, ovulatory dysfunction, and polycystic ovaries. Despite its high prevalence, specific pharmacologic intervention for PCOS is challenging. In this study, we identified artemisinins as anti-PCOS agents. Our finding demonstrated the efficacy of artemisinin derivatives in alleviating PCOS symptoms in both rodent models and human patients, curbing hyperandrogenemia through suppression of ovarian androgen synthesis. Artemisinins promoted cytochrome P450 family 11 subfamily A member 1 (CYP11A1) protein degradation to block androgen overproduction. Mechanistically, artemisinins directly targeted lon peptidase 1 (LONP1), enhanced LONP1-CYP11A1 interaction, and facilitated LONP1-catalyzed CYP11A1 degradation. Overexpression of LONP1 replicated the androgen-lowering effect of artemisinins. Our data suggest that artemisinin application is a promising approach for treating PCOS and highlight the crucial role of the LONP1-CYP11A1 interaction in controlling hyperandrogenism and PCOS occurrence.

Apelin receptor modulation mitigates letrozole-induced polycystic ovarian pathogenesis in mice.

AIMS: Polycystic ovarian syndrome (PCOS) is one of the most common (about 5-20%) reproductive disorders in women of reproductive age; it is characterized by polycystic ovaries, hyperandrogenism, and oligo/ anovulation. The levels and expression of ovarian adipokines are deregulated in the PCOS. Apelin is an adipokine that acts through its receptor (APJ) and is known to express in the various tissues including the ovary. It has also been suggested that apelin and APJ could be targeted as therapeutic adjuncts for the management of PCOS. However, no study has been conducted on the management of PCOS by targeting the apelin system. Thus, we aimed to evaluate its impact on combating PCOS-associated ovarian pathogenesis. METHODS: The current work employed a letrozole-induced-hyperandrogenism PCOS-like mice model to investigate the effects of apelin13 and APJ, antagonist ML221. The PCOS model was induced by oral administration of letrozole (1 mg/kg) for 21 days. A total of four experimental groups were made, control, PCOS control, PCOS + aplein13, and PCOS + ML221. The treatment of apelin13 and ML221 was given from day 22 for two weeks. KEY FINDINGS: The letrozole-induced PCOS-like features such as hyperandrogenism, cystic follicle, decreased corpus luteum, elevated levels of LH/FSH ratio, and up-regulation of ovarian AR expression were ameliorated by apelin13 and ML221 treatment. However, the PCOS-augmented oxidative stress and apoptosis were suppressed by apelin 13 treatments only. ML221 treatment still showed elevated oxidative stress and stimulated apoptosis as reflected by decreased antioxidant enzymes and increased active caspase3 and Bax expression. The expression of ERs was elevated in all groups except control. Furthermore, the PCOS model showed elevated expression of APJ and apelin13 treatment down-regulated its own receptor. Overall, observing the ovarian histology, corpus luteum formation, and decreased androgen levels by both apelin13 and ML221 showed ameliorative effects on the cystic ovary. SIGNIFICANCE: Despite the similar morphological observation of ovarian histology, apelin13 and ML221 exhibited opposite effects on oxidative stress and apoptosis. Therefore, apelin13 (which down-regulates APJ) and ML221 (an APJ antagonist) may have suppressed APJ signalling, which would account for our findings on the mitigation of polycystic ovarian syndrome. In conclusion, both apelin13 and ML221 mediated mitigation have different mechanisms, which need further investigation.

Investigating GABA Neuron-Specific Androgen Receptor Knockout in two Hyperandrogenic Models of PCOS.

Androgen excess is a hallmark feature of polycystic ovary syndrome (PCOS), the most common form of anovulatory infertility. Clinical and preclinical evidence links developmental or chronic exposure to hyperandrogenism with programming and evoking the reproductive and metabolic traits of PCOS. While critical androgen targets remain to be determined, central GABAergic neurons are postulated to be involved. Here, we tested the hypothesis that androgen signaling in GABAergic neurons is critical in PCOS pathogenesis in 2 well-characterized hyperandrogenic mouse models of PCOS. Using cre-lox transgenics, GABA-specific androgen receptor knockout (GABARKO) mice were generated and exposed to either acute prenatal androgen excess (PNA) or chronic peripubertal androgen excess (PPA). Females were phenotyped for reproductive and metabolic features associated with each model and brains of PNA mice were assessed for elevated GABAergic input to gonadotropin-releasing hormone (GnRH) neurons. Reproductive and metabolic dysfunction induced by PPA, including acyclicity, absence of corpora lutea, obesity, adipocyte hypertrophy, and impaired glucose homeostasis, was not different between GABARKO and wild-type (WT) mice. In PNA mice, acyclicity remained in GABARKO mice while ovarian morphology and luteinizing hormone secretion was not significantly impacted by PNA or genotype. However, PNA predictably increased the density of putative GABAergic synapses to GnRH neurons in adult WT mice, and this PNA-induced plasticity was absent in GABARKO mice. Together, these findings suggest that while direct androgen signaling in GABA neurons is largely not required for the development of PCOS-like traits in androgenized models of PCOS, developmental programming of GnRH neuron innervation is dependent upon androgen signaling in GABA neurons.

Hyperandrogenic environment regulates the function of ovarian granulosa cells by modulating macrophage polarization in PCOS.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a common endocrine-metabolic disorder characterized by oligo-anovulation, hyperandrogenism, and polycystic ovaries, with hyperandrogenism being the most prominent feature of PCOS patients. However, whether excessive androgens also exist in the ovarian microenvironment of patients with PCOS, and their modulatory role on ovarian immune homeostasis and ovarian function, is not clear. METHODS: Follicular fluid samples from patients participating in their first in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) treatment were collected. Androgen concentration of follicular fluid was assayed by chemiluminescence, and the macrophage M1:M2 ratio was detected by flow cytometry. In an in vitro model, we examined the regulatory effects of different concentrations of androgen on macrophage differentiation and glucose metabolism levels using qRT-PCR, Simple Western and multi-factor flow cytometry assay. In a co-culture model, we assessed the effect of a hyperandrogenic environment in the presence or absence of macrophages on the function of granulosa cells using qRT-PCR, Simple Western, EdU assay, cell cycle assay, and multi-factor flow cytometry assay. RESULTS: The results showed that a significantly higher androgen level and M1:M2 ratio in the follicular fluid of PCOS patients with hyperandrogenism. The hyperandrogenic environment promoted the expression of pro-inflammatory and glycolysis-related molecules and inhibited the expression of anti-inflammatory and oxidative phosphorylation-related molecules in macrophages. In the presence of macrophages, a hyperandrogenic environment significantly downregulated the function of granulosa cells. CONCLUSION: There is a hyperandrogenic microenvironment in the ovary of PCOS patients with hyperandrogenism. Hyperandrogenic microenvironment can promote the activation of ovarian macrophages to M1, which may be associated with the reprogramming of macrophage glucose metabolism. The increased secretion of pro-inflammatory cytokines by macrophages in the hyperandrogenic microenvironment would impair the normal function of granulosa cells and interfere with normal ovarian follicle growth and development.

Developmental programming: Testosterone excess masculinizes female pancreatic transcriptome and function in sheep.

Hyperandrogenic disorders, such as polycystic ovary syndrome, are often associated with metabolic disruptions such as insulin resistance and hyperinsulinemia. Studies in sheep, a precocial model of translational relevance, provide evidence that in utero exposure to excess testosterone during days 30-90 of gestation (the sexually dimorphic window where males naturally experience elevated androgens) programs insulin resistance and hyperinsulinemia in female offspring. Extending earlier findings that adverse effects of testosterone excess are evident in fetal day 90 pancreas, the end of testosterone treatment, the present study provides evidence that transcriptomic and phenotypic effects of in utero testosterone excess on female pancreas persist after cessation of treatment, suggesting lasting organizational changes, and induce a male-like phenotype in female pancreas. These findings demonstrate that the female pancreas is susceptible to programmed masculinization during the sexually dimorphic window of fetal development and shed light on underlying connections between hyperandrogenism and metabolic homeostasis.

Cellular senescence of granulosa cells in the pathogenesis of polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders in women of reproductive age, but its pathology has not been fully characterized and the optimal treatment strategy remains unclear. Cellular senescence is a permanent state of cell-cycle arrest that can be induced by multiple stresses. Senescent cells contribute to the pathogenesis of various diseases, owing to an alteration in secretory profile, termed 'senescence-associated secretory phenotype' (SASP), including with respect to pro-inflammatory cytokines. Senolytics, a class of drugs that selectively eliminate senescent cells, are now being used clinically, and a combination of dasatinib and quercetin (DQ) has been extensively used as a senolytic. We aimed to investigate whether cellular senescence is involved in the pathology of PCOS and whether DQ treatment has beneficial effects in patients with PCOS. We obtained ovaries from patients with or without PCOS, and established a mouse model of PCOS by injecting dehydroepiandrosterone. The expression of the senescence markers p16INK4a, p21, p53, γH2AX, and senescence-associated ß-galactosidase and the SASP-related factor interleukin-6 was significantly higher in the ovaries of patients with PCOS and PCOS mice than in controls. To evaluate the effects of hyperandrogenism and DQ on cellular senescence in vitro, we stimulated cultured human granulosa cells (GCs) with testosterone and treated them with DQ. The expression of markers of senescence and a SASP-related factor was increased by testosterone, and DQ reduced this increase. DQ reduced the expression of markers of senescence and a SASP-related factor in the ovaries of PCOS mice and improved their morphology. These results indicate that cellular senescence occurs in PCOS. Hyperandrogenism causes cellular senescence in GCs in PCOS, and senolytic treatment reduces the accumulation of senescent GCs and improves ovarian morphology under hyperandrogenism. Thus, DQ might represent a novel therapy for PCOS.

The Search for the Causes of Common Hyperandrogenism, 1965 to Circa 2015.

From 1965 to 2015, immense strides were made into understanding the mechanisms underlying the common androgen excess disorders, premature adrenarche and polycystic ovary syndrome (PCOS). The author reviews the critical discoveries of this era from his perspective investigating these disorders, commencing with his early discoveries of the unique pattern of plasma androgens in premature adrenarche and the elevation of an index of the plasma free testosterone concentration in most hirsute women. The molecular genetic basis, though not the developmental biologic basis, for adrenarche is now known and 11-oxytestosterones shown to be major bioactive adrenal androgens. The evolution of the lines of research into the pathogenesis of PCOS is historically traced: research milestones are cited in the areas of neuroendocrinology, insulin resistance, hyperinsulinism, type 2 diabetes mellitus, folliculogenesis, androgen secretion, obesity, phenotyping, prenatal androgenization, epigenetics, and complex genetics. Large-scale genome-wide association studies led to the 2014 discovery of an unsuspected steroidogenic regulator DENND1A (differentially expressed in normal and neoplastic development). The splice variant DENND1A.V2 is constitutively overexpressed in PCOS theca cells in long-term culture and accounts for their PCOS-like phenotype. The genetics are complex, however: DENND1A intronic variant copy number is related to phenotype severity, and recent data indicate that rare variants in a DENND1A regulatory network and other genes are related to PCOS. Obesity exacerbates PCOS manifestations via insulin resistance and proinflammatory cytokine excess; excess adipose tissue also forms testosterone. Polycystic ovaries in 40 percent of apparently normal women lie on the PCOS functional spectrum. Much remains to be learned.

Increased oxidative stress is associated with hyperandrogenemia in polycystic ovary syndrome evidenced by oxidized lipoproteins stimulating rat ovarian androgen synthesis in vitro.

PURPOSE: To determine the relationship between serum total testosterone (TT) levels and oxidative stress indices in patients with polycystic ovary syndrome (PCOS), and to investigate the effect of oxidative stress on androgen synthesis and its mechanism in rat ovarian theca-interstitial (T-I) cells. METHODS: Clinical, hormonal, metabolic, and oxidative stress parameters were analyzed in a cross-sectional case-control study including 626 patients with PCOS and 296 controls. The effects of oxidized low-density lipoprotein (ox-LDL) and oxidized high-density lipoprotein (ox-HDL) on cell proliferation, TT secretion, and expression of key enzymes involved in testosterone synthesis were evaluated in T-I cells. RESULTS: Serum TT levels were elevated with an increase in ox-LDL levels, whereas glutathione concentrations were lower in the high-TT subgroup than in the low-TT subgroup. The average ovarian volume and ox-LDL and malondialdehyde levels were significant predictors of TT levels in the multivariate regression models. In a rat ovarian T-I cell model, lipoprotein and oxidized lipoprotein treatments stimulated proliferation and promoted testosterone secretion. The mRNA and protein levels of 17α-hydroxylase were significantly higher in oxidized lipoprotein-treated cells than those in lipoprotein-treated cells. The mRNA levels of cholesterol side chain cleavage enzyme and steroidogenic acute regulatory protein were also significantly higher in ox-HDL-treated cells than in HDL-treated cells. CONCLUSIONS: Oxidative stress can promote androgen production by up-regulating the expression of testosterone synthesis-related enzymes in vitro and may be an essential factor in elevating serum TT levels in patients with PCOS.

Alterations in nonesterified free fatty acid trafficking rather than hyperandrogenism contribute to metabolic health in obese women with polycystic ovary syndrome.

OBJECTIVE: To determine whether alterations in nonesterified fatty acid (NEFA) dynamics or degree of hyperandrogenism (HA) contribute to the difference in insulin sensitivity between women with metabolically healthy obese polycystic ovary syndrome (PCOS) (MHO-PCOS) and women with metabolically unhealthy obese PCOS (MUO-PCOS). DESIGN: Prospective cross-sectional study. SETTING: Tertiary-care academic center. PATIENTS: One hundred twenty-five obese women with PCOS. INTERVENTION: Consecutive obese (body mass index [BMI] ≥ 30 kg/m2) oligo-ovulatory women (n = 125) with PCOS underwent an oral glucose tolerance test and a subgroup of 16 participants underwent a modified frequently sampled intravenous glucose tolerance test to determine insulin-glucose and -NEFA dynamics. MAIN OUTCOME MEASURES: Degree of insulin resistance (IR) in adipose tissue (AT) basally (Adipo-IR) and dynamically (the nadir in NEFA levels observed [NEFAnadir], the time it took for NEFA levels to reach nadir [TIMEnadir], and the percent suppression in plasma NEFA levels from baseline to nadir [%NEFAsupp]); peak lipolysis rate (SNEFA) and peak rate of NEFA disposal from plasma pool (KNEFA); whole-body insulin-glucose interaction (acute response of insulin to glucose [AIRg], insulin sensitivity index [Si], glucose effectiveness [Sg], and disposition index [Di]); and HA (hirsutism score, total and free testosterone levels, and dehydroepiandrosterone sulfate levels). RESULTS: A total of 85 (68%) women were MUO-PCOS and 40 (32%) were MHO-PCOS using the homeostasis model of assessment of IR. Subjects with MUO-PCOS and MHO-PCOS did not differ in mean age, BMI, waist-to-hip ratio, HA, and lipoprotein levels. By a modified frequently sampled intravenous glucose tolerance test, eight women with MUO-PCOS had lesser Si, KNEFA, and the percent suppression in plasma NEFA levels from baseline to nadir (%NEFAsupp) and greater TIMEnadir, NEFAnadir, and baseline adipose tissue IR index (Adipo-IR) than eight subjects with MHO-PCOS, but similar fasting NEFA levels and SNEFA. Women with MUO-PCOS had a higher homeostasis model of assessment-ß% and fasting insulin levels than women with MHO-PCOS. In bivalent analysis, Si correlated strongly and negatively with Adipo-IR and NEFAnadir, weakly and negatively with TIMEnadir, and positively with KNEFA and %NEFAsupp, in women with MUO-PCOS only. CONCLUSION: Independent of age and BMI, women with MUO-PCOS have reduced NEFA uptake and altered insulin-mediated NEFA suppression, but no difference in HA, compared with women with MHO-PCOS. Altered insulin-mediated NEFA suppression, rather than HA or lipolysis rate, contributes to variations in insulin sensitivity among obese women with PCOS.

Discriminatory Value of Steroid Hormones on Polycystic Ovary Syndrome and Clustering of Hyperandrogenism and Metabolic Factors.

OBJECTIVE: We determined (1) if 11-oxygenated androgens better identify polycystic ovary syndrome (PCOS) diagnosis in women with obesity compared to total or free testosterone (T) and free androgen index; (2) how biochemical hyperandrogenism and metabolic factors cluster in a cohort of women with infertility and obesity. METHODS: Women with obesity and PCOS comprised the study group (N = 132). Ovulatory women with obesity and idiopathic, tubal or male factor infertility were the control group (N = 83). Steroid hormones were measured by means of liquid chromatography tandem mass spectrometry. Receiver operating characteristic curves and principal component analysis were used. RESULTS: Women with obesity and PCOS had higher 11-ketotestosterone (11 KT) (1.22 nmol/L [0.84; 1.65] vs 1.05 [0.78; 1.35], P = .04) compared to controls, but not 11ß-hydroxyandrostenedione 4.30 [2.87; 5.92] vs 4.06 [3.22; 5.73], P = .44). 11-ketotestosterone (area under the curve: 0.59) did not better discriminate PCOS in women with obesity compared to: total T (0.84), free T (0.91), and free androgen index (0.85). We identified 4 principal components (PCs) in the PCOS group (72.1% explained variance): (1) insulin resistance status; (2) blood pressure; (3) obesity; (4) androgen status and 4 PCs in the control group (68.7% explained variance) with variables representing metabolism being dispersed in component 2, 3, and 4. CONCLUSIONS: Eleven-oxygenated androgens do not aid in the diagnosis of PCOS in women with obesity. Insulin resistance is the strongest PC in the PCOS group. There is no major dominant characteristic that defines obese non-PCOS women.

Metabolic dysregulation and gut dysbiosis linked to hyperandrogenism in female mice.

INTRODUCTION: Polycystic ovary syndrome (PCOS) is a common endocrine pathology in women. In addition to infertility, women with PCOS have metabolic dysregulation which predisposes them to Type 2 diabetes, cardiovascular disease and non-alcoholic fatty liver disease. Moreover, women with PCOS have changes in their gut microbial community that may be indicative of dysbiosis. While hyperandrogenism is associated with both the development of metabolic dysfunction and gut dysbiosis in females, the mechanisms involved are not well understood. METHODS: We used dihydrotestosterone (DHT) and ovariectomy (OVX) mouse models coupled with metabolic assessments and 16S rRNA gene sequencing to explore the contributions of hyperandrogenism and oestrogen deficiency to the development of insulin resistance and gut microbial dysbiosis in pubertal female mice. RESULTS: We demonstrated that, while DHT treatment or OVX alone were insufficient to induce insulin resistance during the pubertal-to-adult transition, combining OVX with DHT resulted in insulin resistance similar to that observed in letrozole-treated mice with elevated testosterone and decreased oestrogen levels. In addition, our results showed that OVX and DHT in combination resulted in a distinct shift in the gut microbiome compared to DHT or OVX alone, suggesting that the substantial metabolic dysregulation occurring in the OVX + DHT model was accompanied by unique changes in the abundances of gut bacteria including S24-7, Rikenellaceae and Mucispirillum schaedleri. CONCLUSIONS: While hyperandrogenism plays an important role in the development of metabolic dysregulation in female mice, our results indicate that investigation into additional factors influencing insulin resistance and the gut microbiome during the pubertal-to-adult transition could provide additional insight into the pathophysiology of PCOS.

Defective Uterine Spiral Artery Remodeling and Placental Senescence in a Pregnant Rat Model of Polycystic Ovary Syndrome.

Pregnancy-related problems have been linked to impairments in maternal uterine spiral artery (SpA) remodeling. The mechanisms underlying this association are still unclear. It is also unclear whether hyperandrogenism and insulin resistance, the two common manifestations of polycystic ovary syndrome, affect uterine SpA remodeling. We verified previous work in which exposure to 5-dihydrotestosterone (DHT) and insulin (INS) in rats during pregnancy resulted in hyperandrogenism, insulin intolerance, and higher fetal mortality. Exposure to DHT and INS dysregulated the expression of angiogenesis-related genes in the uterus and placenta and also decreased expression of endothelial nitric oxide synthase and matrix metallopeptidases 2 and 9, increased fibrotic collagen deposits in the uterus, and reduced expression of marker genes for SpA-associated trophoblast giant cells. These changes were related to a greater proportion of unremodeled uterine SpAs and a smaller proportion of highly remodeled arteries in DHT + INS-exposed rats. Placentas from DHT + INS-exposed rats exhibited decreased basal and labyrinth zone regions, reduced maternal blood spaces, diminished labyrinth vascularity, and an imbalance in the abundance of vascular and smooth muscle proteins. Furthermore, placentas from DHT + INS-exposed rats showed expression of placental insufficiency markers and a significant increase in cell senescence-associated protein levels. Altogether, this work demonstrates that increased pregnancy complications in polycystic ovary syndrome may be mediated by problems with uterine SpA remodeling, placental functionality, and placental senescence.

GC-MS analysis and in silico docking of constituents of Cinnamomum malabatrum against CYP450 17α and CYP450 19 (Aromatase)- Key targets for hyperandrogenism.

Poly cystic ovary syndrome (PCOS) is considered as one of the common hormonal disorders affecting 6-20% of women in their reproductive age with characteristic features include anovulatory infertility, hyperandrogenism, cystic follicles and insulin resistance. The gene CYP play an important role in pathophysiology of hyperandrogenism associated with PCOS. An elevated androgens are reported in PCOS condition due to overexpression of the enzyme CYP450 17 α: . As well as diminished levels of aromatase (CYP450 19) were observed in several hyperandrogenic PCOS patients. The powdered leafy material of Cinnamomum malabatrum was subjected to Soxhlet extraction. The plant extract was subjected to Gas chromatography-MS analysis (GC-MS), and the chromatogram obtained revealed the presence of active chemical constituents like 1(10),9(11)-B-Homolanistadiene for the first time and other potential compounds. Hypothesis has raised to interpret the efficiency of phytoconstituents of Cinnamomum malabatrum on these enzyme targets and which may be a novel drug candidate for the treatment and maintenance of hyperandrogenism associated with PCOS. Thus, the results obtained from the in-silico study of Cinnamomum malabatrum leaf extract using computational approaches indicate that the phytoconstituents have good affinities for the selected two key targets. ADME and PASS studies has been performed for active phytoconstituents homolanistadiene, ß-sitosterol, cycloartenol and a pyrazole derivative, and results revealed the Lipinski drug-likeness and pharmacological potential. In conclusion, this work throws a new insight into the possibility of the active phytoconstituents on binding the two active CYP45017 α and CYP45019 aromatase enzymes which facilitates development of novel compounds for hyperandrogenism associated with PCOS.

Single-Cell RNA-Seq Identifies Pathways and Genes Contributing to the Hyperandrogenemia Associated with Polycystic Ovary Syndrome.

Polycystic ovary syndrome (PCOS) is a common endocrine disorder characterized by hyperandrogenemia of ovarian thecal cell origin, resulting in anovulation/oligo-ovulation and infertility. Our previous studies established that ovarian theca cells isolated and propagated from ovaries of normal ovulatory women and women with PCOS have distinctive molecular and cellular signatures that underlie the increased androgen biosynthesis in PCOS. To evaluate differences between gene expression in single-cells from passaged cultures of theca cells from ovaries of normal ovulatory women and women with PCOS, we performed single-cell RNA sequencing (scRNA-seq). Results from these studies revealed differentially expressed pathways and genes involved in the acquisition of cholesterol, the precursor of steroid hormones, and steroidogenesis. Bulk RNA-seq and microarray studies confirmed the theca cell differential gene expression profiles. The expression profiles appear to be directed largely by increased levels or activity of the transcription factors SREBF1, which regulates genes involved in cholesterol acquisition (LDLR, LIPA, NPC1, CYP11A1, FDX1, and FDXR), and GATA6, which regulates expression of genes encoding steroidogenic enzymes (CYP17A1) in concert with other differentially expressed transcription factors (SP1, NR5A2). This study provides insights into the molecular mechanisms underlying the hyperandrogenemia associated with PCOS and highlights potential targets for molecular diagnosis and therapeutic intervention.

Effects of early life overnutrition and hyperandrogenism on spatial learning and memory in a rat model of polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a complex disorder characterized by endocrine and metabolic abnormalities such as obesity and insulin resistance. PCOS is also associated with psychiatric disorders and cognitive impairment. The animal model of PCOS was induced by treating rats with 5α-dihydrotestosterone (5α-DHT) and additionally modified to induce adiposity by litter size reduction (LSR). Spatial learning and memory were assessed using the Barnes Maze test, and striatal markers of synaptic plasticity were analyzed. Striatal insulin signaling was estimated by the levels of insulin receptor substrate 1 (IRS1), its inhibitory phosphorylation at Ser307, and glycogen synthase kinase-3α/ß (GSK3α/ß) activity. Both LSR and DHT treatment significantly decreased striatal protein levels of IRS1, followed by increased GSK3α/ß activity in small litters. Results of the behavioral study showed that LSR had a negative effect on learning rate and memory retention, whereas DHT treatment did not induce impairment in memory formation. While protein levels of synaptophysin, GAP43, and postsynaptic density protein 95 (PSD-95) were not altered by the treatments, DHT treatment induced an increase in phosphorylation of PSD-95 at Ser295 in both normal and small litters. This study revealed that LSR and DHT treatment suppressed insulin signaling by downregulating IRS1 in the striatum. However, DHT treatment did not have an adverse effect on learning and memory, probably due to compensatory elevation in pPSD-95-Ser295, which had a positive effect on synaptic strength. This implies that hyperandrogenemia in this setting does not represent a threat to spatial learning and memory, opposite to the effect of overnutrition-related adiposity.

Deletion of Androgen Receptors From Kisspeptin Neurons Prevents PCOS Features in a Letrozole Mouse Model.

Polycystic ovarian syndrome (PCOS) is the leading cause of anovulatory infertility and is a heterogenous condition associated with a range of reproductive and metabolic impairments. While its etiology remains unclear, hyperandrogenism and impaired steroid negative feedback have been identified as key factors underpinning the development of PCOS-like features both clinically and in animal models. We tested the hypothesis that androgen signaling in kisspeptin-expressing neurons, which are key drivers of the neuroendocrine reproductive axis, is critically involved in PCOS pathogenesis. To this end, we used a previously validated letrozole (LET)-induced hyperandrogenic mouse model of PCOS in conjunction with Cre-lox technology to generate female mice exhibiting kisspeptin-specific deletion of androgen receptor (KARKO mice) to test whether LET-treated KARKO females are protected from the development of reproductive and metabolic PCOS-like features. LET-treated mice exhibited hyperandrogenism, and KARKO mice exhibited a significant reduction in the coexpression of kisspeptin and androgen receptor mRNA compared to controls. In support of our hypothesis, LET-treated KARKO mice exhibited improved estrous cyclicity, ovarian morphology, and insulin sensitivity in comparison to LET-treated control females. However, KARKO mice were not fully protected from the effects of LET-induced hyperandrogenism and still exhibited reduced corpora lutea numbers and increased body weight gain. These data indicate that increased androgen signaling in kisspeptin-expressing neurons plays a critical role in PCOS pathogenesis but highlight that other mechanisms are also involved.

Characterization of trophoblast mitochondrial function and responses to testosterone treatment in ACH-3P cells.

INTRODUCTION: Trophoblast mitochondria play important roles in placental energy metabolism, physiology and pathophysiology. Hyperandrogenism has been associated with mitochondrial abnormalities in pregnancy disorders such as pre-eclampsia, gestational diabetes, and intrauterine growth restriction, but the direct impacts of androgen exposure on placental mitochondrial function are unknown. Given the inherent limitations of studying the human placenta during pregnancy, trophoblast cell lines are routinely used to model placental biology in vitro. The aim of this study was to characterize mitochondrial respiratory function in four commonly used trophoblast cell lines to provide a basis for selecting one well-suited to investigating the impact of androgens on trophoblast mitochondrial function. METHODS: Androgen receptor expression, mitochondrial respiration (JO2) and reactive oxygen species (ROS) release rates were evaluated in three human trophoblast cell lines (ACH-3P, BeWo and Swan-71) and one immortalized ovine trophoblast line (iOTR) under basal and substrate-stimulated conditions using high-resolution fluorespirometry. RESULTS: ACH-3P cells exhibited the greatest mitochondrial respiratory capacity and coupling efficiency of the four trophoblast lines tested, along with robust expression of androgen receptor protein that was found to co-localize with mitochondria by immunoblot and immunofluorescence. Acute testosterone administration (10 nM) tended to decrease ACH-3P mitochondrial JO2 and increase ROS release, while chronic (7 days) testosterone exposure increased expression of mitochondrial proteins, JO2, and ROS release. DISCUSSION: These studies establish ACH-3P as a suitable cell line for investigating trophoblast mitochondrial function, and provide foundational evidence supporting links between hyperandrogenism and placental mitochondrial ROS production with potential relevance to several common pregnancy disorders.