Follicular Growth Arrest Research Articles

Impact of Prestimulation Treatment

Polycystic ovarian syndrome (PCOS) is one of the commonest endocrine disorders encountered in women of reproductive age. It is characterised by an altered Androgen metabolism which leads to irregular menses, anovulation and hyperandrogenism (HA). It is associated with hyperinsulinaemia and obesity which aggravate the HA status. Ovulation induction and controlled ovarian stimulation pose a unique challenge in these patients. The endocrine disturbance leads to arrest of follicular growth and patients may either not respond to OI drugs are may hyper-respond leading to a risk of OHSS. The best strategy to improve outcome of IVF in PCOS patients is still under consideration. This lecture will address pre-treatment (IVF) drugs and strategies that ameliorate HA, decrease the risk of OHSS and improve cycle outcome. Selection of the safest and most effective OS protocols will be discussed. Use of segmental IVF in PCOS patients has almost eliminated OHSS- it involves using an antagonist protocol with a GnRH agonist trigger and a freeze all strategy. The pros and cons of this strategy will be discussed. We will also look at the current status of Metformin Inositols, Vit D in PCOS patients and their role in improving IVF outcome. Use of pre-treatment hormones/OCP’s and their effect on IVF outcome will be deliberated. Lifestyle change and weight reduction are important to improve the HI & HA status and have become a fundamental requirement in these young women. The challenge patients face in achieving weight reduction goals due to the inherent metabolic disturbances needs to be recognised. At the end of the lecture the delegate would be able to have a better understanding of the current status of various pre-treatment strategies used in PCOS patients undergoing IVF.

Androgen-induced exosomal miR-379-5p release determines granulosa cell fate: cellular mechanism involved in polycystic ovaries

Polycystic ovarian syndrome (PCOS) is a complex multi-factorial syndrome associated with androgen excess and anovulatory infertility. In the current study, we investigated the role of dihydrotestosterone-induced exosomal miR-379-5p release in determining the destiny of the developing follicles. Our hypothesis was that androgen regulates granulosa cell miR-379-5p content by facilitating its exosomal release in a follicular-stage dependent manner, a process which determines granulosa cell fate. Compared to human non-PCOS subjects, individuals with PCOS exhibit higher follicular fluid free testosterone levels, lower exosomal miR-379-5p content and granulosa cell proliferation. Androgenized rats exhibited lower granulosa cell miR-379-5p but higher phosphoinositide-dependent kinase-1 (PDK1; a miR-379-5p target) content and proliferation. Androgen reduced granulosa cell miR-379-5p content by increasing its exosomal release in preantral follicles, but not in antral follicles in vitro. Studies with an exosomal release inhibitor confirmed that androgen-induced exosomal miR-379-5p release decreased granulosa cell miR-379-5p content and proliferation. Ovarian overexpression of miR-379-5p suppressed granulosa cell proliferation, and basal and androgen-induced preantral follicle growth in vivo. These findings suggest that increased exosomal miR-379-5p release in granulosa cells is a proliferative response to androgenic stimulation specific for the preantral stage of follicle development and that dysregulation of this response at the antral stage is associated with follicular growth arrest, as observed in human PCOS.

Sex-Dependent Bioaccumulation of Nano Zinc Oxide and Its Adverse Effects on Sexual Behavior and Reproduction in Japanese Medaka.

This study investigated the adverse effects of 200 nm zinc oxide particles (nZnO) on sexual behavior and reproduction in Japanese medaka in comparison with ZnSO4 and correlated the consequences with the bioaccumulation pattern of the particles in associated organs. nZnO exposure impaired sexual and territorial behaviors and affected fertility by altering sperm viability and motility in males through reactive oxygen species (ROS) induction. Conversely, none of these effects other than behavior loss was seen in males exposed to ZnSO4. nZnO exposure to females induced ROS in ovaries, causing follicular growth arrest, atresia, and subfertility. Further, sex-steroid levels were altered by both nZnO and ZnSO4 in males and by nZnO but not ZnSO4 in females. Biodistribution studies revealed the deposition of nZnO as particulate matter in the brain, gills, gut, kidney, and ovary. Particle accumulation in the brain was sex specific, as the particles were found in the brain of males but not that of females. A similar trend was seen for zinc levels in males and females exposed to ZnSO4. Importantly, the female sex hormone, 17β-estradiol was found to prevent nZnO accumulation in the female brain, emphasizing the need for biodistribution profiling of nanoparticle-based drug delivery vehicles separately in males and females before they are commercialized. This study has demonstrated that the toxic effects of nZnO on the reproductive system were mainly caused by ROS induction, while zinc ions were predominantly responsible for the adverse impact of ZnSO4.

Identification of the similarly expressed genes in patients with polycystic ovary syndrome and transsexuals.

Polycystic ovary syndrome (PCOS) is a common female infertility, which may be caused by excessive androgen, but its mechanism remains unknown. Transsexuals are women who take androgen drugs for a long time, and gradually have male signs. Their ovaries may have received high concentrations of androgen, which leads to the failure of ovarian reproductive function. Therefore, we searched the relevant data of PCOS and transsexuals in gene expression omnibus database, used limma package to identify the most similarly genes, and then analyzed the possible mechanism of PCOS through gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) pathway analysis. Then, the protein-protein interaction network was constructed by searching the String database, and the top 5 hub genes were identified by the cytohubba plug-in of Cytoscape. Finally, ubiquitin conjugating enzyme E2 E1 (UBE2E1), ubiquitin C (UBC), transcription elongation factor B subunit 1 (TCEB1), ubiquitin conjugating enzyme E2 N (UBE2N), and ring finger protein 7 (RNF7) genes were identified as the most similarly expressed genes between PCOS and Transsexuals. They may cause the ubiquitination of androgen receptor and eventually lead to sinus follicular growth arrest. In conclusion, 5 Central genes were identified in PCOS and transsexuals. These genes can be used as targets for early diagnosis or treatment of PCOS.

Improving the sexual activity and reproduction of female zebrafish with high testosterone levels

High levels of testosterone cause clinical symptoms in female reproduction and possibly, alterations in sexuality. Yet, the underlying mechanisms remain to be examined. Here, we report a study that investigates the effects of testosterone in follicle development and sexual mating using zebrafish models. We developed an acute zebrafish model with high testosterone levels by exposing young female zebrafish to testosterone dissolved in swimming water. After given a high concentration of testosterone treatment (e.g., 100 ng/ml), the fish showed hallmark pathological symptoms similar to those displayed in patients with polycystic ovary syndrome (PCOS), such as follicular growth-arrest, rare ovulation, ovary enlargement, decrease in reproduction, and down regulation of the expression of some PCOS susceptible genes, such as Tox3. These fish are referred to as the PCOS fish. By monitoring mating-like swimming behaviors, we measured the sexual activity of PCOS zebrafish. In general, the PCOS fish showed no desire to interact with males. As a consequence, their mating rate was decreased as compared to control animals. The sexuality levels of PCOS fish, however, could be improved after short periods of rearing in conditions that lack of males. After only 3 days of rearing alone, the PCOS fish showed an increase in sexuality levels and displayed characteristic swimming patterns for mating. After 30 days of separation from males, not only the sexual activity, but also the mating rate was improved in the PCOS fish. Together, the data suggests that zebrafish can serve as a new type of research model to further develop strategies for the treatment of reproductive disorders, such as those related to PCOS.

Direct impact of gonadotropins on glucose uptake and storage in preovulatory granulosa cells: Implications in the pathogenesis of polycystic ovary syndrome

BackgroundPolycystic ovary syndrome (PCOS) is often associated with higher levels of LH, and arrested ovarian follicular growth. The direct impact of high LH on FSH mediated metabolic responses in PCOS patients is not clearly understood. MethodIn order to investigate the impact of FSH and LH on glucose metabolism in preovulatory granulosa cells (GCs), we used [U14C]-2 deoxyglucose, D-[U14C]-glucose or 2-NBD glucose to analyse glucose uptake and its incorporation into glycogen. To reproduce the high androgenic potential in PCOS patients, we administered hCG both in vitro and in vivo. The role of IRS-2/PI3K/Akt2 pathway was studied after knockdown with specific siRNA. Immunoprecipitation and specific assays were used for the assessment of IRS-2, glycogen synthase and protein phosphatase 1. Furthermore, we examined the in vivo effects of hCG on FSH mediated glycogen increase in normal and PCOS rat model. HEK293 cells co-expressing FSHR and LHR were used to demonstrate glucose uptake and BRET change by FSH and hCG. ResultsIn normal human and rat granulosa cells, FSH is more potent than hCG in stimulating glucose uptake, however glycogen synthesis was significantly upregulated only by FSH through increase in activity of glycogen synthase via IRS-2/PI3K/Akt2 pathway. On the contrary, an impaired FSH-stimulated glucose uptake and glycogen synthesis in granulosa cells of PCOS-patients indicated a selective defect in FSHR activation. Further, in normal human granulosa cells, and in immature rat model, the impact of hCG on FSH responses was such that it inhibited the FSH-mediated glucose uptake as well as glycogen synthesis through inhibition of FSH-stimulated IRS-2 expression. These findings were further validated in HEK293 cells overexpressing Flag-LHR and HA-FSHR, where high hCG inhibited the FSH-stimulated glucose uptake. Notably, an increased BRET change was observed in HEK293 cells expressing FSHR-Rluc8 and LHR-Venus possibly suggesting increased heteromerization of LHR and FSHR in the presence of both hCG and FSH in comparison to FSH or hCG alone. ConclusionOur findings confirm a selective attenuation of metabolic responses to FSH such as glucose uptake and glycogen synthesis by high activation level of LHR leading to the inhibition of IRS-2 pathway, resulting in depleted glycogen stores and follicular growth arrest in PCOS patients.

Significance of pro-angiogenic estrogen metabolites in normal follicular development and follicular growth arrest in polycystic ovary syndrome.

Do alterations in pro- and anti-angiogenic estrogen metabolites in follicular fluid (FF) contribute to the follicular growth arrest and anovulation associated with polycystic ovary syndrome (PCOS)? FF of PCOS women with anovulation have reduced levels of pro-angiogenic estrogen metabolites (EMs) and vascular endothelial growth factor (VEGF) compared to that of fertile women with regular menstrual cycles, but exogenous gonadotropins increase the pro-angiogenic EMs and VEGF levels in PCOS women. PCOS is characterized by the arrest of follicular development that leads to chronic anovulation. Follicular arrest is generally associated with elevated plasma levels of luteinizing hormone (LH), androgens and anti-Mullerian hormone (AMH). There is also reduced angiogenesis in the follicles of PCOS women compared to those of normal cycling women. It is known that angiogenesis is a critical factor during follicular development. We and other investigators have explored the role of EMs in ovarian angiogenesis, particularly in human corpus luteum function, showing that 4-hydroxyestrone (4-OHE1) and 16-ketoestradiol (16-kE2) have pro-angiogenic effects while 2-methoxyestradiol (2-ME2) and 2-methoxyestrone (2-ME1) have anti-angiogenic effects. Additionally, 2-hydroxyestradiol (2-OHE2), which is produced in the ovary, has proliferative and pro-angiogenic properties. We hypothesized that EMs could be involved in angiogenesis necessary for ovarian follicular development in fertile women, and that dysregulation of these factors may contribute to follicular arrest in PCOS. The relationship between EMs, VEGF and AMH in the pathophysiology of follicular arrest in PCOS has not been previously studied at a follicular level in anovulatory women without ovulation induction. This is a comparative experimental study of serum and FF collected from different sized follicles (antral ˂10 mm and dominant ˃16 mm) of women with and without ovarian stimulation. The study included women with regular menstrual cycles who were proven to be fertile (n = 20) and PCOS women with follicular arrest who were candidates for ovarian drilling (n = 17), as well as other patients requiring ovarian stimulation, i.e. control women undergoing IVF for male factor infertility (n = 12) and PCOS women undergoing IVF (n = 17). In vitro studies were carried out on granulosa-lutein cells (GCs) obtained from subsets of women undergoing IVF for male factor infertility (n = 6) and PCOS women undergoing IVF (n = 6). GCs were maintained in culture for up to 6 days. Intrafollicular estradiol, estrone and EMs concentrations were determined by high performance liquid chromatography-mass spectrometry. Testosterone in serum was measured by RIA, and LH, FSH and sex hormone-binding globulin in serum were measured with IRMA kits. AMH was determined in serum and FF by enzyme linked immunosorbant assay (ELISA). VEGF levels were measured in FF and conditioned medium by ELISA. Conditioned medium were obtained from cultured GCs. The angiogenic potential was assessed by in vitro angiogenic assays. Pro-angiogenic EMs (4-OHE1, 16-kE2 and 2-OHE2) and VEGF were lower in FF of antral follicles of PCOS women with follicular arrest compared those of fertile women with ovulatory cycles (P < 0.05). In contrast, higher concentrations of AMH were found in FF of antral follicles from PCOS women with follicular arrest compared to those of fertile women with ovulatory cycles (P < 0.05). Exogenous gonadotropins used in IVF increased pro-angiogenic EMs and VEGF production in PCOS women, reaching similar profiles compared to control women receiving gonadotropins in their IVF treatment for male factor infertility. The pro-angiogenic EM 2-OHE2 increased the angiogenic potential and VEGF levels of GCs from PCOS women compared to the basal condition (P < 0.05). These findings suggest that there is a role for pro-angiogenic EMs in the control of follicular VEGF production. The limitations include the possibility that in vitro analysis of GCs might not reflect the in vivo mechanisms involved in the pro-angiogenic action of 2-OHE2 since GCs obtained at the time of oocyte retrieval belong to a very early stage of the luteal phase and might not be representative of GCs during follicular growth. Therefore, our findings do not conclusively rule out the possibility that other in vivo mechanisms also account for defective angiogenesis observed in PCOS. The present study highlights the significance of EMs, angiogenic factors and AMH and their interaction in the pathophysiology of follicular development in PCOS. This study provides new insights into the role of pro-angiogenic factors in follicular arrest in PCOS. This study was funded by CONICYT/FONDECYT 1140693 and NIH grant R01HD083323. All authors declare no conflict of interest. N/A.

R-spondin2 signaling is required for oocyte-driven intercellular communication and follicular growth

R-spondin2 (RSPO2) is a member of the R-spondin family, which are secreted activators of the WNT/β-catenin (CTNNB1) signaling pathway. In the mouse postnatal ovary, WNT/CTNNB1 signaling is active in the oocyte and in the neighboring supporting cells, the granulosa cells. Although the role of Rspo2 has been previously studied using in vitro experiments, the results are conflicting and the in vivo ovarian function of Rspo2 remains unclear. In the present study, we found that RSPO2/Rspo2 expression is restricted to the oocyte of developing follicles in both human and mouse ovaries from the beginning of the follicular growth. In mice, genetic deletion of Rspo2 does not impair oocyte growth, but instead prevents cell cycle progression of neighboring granulosa cells, thus resulting in an arrest of follicular growth. We further show this cell cycle arrest to be independent of growth promoting GDF9 signaling, but rather associated with a downregulation of WNT/CTNNB1 signaling in granulosa cells. To confirm the contribution of WNT/CTNNB1 signaling in granulosa cell proliferation, we induced cell type specific deletion of Ctnnb1 postnatally. Strikingly, follicles lacking Ctnnb1 failed to develop beyond the primary stage. These results show that RSPO2 acts in a paracrine manner to sustain granulosa cell proliferation in early developing follicles. Taken together, our data demonstrate that the activation of WNT/CTNNB1 signaling by RSPO2 is essential for oocyte-granulosa cell interactions that drive maturation of the ovarian follicles and eventually female fertility.

Ovarian Expression of Adipokines in Polycystic Ovary Syndrome: A Role for Chemerin, Omentin, and Apelin in Follicular Growth Arrest and Ovulatory Dysfunction?

Adipokines are a potential link between reproduction and energy metabolism and could partly explain some infertilities related to some pathophysiology, such as polycystic ovary syndrome (PCOS). However, adipokines were predominantly assessed in blood samples, while very little is known concerning their variations in follicular fluid (FF) and ovarian granulosa cells (GCs) of PCOS women. Thus, the objectives of our study were to investigate adiponectin, chemerin, resistin, visfatin, omentin, and apelin ovarian expression in PCOS women in comparison with controls and women with only a polycystic ovary morphology. In total, 78 women undergoing an in vitro fertilization procedure were divided into three groups: 23 PCOS women, 28 women presenting only ≥12 follicles per ovary (ECHO group), and 27 control women. Each group almost equally included normal weight and obese women. Follicular fluid (FF) concentration and granulosa cells (GCs) mRNA expression of adipokines and their receptors were assessed by ELISA and RT-qPCR, respectively. Omentin levels in FF and GC were higher in PCOS than in ECHO and control women, while apelin expression was increased in both PCOS and ECHO groups. FF chemerin concentration was predominant in normal-weight PCOS women compared to BMI (Body Mass Index)-matched ECHO and control women, while GC mRNA levels were higher in the obese PCOS group than in the ECHO one. Compared to PCOS, ECHO women had increased FF adiponectin concentrations and lower plasma AMH levels. The FF concentration of all adipokines was higher in obese subjects except for adiponectin, predominant in normal-weight women. In conclusion, women with PCOS expressed higher GC chemerin and omentin, whereas the ECHO group presented higher levels of FF adiponectin and apelin and lower plasma AMH and LH concentrations. Chemerin, omentin, and apelin expression was differently regulated in women with PCOS, suggesting their possible role in follicular growth arrest and ovulatory dysfunction characterizing PCOS pathogenesis.

MON-202 Changes in the Expression of Epigenetic Enzymes Induced by Prenatal Testosterone Excess May Underlie the Antral Follicular Defects in the Sheep Model of PCOS

Prenatal testosterone (T) excess leads to oligo/anovulation and multifollicular ovary in sheep, characteristics of women with PCOS. Persistence of antral follicles - the result of reduced atresia, follicular growth arrest and premature luteinization, contribute at least in part to the multifollicular ovarian morphology. Decrease in aromatase (CYP19) in granulosa cells and 17alpha-hydroxylase (CYP17) in theca cells and increase in anti-Mullerian hormone (AMH) in granulosa cells may contribute to the antral follicular persistence. Observations that histone H3K9 hypermethylation decreases cumulus cell CYP19 (Journal of Assisted Reproduction and Genetics 33: 1105-1113, 2016), AMH gene hypermethylation reduces granulosa cell AMH (Oncotarget 6: 3627-3643, 2015), and inhibition of histone deacetylation reduces theca cell CYP17 expression (PloS One 7: e49553, 2012) suggest that epigenetic changes are involved. We hypothesized that prenatal T excess induced changes in expression of CYP17, CYP19 and AMH are facilitated by changes in expression of key epigenetic enzymes. Granulosa / theca cells were isolated via laser capture microdissection from antral follicles of 21 months-of-age control (n=5) and prenatal T-treated (100mg im. twice weekly from gestational day 30 to 90; term: 147 days; n=6) sheep. Expression of methylation / demethylation and / or acetylation / deacetylation enzymes were determined by real time PCR and data analyzed by Student’s t-test and Cohen’s effect size analysis. Prenatal T excess decreased (p < 0.05) histone demethylase KDM1A in granulosa and theca cells, histone acetylase (HDAC) 3 in theca cells, and increased histone methyltransferase SMYD3 in the theca cells. Prenatal T excess also induced large magnitude (1) increases in DNA methyltransferase (DNMT) 1, histone methyltransferases EZH2 and SUV39H1, and histone deacetylase HDAC1 in granulosa cells; (2) decrease in DNA methyltransferase DNMT3B, KDM1A, and HDAC3 in granulosa cells; (3) increase in SMYD3 and HDAC1 in theca cells, and (4) decrease in KDM1A and HDAC3 in theca cells. While decrease in KDM1A and increase in SUV39H1 leading to H3K9 hypermethylation may account for reduced granulosa cell expression of CYP19, downregulation of HDAC3 may underlie the decrease in thecal cell expression of CYP17. Decreased DNMT3B expression may contribute to hypomethylation of AMH gene and consequent increase in AMH expression in granulosa cells. These findings suggest that changes in expression of key genes involved in the development of multifollicular phenotype are likely mediated via granulosa and theca cell specific changes in epigenetic enzyme expression. Because the ovarian attributes of prenatal T-treated sheep parallel that seen in PCOS women, these observations may be of translational relevance. Supported by NIH PO1HD44232.

The Influence of Pentraxin 3 on the Ovarian Function and Its Impact on Fertility.

Follicular development is a highly coordinated process that in humans takes more than 6 months. Pituitary gonadotropins and a variety of locally produced growth factors and cytokines are involved in determining a precise sequence of changes in cell metabolism, proliferation, vascularization, and matrix remodeling in order to obtain a follicle with full ovulatory and steroidogenic capability. A low-grade inflammation can alter such processes leading to premature arrest of follicular growth and female reproductive failure. On the other hand, factors that are involved in inflammatory response as well as in innate immunity are physiologically upregulated in the follicle at the final stage of maturation and play an essential role in ovulation and fertilization. The generation of pentraxin 3 (PTX3) deficient mice provided the first evidence that this humoral pattern recognition molecule of the innate immunity has a non-redundant role in female fertility. The expression, localization, and molecular interactions of PTX3 in the periovulatory follicle have been extensively studied in the last 10 years. In this review, we summarize findings demonstrating that PTX3 is synthesized before ovulation by cells surrounding the oocyte and actively participates in the organization of the hyaluronan-rich provisional matrix required for successful fertilization. Data in humans tend to confirm these findings, indicating PTX3 as a biomarker of oocyte quality. Moreover, we discuss the emerging evidence that in humans altered PTX3 systemic levels, determined by genetic variations and/or low-grade chronic inflammation, can also impact the growth and development of the follicle and affect the incidence of ovarian disorders.

Regulation of androgen receptor signaling by ubiquitination during folliculogenesis and its possible dysregulation in polycystic ovarian syndrome

Although chronic hyperandrogenism suppresses antral follicular development, a phenomenon often observed in polycystic ovarian syndrome (PCOS), whether and how deregulation of androgen receptor (AR) signaling is involved, is not well understood. In the present study, we examined the role of ring finger protein 6 (RNF6) in AR ubiquitination and the possible dysregulation in the expression and actions of growth differentiation factor 9 (GDF9) and kit-ligand (Kitlg) in a chronic androgenized PCOS rat model. 5α-dihydrotestosterone (DHT) treatment in vivo inhibited antral follicle growth, a response mediated through increased RNF6 content, suppressed K63- but increased K48-linked AR ubiquitination as well as the mRNA expression and content of soluble KIT-L (sKitlg) and content of GDF9. These androgenic responses were attenuated by gonadotropin treatment in vivo. Growth of antral follicles from DHT-treated rats in vitro was significantly slower when compared to those of control but was significantly enhanced by exogenous GDF9, suggesting the DHT-induced antral follicular growth arrest is in part the results of GDF9 suppression. Our findings indicate how hyperandrogenism modulates RNF6 content and subsequently AR ubiquitination, resulting in antral follicle growth arrest in a chronically androgenized PCOS rat model.

Physiologic Course of Female Reproductive Function: A Molecular Look into the Prologue of Life.

The genetic, endocrine, and metabolic mechanisms underlying female reproduction are numerous and sophisticated, displaying complex functional evolution throughout a woman's lifetime. This vital course may be systematized in three subsequent stages: prenatal development of ovaries and germ cells up until in utero arrest of follicular growth and the ensuing interim suspension of gonadal function; onset of reproductive maturity through puberty, with reinitiation of both gonadal and adrenal activity; and adult functionality of the ovarian cycle which permits ovulation, a key event in female fertility, and dictates concurrent modifications in the endometrium and other ovarian hormone-sensitive tissues. Indeed, the ultimate goal of this physiologic progression is to achieve ovulation and offer an adequate environment for the installation of gestation, the consummation of female fertility. Strict regulation of these processes is important, as disruptions at any point in this evolution may equate a myriad of endocrine-metabolic disturbances for women and adverse consequences on offspring both during pregnancy and postpartum. This review offers a summary of pivotal aspects concerning the physiologic course of female reproductive function.

Comparison of gonadotropin-releasing hormone agonist with GnRH antagonist in polycystic ovary syndrome patients undergoing in vitro fertilization cycle: Retrospective analysis from a tertiary center and review of literature.

INTRODUCTION:Polycystic ovary syndrome (PCOS) is one of the most common infertility factor for which women are enrolled in in vitro fertilization (IVF) technique. In the recent years, gonadotropin releasing hormone antagonist protocol has emerged as the protocol of choice for controlled ovarian hyperstimulation in these patients.OBJECTIVES:The objective of the present study is to compare conventional long agonist protocol with fixed antagonist protocol in PCOS patients undergoing IVF cycle.MATERIALS AND METHODS:Retrospective analysis of 4 years data of a single center from northern India. Totally 81 patients who had long agonist protocol were compared with 36 patients with similar baseline characteristics who had antagonist protocol.RESULT:Total dose of gonadotropin required was significantly lower (P - 0.004) in the antagonist group. There was no significant difference in pregnancy rate or incidence of ovarian hyperstimulation syndrome between two groups. Cycle cancellation due to arrest of follicular growth was significantly higher in the antagonist group (P - 0.027).CONCLUSION:More randomized control trials and meta-analysis are required before replacing conventional long agonist protocol with antagonist protocol in patients with polycystic ovary syndrome.

Chemerin Suppresses Ovarian Follicular Development and Its Potential Involvement in Follicular Arrest in Rats Treated Chronically With Dihydrotestosterone

In the present study, we have investigated the cellular mechanisms of androgen-induced antral follicular growth arrest and the possible involvement of chemerin and its receptor chemokine-like receptor 1 (CMKLR1) in this process, using a chronically androgenized rat model. We hypothesize that hyperandrogenism induces antral follicle growth arrest via the action of chemerin and ovarian structural changes, resulting from granulosa cell and oocyte apoptosis and theca cell survival. Dihydrotestosterone (DHT) treatment resulted in increased expression of chemerin and CMKLR1 in antral follicles, absence of corpus luteum, and increased atypical follicles. Addition of chemerin to follicle cultures induced granulosa cell apoptosis and suppressed basal, FSH- and growth differentiation factor-9-stimulated follicular growth. DHT down-regulated aromatase expression and increased active caspase-3 content and DNA fragmentation in granulosa cells in vivo. These changes were accompanied by higher phosphatase and tensin homolog and lower phospho-Akt (Ser473) content in antral follicles and higher calpain expression and down-regulation of cytoskeletal proteins in atypical follicles, which were constituted predominantly of theca cells. DHT also activated granulosa cell caspase-3, decreased X-linked inhibitor of apoptosis protein, poly(ADP-ribose) polymerase, and phospho-Akt contents and induced apoptosis in vitro, responses readily attenuated by forced X-linked inhibitor of apoptosis protein expression. These findings are consistent with our hypothesis that antral follicular growth arrest in DHT-treated rats results from increased chemerin expression and action, as well as changes in follicular cell fate and structure, which are a consequence of dysregulated interactions of pro-survival and pro-apoptotic modulators in a cell-specific manner. Our observations suggest that this chronically androgenized rat model may be useful for studies on the long-term effects of androgens on folliculogenesis and may have implications for the female reproductive disorders associated with hyperandrogenism.

Differential Gene Expression in Granulosa Cells from Polycystic Ovary Syndrome Patients with and without Insulin Resistance: Identification of Susceptibility Gene Sets through Network Analysis

Polycystic ovary syndrome (PCOS) is a heterogeneous, genetically complex, endocrine disorder of uncertain etiology in women. Our aim was to compare the gene expression profiles in stimulated granulosa cells of PCOS women with and without insulin resistance vs. matched controls. This study included 12 normal ovulatory women (controls), 12 women with PCOS without evidence for insulin resistance (PCOS non-IR), and 16 women with insulin resistance (PCOS-IR) undergoing in vitro fertilization. Granulosa cell gene expression profiling was accomplished using Affymetrix Human Genome-U133 arrays. Differentially expressed genes were classified according to gene ontology using ingenuity pathway analysis tools. Microarray results for selected genes were confirmed by real-time quantitative PCR. A total of 211 genes were differentially expressed in PCOS non-IR and PCOS-IR granulosa cells (fold change≥1.5; P≤0.001) vs. matched controls. Diabetes mellitus and inflammation genes were significantly increased in PCOS-IR patients. Real-time quantitative PCR confirmed higher expression of NCF2 (2.13-fold), TCF7L2 (1.92-fold), and SERPINA1 (5.35-fold). Increased expression of inflammation genes ITGAX (3.68-fold) and TAB2 (1.86-fold) was confirmed in PCOS non-IR. Different cardiometabolic disease genes were differentially expressed in the two groups. Decreased expression of CAV1 (-3.58-fold) in PCOS non-IR and SPARC (-1.88-fold) in PCOS-IR was confirmed. Differential expression of genes involved in TGF-β signaling (IGF2R, increased; and HAS2, decreased), and oxidative stress (TXNIP, increased) was confirmed in both groups. Microarray analysis demonstrated differential expression of genes linked to diabetes mellitus, inflammation, cardiovascular diseases, and infertility in the granulosa cells of PCOS women with and without insulin resistance. Because these dysregulated genes are also involved in oxidative stress, lipid metabolism, and insulin signaling, we hypothesize that these genes may be involved in follicular growth arrest and metabolic disorders associated with the different phenotypes of PCOS.

Catechol-O-methyltransferase (COMT) single nucleotide polymorphisms and haplotypes are not major risk factors for polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is an endocrine disorder that affects 5–8% of reproductive age women. The primary features of PCOS are hyperandrogenemia, chronic anovulation and infertility. It has been suggested that defects in ovarian steroid metabolism contribute to the follicular growth arrest and abnormal production of ovarian steroid hormones that are characteristic of PCOS. 2-Methoxyestradiol (2-ME) is formed by the action of catechol-O-methyltransferase (COMT) on 2-hydroxyestradiol. COMT expression is increased in the follicles and ovarian stroma of women with PCOS. Moreover, 2-ME decreases granulosa cell proliferation and steroidogenesis, raising the possibility that ovarian dysfunction associated with PCOS is due, in part, to increased synthesis of 2-ME resulting from increased COMT activity. Four single-nucleotide polymorphisms (SNPs) (rs6269, rs4633, rs4818, rs4680) in the COMT gene characterize haplotypes, which are associated with large variations in COMT enzymatic activity. The aim of this study was to determine whether individual COMT SNPs and the COMT haplotypes are associated with PCOS using a family-based test of association and linkage. Additionally, we examined the relationships between COMT SNPs and haplotypes with quantitative variables usually assessed in the evaluation of women with PCOS. There were no significant correlations between genotype and total testosterone, non-SHBG bound testosterone and BMI. However, we found that the prolactin level in women with PCOS varied significantly with COMT haplotype, and suggest that this association reflects a genetic factor influencing the stress response. Our findings suggest that common variants and haplotypes of the COMT gene are not major contributors to risk for PCOS, but that COMT genotype may influence prolactin levels.

No difference in the aneuploidy rate of cleavage stage embryos in gonadotropin releasing hormone antagonists (GnRH-ant) cycles when compared to the agonists (GnRH-a) cycles

OBJECTIVE: Many studies have documented a slight decrease in the clinical pregnancy rates with the use of GnRH-ant when compared to GnRH-a cycles. It is possible that the transient arrest of follicular growth after GnRH-ant administration and subsequent rescue by exogenous gonadotrophins can lead to subtle genomic disruption. This study aims at comparing the aneuploidy rate of cleavage embryos during GnRH-ant cycles to the rate of aneuploidy in GnRH-a cycles.

Lutropin alpha, recombinant human luteinizing hormone, for the stimulation of follicular development in profoundly LH-deficient hypogonadotropic hypogonadal women: a review

Lutropin alpha, recombinant human luteinizing hormone, for the stimulation of follicular development in profoundly LH-deficient hypogonadotropic hypogonadal women: a review Bernd Th Krause1, Ralf Ohlinger2, Annette Haase31Center for Endocrinology and Reproductive Medicine, MVZ Uhlandstr, Berlin, Germany; 2Ernst-Moritz-Arndt-University, Department of Gynecology and Obstetrics, Greifswald, Germany; 3Uhlandstr. 162, 10719 BerlinAbstract: Hypogonadotropic hypogonadism is defined as a medical condition with low or undetectable gonadotropin secretion, associated with a complete arrest of follicular growth and very low estradiol. The main cause can be traced back to an irregular or absent hypothalamic GnRH secretion, whereas only a minority suffers from a pituitary disorder. The choice of treatment to reverse this situation is a pulsatile GnRH application or a direct ovarian stimulation using gonadotropin injections. The goal is to achieve a proper ovarian function in these cases for a short time to allow ovulation and chance of pregnancy. Since the pulsatile GnRH treatment lost its former importance, several gonadotropins are in use to stimulate follicular growth, such as urine-derived human menopausal gonadotropin, highly purified follicle stimulating hormone (FSH) or recombinant FSH, all with different success. The introduction of recombinant luteinizing hormone (LH) and FSH provided an opportunity to investigate the distinct influences of LH and FSH alone and in combination on follicular growth in monofollicular ovulation induction cycles, and additionally on oocyte maturation, fertilization competence of the oocyte and embryo quality in downregulated IVF patients. Whereas FSH was known to be indispensable for normal follicular growth, the role of LH remained questionable. Downregulated IVF patients with this short-term gonadotropin depletion displayed no advance in stimulation success with the use of recombinant LH. Patients with hypogonadotropic hypogonadism undergoing monofollicular stimulation for ovulation induction showed clearly a specific role and need for both hormones in normal follicular growth. Therefore, a combined stimulation with FSH and LH seems to be the best treatment choice. In the first half of the stimulation cycle the FSH dosage should exceed that of LH by 2:1, with an inverse ratio for the second half.Keywords: hypogonadotropic hypogonadism, ovarian stimulation, ovulation induction, recombinantluteinizing hormone

Lutropin alpha, recombinant human luteinizing hormone, for the stimulation of follicular development in profoundly LH-deficient hypogonadotropic hypogonadal women: a review.

Hypogonadotropic hypogonadism is defined as a medical condition with low or undetectable gonadotropin secretion, associated with a complete arrest of follicular growth and very low estradiol. The main cause can be traced back to an irregular or absent hypothalamic GnRH secretion, whereas only a minority suffers from a pituitary disorder. The choice of treatment to reverse this situation is a pulsatile GnRH application or a direct ovarian stimulation using gonadotropin injections. The goal is to achieve a proper ovarian function in these cases for a short time to allow ovulation and chance of pregnancy. Since the pulsatile GnRH treatment lost its former importance, several gonadotropins are in use to stimulate follicular growth, such as urine-derived human menopausal gonadotropin, highly purified follicle stimulating hormone (FSH) or recombinant FSH, all with different success. The introduction of recombinant luteinizing hormone (LH) and FSH provided an opportunity to investigate the distinct influences of LH and FSH alone and in combination on follicular growth in monofollicular ovulation induction cycles, and additionally on oocyte maturation, fertilization competence of the oocyte and embryo quality in downregulated IVF patients. Whereas FSH was known to be indispensable for normal follicular growth, the role of LH remained questionable. Downregulated IVF patients with this short-term gonadotropin depletion displayed no advance in stimulation success with the use of recombinant LH. Patients with hypogonadotropic hypogonadism undergoing monofollicular stimulation for ovulation induction showed clearly a specific role and need for both hormones in normal follicular growth. Therefore, a combined stimulation with FSH and LH seems to be the best treatment choice. In the first half of the stimulation cycle the FSH dosage should exceed that of LH by 2:1, with an inverse ratio for the second half.