Normal Ovarian Function Research Articles

Do cancer therapies damage the uterus and compromise fertility?

As cancer survival rates improve, understanding and preventing the adverse off-target and long-term impacts of cancer treatments, including impacts on fertility, have become increasingly important. Cancer therapy-mediated damage to the ovary and depletion of the primordial follicle reserve are well characterised. However, our knowledge of the full extent of damage to the rest of the female reproductive tract, in particular the uterus, is limited. Improving our understanding of the off-target effects of cancer therapies on the entire female reproductive tract is a critical step towards developing truly effective strategies to protect the fertility of cancer survivors. The objective of this narrative review was to critically evaluate the available literature regarding the capacity for the uterus to sustain a healthy pregnancy following exposure to radiotherapy or chemotherapy. The authors performed PubMed (Medline) searches using the following key words: uterus, cancer survivors, radiotherapy, chemotherapy, pregnancy outcome, fertility preservation, infertility. There were no limits placed on time of publication. Overall, there were major limitations to the current available literature, meaning that interpretations should be taken with caution. Despite these drawbacks, data suggest that the uterus may sustain off-target damage, with the extent of damage dependent on the type of cancer treatment and patient age. Specifically, uterine growth is stunted and resistant to hormone replacement therapy in prepubertal girls receiving abdominal, pelvic or whole-body radiotherapy. In contrast, females treated with radiotherapy post-puberty can benefit from hormone replacement therapy, as demonstrated by increased uterine volume and function. No live births have been reported in women previously exposed to radiotherapy after transplantation of cryopreserved ovarian tissue, even when menstruation returns. However, this technique has proven to be a successful fertility preservation method for women previously treated with chemotherapy. Obstetricians commonly report that women who maintain sufficient ovarian function can achieve pregnancy naturally following radiotherapy, but they have thin and/or fibrotic myometrium at delivery, compromising safe delivery and subsequent pregnancy. Furthermore, women exposed to either radiotherapy or chemotherapy have a higher prevalence of preterm birth and low birth weight infants, even in those with normal ovarian function or when oocyte donation is utilised. The mechanisms of potential uterine damage are poorly understood. While the myometrium, vasculature and endometrial progenitor cells are possibly targets, further studies are clearly required and well-controlled animal models could provide the best avenue for these types of future investigations. Female cancer survivors experience greater rates of early pregnancy loss and complications, suggesting that cancer therapy-induced damage to the uterus contributes to infertility. Despite clinical reports dating back to 1989, we highlight a surprising lack of detail in the literature regarding the precise nature and extent of off-target damage inflicted to the uterus in response to cancer therapies. Young women requiring cancer treatment, and the clinicians treating them, must be equipped with accurate information to aid informed decision-making regarding cancer treatment regimens as well as the development and use of effective fertility preservation measures. As the current literature on the impacts of cancer treatments is limited, we hope that our narrative review on this subject will stimulate more research in this important field.

Analysis of the MicroRNA Profile by Sequencing in Ovarian Granular Cells from Women Suffering Fluorosis with Reproductive Dysfunction

Excessive intake of fluoride may cause female reproductive dysfunction but pathological mechanism is unclear. The miRNAs in follicular fluid are a class of small non-coding RNAs from granulosa cells. The aim of this study is to examine the differential expressions of miRNAs in ovarian granulosa cells from women suffering from fluorosis and infertility. Five fluorosis women suffering infertility and 5 non-fluorosis (normal ovarian secretory function control) women were included as two groups. These two groups were indexed by serum and urine fluoride (F) levels as high F group and control group. The concentrations of estradiol (E2), progesterone (P), human chorionic gonadotropin (HCG), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) were measured by ELISA kits. The characteristics of menstruation from each woman were collected. The total RNA was isolated from granulosa cells for sequencing. The validation was completed by RT-qPCR. There was no significant difference between the two groups on age, the levels of E2, FSH, P, HCG, and LH in serum, and the characteristics of menstruation. A total of 37 miRNAs were upregulated and 40 miRNAs downregulated in follicular fluid from granulosa cells in the high F group compared with that in the control group. The expression of hsa-miR-29b increased in fluorosis patients identified by the miRNA sequence analysis. The distinct expression patterns of miRNAs in women requiring IVF from fluorosis area provide a new direction for molecular mechanism of fluorosis caused reproductive dysfunction.

Analysis of NR5A1 in 142 patients with premature ovarian insufficiency, diminished ovarian reserve, or unexplained infertility

Ovarian deficiency, including diminished ovarian reserve and premature ovarian insufficiency, represents one of the main causes of female infertility. Little is known of the genetic basis of diminished ovarian reserve, while premature ovarian insufficiency often has a genetic basis, with genes affecting various processes. NR5A1 is a key gene required for gonadal function, and variants are associated with a wide phenotypic spectrum of disorders of sexual development, and are found in 0.26–8% of patients with premature ovarian insufficiency. As there is some debate about the extent of involvement of NR5A1 in the pathogenesis of ovarian deficiency, we performed an in-depth analysis of NR5A1 variants detected in a cohort of 142 patients with premature ovarian insufficiency, diminished ovarian reserve, or unexplained infertility associated with normal ovarian function. We identified rare non-synonymous protein-altering variants in 2.8 % of women with ovarian deficiency and no such variants in our small cohort of women with infertility but normal ovarian function. We observed previously reported variants associated with premature ovarian insufficiency in patients with diminished ovarian reserve, highlighting a genetic relationship between these conditions. We confirmed functional impairment resulting from a p.Val15Met variant, detected for the first time in a patient with premature ovarian insufficiency. The remaining variants were associated with preserved transcriptional activity and localization of NR5A1, indicating that rare NR5A1 variants may be incorrectly curated if functional studies are not undertaken, and/or that NR5A1 variants may have only a subtle impact on protein function and/or confer risk of ovarian deficiency via oligogenic inheritance.

CD24: a marker of granulosa cell subpopulation and a mediator of ovulation

Granulosa cells (GCs) play a critical role in driving the formation of ovarian follicles and building the cumulus-oocyte complex surrounding the ovum. We are particularly interested in assessing oocyte quality by examining the detailed gene expression profiles of human cumulus single cells. Using single-cell RNAseq techniques, we extensively investigated the single-cell transcriptomes of the cumulus GC populations from two women with normal ovarian function. This allowed us to elucidate the endogenous heterogeneity of GCs by uncovering the hidden GC subpopulation. The subsequent validation results suggest that CD24(+) GCs are essential for triggering ovulation. Treatment with human chorionic gonadotropin (hCG) significantly increases the expression of CD24 in GCs. CD24 in cultured human GCs is associated with hCG-induced upregulation of prostaglandin synthase (ARK1C1, PTGS2, PTGES, and PLA2G4A) and prostaglandin transporter (SLCO2A1 and ABCC4) expression, through supporting the EGFR-ERK1/2 pathway. In addition, it was observed that the fraction of CD24(+) cumulus GCs decreases in PCOS patients compared to that of controls. Altogether, the results support the finding that CD24 is an important mediator of ovulation and that it may also be used for therapeutic target of ovulatory disorders.

Effect of radiotherapy on ovarian function in patients undergoing radical surgery for cervical cancer

Objective The correlation between dosimetric parameters of transposed ovary and different clinical ovarian functional status was assessed in young patients with cervical cancer who needed adjuvant radiotherapy after radical resection of the ⅠB1-ⅡA2 phase of preserved and transposed ovaries. Methods The function of transposed ovary and relevant clinical symptoms in 86 patients before and 2 years after treatment between 2015 and 2017 were retrospectively analyzed, and the correlation between the dosimetric parameters and functional status of transposed ovaries during radiotherapy was evaluated. Different in vitro measures were adopted to protect the transposed ovaries during postoperative radiotherapy including 68 cases of IMRT or VMAT and 18 cases of two-dimensional and other central radiotherapy. Results The nearest distance between ovary and PTV was negatively correlated with the ovarian dose ≥V5 Gy (P=0.025). V8 Gy and Dmean were positively correlated with FSH after treatment (P=0.011, 0.020). The larger the volume of V8 Gy and the large Dmean, the higher the FSH, the worse the ovarian function. In two-dimensional technology, the ovarian dose ≥V5 Gy was significantly lower than that in three-dimensional technique. The average age of those with normal ovarian function after treatment was 33.4 years, whereas the average age of women with ovarian failure was 39.6 years (P=0.007). The number of preserved ovaries and whether synchronous chemotherapy was delivered were similar in patients with different ovarian status, which were correlated with the levels of FSH and E2 (Estradiol) before treatment, that is, the higher the level of FSH before treatment, the lower the E2 of ovarian FSH after treatment, and the higher the level of FSH after treatment, the lower the level of ovarian E2. Patients who retained their ovaries before treatment but suffered from ovarian failure received neoadjuvant chemotherapy with a slightly higher age. Conclusions Age, V8 Gy and Dmean of the transposed ovary, the shortest distance between transposed ovary and PTV, whether neoadjuvant chemotherapy was delivered before surgery and radiotherapy technique affect the protection of the function of transposed ovary. Key words: Cervical neoplasm/radiotherapy; Ovarian transposition; Ovarian function

Reproductive Health in Xeroderma Pigmentosum: Features of Premature Aging.

To assess the age at menarche and menopause of women with xeroderma pigmentosum, a DNA repair disease with premature aging, in a longitudinal natural history study. We conducted a natural history study that reviewed medical records for gynecologic and reproductive health of all female patients with xeroderma pigmentosum aged older than 9 years examined at the National Institutes of Health (NIH). We performed gynecologic and laboratory examinations on a subset of the patients. Women in a second subset, who could not be examined, were interviewed using a questionnaire. Women who were deceased or lost to follow-up formed a third subset. Sixty females with xeroderma pigmentosum aged older than 9 years (median 29 years, range 10-61 years) were evaluated at the NIH from 1971 to 2018. Of these 60, 31 had history, questionnaire, record review, and gynecologic evaluation; 14 had record review and questionnaire interview by telephone; and 15 had only NIH record review. Menarche in females with xeroderma pigmentosum occurred at a median age of 12.0 years (range 9-17 years), which was comparable with the U.S. general population. Among the 18 patients with menopause, the median age at menopause of 29.5 years (range 18-49.5 years) was more than 20 years younger than in the U.S. general population (52.9 years). Premature menopause (before age 40 years) occurred in 14 of the 45 (31%) women aged 18 years or older, and primary ovarian insufficiency was documented in nine of them. There were 32 live births among 21 of the women, five of whom subsequently developed premature menopause. Females with xeroderma pigmentosum in our study had a normal age at menarche and were fertile but had increased incidence of premature menopause. Premature menopause, a symptom of premature aging, should be considered for gynecologic and reproductive health as well as implicating DNA repair in maintaining normal ovarian function. ClinicalTrials.gov, NCT00001813.

Novel HARS2 missense variants identified in individuals with sensorineural hearing impairment and Perrault syndrome

Biallelic variants in HARS2 have been associated with Perrault syndrome, characterized by sensorineural hearing impairment and premature ovarian insufficiency. Here we report three novel families, compound heterozygous for missense variants in HARS2 identified by next-generation sequencing, namely c.172A > G (p.Lys58Glu) and c.448C > T (p.Arg150Cys) identified in two sisters aged 13 and 16 years and their older brother, c.448C > T (p.Arg150Cys) and c.980G > A (p.Arg327Gln) identified in a seven year old girl, and finally c.137T > A (p.Leu46Gln) and c.259C > T (p.Arg87Cys) identified in a 32 year old woman. Clinically, all five individuals presented with early onset, rapidly progressive hearing impairment. Whereas the oldest female fulfilled the criteria of Perrault syndrome, the three younger females, aged 7, 13 and 16, all had apparently normal ovarian function, apart from irregular menstrual periods in the oldest female at age 16. The present report expands the list of HARS2 variants and helps gain further knowledge to the phenotype.

Rapid In Situ Action of Estradiol 17β on Ion Transporter Function in Brain Segments of Female Mozambique Tilapia ( Oreochromis mossambicus Peters)

Being the principal estrogen, estradiol 17I² (E 2 ) is essential for normal ovarian function in the vertebrates including fishes. Besides its primary role in reproduction, E 2 is also known for its role in many other physiological processes including water and mineral balance. However, it is uncertain, how E 2Â regulates ion-specific ATPases that drive Na + , K + , H + , Ca 2+ and Mg 2+ transport in fish brain. We, therefore, examined the short-term in situ action of E 2 on ion transporter function in the brain segments of freshwater female Mozambique tilapia Oreochromis mossambicus . Tilapia were perfused with increasing doses of E 2 (10 -9 , 10 -8 and 10 -7 M) for 20 min and sampled for determining Na + /K + -ATPase, H + -ATPase, Ca 2+ -ATPase, and Mg 2+ -ATPase activities in the prosencephalon (PC), mesencephalon (MC) and metencephalon (MeC) segments of brain. Dose-dependent increase in Na + /K + - and Ca 2+ -dependent transporter activities after E 2 perfusion were found in PC. In MC, E 2 treatment, however, produced significant increase in Mg 2+ , Ca 2+ and H + transport activities in mitochondria but decreased Na + /K + - and I½H + transporter activities. On the contrary, in MeC, E 2 administration while producing increase in Na + /K + -, mitochondrial- and I½H + -transport, lowered cytosolic and mitochondrial Ca 2+ transport. Taken together, the data indicate that E 2 has rapid and direct action on ion transporter function that corresponds to the differential activation/inactivation of neuronal clusters in the brain segments of female freshwater tilapia.

Ovarian transposition before radiotherapy in cervical cancer patients: functional outcome and the adequate dose constraint

BackgroundThe data regarding a transposed ovary in intensity-modulated radiotherapy (IMRT) are not sufficient. Here we aim to investigate the adequate dose constraint of ovarian transposition before radiotherapy in cervical cancer patients.MethodsThis was a retrospective analysis of 118 patients with cervical cancer who received a radical hysterectomy and ovarian transposition before pelvic irradiation from April 2012 to July 2017. A total of 105 patients underwent IMRT with a limited radiation dose to the ovaries; 48 of these patients received unilateral ovary limitation, while 57 received bilateral ovary limitations. Patient follow up regarding sex hormone levels (estrogen [E2], follicle stimulating hormone [FSH]) and menopausal symptoms was completed one year after their radiation therapy.ResultsA total of 41 out of 105 patients (39.0%) who underwent IMRT with a limited radiation dose to the ovaries preserved their normal ovarian function. The cutoff dose of comparatively lower side ovarian maximum dose was 9.985Gy and the cutoff of mean dose was 5.32Gy. The optimal dose–volume constrains to ovaries was V5.5 < 29.65%. Age ≤ 38 (P = 0.001) was an independent predictors of ovarian function, while limited ovarian side numbers were excluded.ConclusionUsing IMRT, preservation of ovarian function was possible when the limited dose was as low as possible to the ovaries regardless of bilateral or unilateral limitation to the ovaries. The ovarian maximum dose of less than 9.985Gy, the mean dose less than 5.32Gy and V5.5 < 29.65% could be better at preventing ovarian dysfunction. Patients younger than 38 years old were more likely to keep normal ovarian function while limited ovarian side numbers did not appear to exert an obvious effect.

Does polycystic ovary syndrome affect morphokinetics or abnormalities in early embryonic development?

ObjectivesThis study aimed to investigate whether oocyte retrieval from PCOS patients affects the morphokinetics or the incidence of abnormalities in early embryonic development, using time-lapse imaging analysis.MethodsThis was a prospective study in total of 39 PCOS patients and 67 women with normal ovarian function, underwent a GnRH antagonist protocol of controlled ovarian stimulation and fertilization by ICSI. 402 zygotes from the PCOS group and 449 zygotes from the control group were observed by time-lapse monitoring for 48 h following sperm injection.ResultsPatients with PCOS showed a significantly higher number of retrieved oocytes, and a significantly higher number of metaphase II oocytes per cycle than that of the non-PCOS group (18.85 ± 9.41 vs. 11.48 ± 5.51 and 14.97 ± 7.43 vs. 9.51 ± 4.7, respectively). However, oocyte maturation rate and morphologically assessed embryo quality did not differ between two groups. After 48 h of the embryo culture using time-lapse video recording, most of the embryos in both groups had reached the 4-cell stage (353/449 vs. 314/402 embryos). There were no significant differences between the control and PCOS group regarding the time-points of the successive events in early embryonic development from the appearance of 2 polar bodies to the 6-cell stage. However, the percentage of t2 stages which fell in “optimal range” (>24 h and ≤28 h) was significantly lower in the PCOS group than in the control group (37.8% vs. 48.1%, P = 0.004). The proportion of embryos manifesting abnormal fertility, multinucleation, direct cleavage and reverse cleavage were not significantly different between the two groups.ConclusionOur data indicated that PCOS does not affect the morphokinetics or incidence of abnormalities in early embryonic development.

Review of Hormone Replacement Therapy in Girls and Adolescents with Hypogonadism.

Girls with either hypo- or hypergonadotropic hypogonadism need treatment with estrogens to initiate puberty and maintain a normal hormonal milieu. The focus of this review is hormone replacement treatment in girls with hypogonadism, to initiate and progress through puberty, and to maintain a healthy hormonal milieu in women. It also addresses what is known in the literature regarding estrogen levels in girls and women, instructive cases, practical tables for reference and application, and thoughts on future directions in this area. It represents a thorough literature review with author opinions and recommendations. Girls with normal ovarian function begin puberty on average at 10.5years old, although there is variation according to ethnicity and degree of excess weight gain. The aim of estrogen therapy to initiate puberty is to mimic normal onset and rate of progression. On the basis of the currently available literature, when a diagnosis of hypogonadism is established, we recommend initiating treatment between age 11 and 12years of age, with dose increases approximately every 6months until adult levels are reached. In some situations, treatment may be delayed to allow time for diagnosis or permit more time for linear growth, or address unique risks found in girls treated for various cancers or blood disorders. When adult dosing is reached, progestins are also used to protect uterine health. This can be combined sequentially, allowing regular menstruation, or combined continuously when menstrual bleeding is not preferred. Treatment is continued until the average age of menopause, again with various considerations for longer or shorter duration on the basis of risk-benefit ratios. Transdermal estrogens are considered the most physiologic replacement and theoretically might have fewer associated risks. We review what is known about risks and outcomes and areas for future research.

A cohort study of both human menopausal gonadotropin (HMG) and recombinant luteinizing hormone addition at early follicular stage in in vitro fertilization outcome: A STROBE-compliant study.

At present, the precise role of human menopausal gonadotropin (HMG) and recombinant luteinizing hormone (rLH) supplementation at an early time of follicular phase on in vitro fertilization (IVF)/intra cytoplasmatic sperm injection (ICSI) outcomes remains uncertain.Here infertile women of normal ovarian function undergoing their first cycle of IVF/ICSI were studied and were randomly allocated into 3 groups. Group 1, ovarian stimulation with 150IU recombinant follicle-stimulating hormone (FSH) alone. Group 2, patients received 75IU rFSH and 75IU HMG. Group 3 patients were given 150IU rFSH and 75IU rLH.There were no significant differences in the clinical characteristics, ovarian response, the biochemical, clinical and ongoing pregnancy rates among the 3 groups. No significant differences were found in biochemical, clinical and ongoing pregnancy rates between the patients whose LH levels were lower than 0.75 mIU/ml and those above this threshold in group 1. Furthermore, there were also no significant differences in biochemical, clinical and ongoing pregnancy rates among the 3 group patients whose LH level lower than 0.75 mIU/ml.The results showed that either the addition of HMG or rLH supplementation at an early time of follicular phase produce no significant benefit on IVF outcome in patients with normal ovarian function.

SUN-194 Erasing the Methyl Mark: Mechanism of Androgen Action in the Ovary

Androgen actions through androgen receptor (AR) are critical for follicle growth and female fertility. While excess androgen level leads to polycystic ovary syndrome (PCOS), a certain amount of direct androgen action is critical for normal ovarian function. Thus, with respect to androgen actions in the ovary, balance is key. However, how androgens regulate ovarian physiology or its underlying mechanism is still poorly understood. Androgen actions are mediated through ‘nuclear/genomic’ or ‘extra-nuclear/non-genomic’ actions of AR. Recently we reported that androgens through both the extra-nuclear and nuclear pathways inhibit the expression and activity of Ezh2, (a histone methyl-transferase involved in gene repression mark, H3K27me3) to induce different genes that are imperative for ovarian function. Here, we further extended our study to elucidate other histone modifications induced by androgens and its downstream physiological effect in follicular development. In addition to Ezh2, we have found that androgens also regulated the expression of a histone demethylase called Jumonji domain containing protein 3 (JMJD3/KDM6B), that is responsible of removing H3K27me3 mark. Results showed that DHT in GCs increased hypoxia-inducible factor 1 alpha (HIF1α) protein levels through the PI3K/Akt pathway, which in turn induced JMJD3 expression. We found that DHT had not affect HIF1α mRNA but increased HIF1α protein levels. This effect of DHT on HIF1α protein level was blocked by flutamide (AR inhibitor), LY294002 (PI3K inhibitor) as well as cyclohexamide (protein translational inhibitor). Interestingly the 5’ prime end of JMJD3 have HIF1α response elements (HRE) and it is well established that several JMJDs are induced by HIF1α under both normal conditions and hypoxic stress. Our studies showed that DHT treatment induced JMJD3 expression and decreased H3K27me3 levels. Inhibition of AR, PI3K/AKT pathway and siRNA-mediated knockdown of HIF1α attenuated androgen-induced expression of JMJD3. Moreover, ChIP studies showed increased binding of HIF1α on the JMJD3 promoter region with DHT treatment. In our efforts to understand the downstream effects of androgen-induced JMJD3 expression, we found Dicer1 was upregulated upon DHT treatment to mouse granulosa cells. Given the critical role of Dicer in follicular development, ovarian physiology and female fertility, in general, along with the importance of androgens in ovarian physiology, we propose that regulation of Dicer is one of the underlying mechanisms of androgen actions in the ovary. Androgens in a transcription-independent manner, through the PI3K/Akt signaling pathway, enhance HIF1α protein synthesis under normoxic conditions which in turn regulate the expression of Dicer through JMJD3. Altogether, this study provides a mechanistic insight of androgen actions in ovarian function by epigenetic modulation.

SAT-214 Satoyoshi Syndrome: A Rare Cause of Premature Ovarian Failure

Introduction: Satoyoshi syndrome is a rare multisystemic disorder characterized by painful muscle spasms, alopecia universalis and diarrhea. Endocrine abnormalities associated with this syndrome include delayed puberty, primary amenorrhea and hypoplastic uterus and ovaries. It was first reported in 1967 in Japan with the majority of the cases affecting the Asian population, but cases in other ethnic groups have also been reported. It is a sporadic disease, with female to male ratio of two to one and mean age of onset of 10 years. The underlying etiology is unknown and is thought to be of an autoimmune nature. Case presentation: We report a case of a 27 year old Asian American female with Satoyoshi syndrome that was diagnosed at age 12, when she started to suffer from diffuse muscle spasms mostly in the neck, jaw, and lower extremities along with diarrhea and loss of hair. She received intravenous immunoglobulins (IVIG) and systemic corticosteroid therapy in the past and is currently on annual Rituximab infusion. She also takes Carbamazepine and Imodium for muscle contractions and diarrhea respectively. She had menarche at age 11 and recalls having one menstrual period only. This was followed by amenorrhea and hot flashes, both resolved after starting estrogen and progesterone therapy. She was also noted to have low bone density for which she takes vitamin D. For the past few years, she has been on Norethindrone acetate therapy and reported regular periods on it. She had normal anti-müllerian hormone level two years ago. On physical examination, she had diffuse muscle contractions most notably in the muscles of mastication. She also had global thinning of the hair without patches of hair loss. Breast examination showed Tanner stage V for breast development and pelvic examination showed Tanner stage IV for pubic hair growth. The remainder of her physical exam was normal. Her latest lab work showed luteinizing hormone level of 7.15 mIU/ml, follicular stimulating hormone of 9.60 mIU/ml and estradiol level of 33.4 pg/ml suggestive of normal ovarian function. Conclusion: Our patient had all the characteristic features of Satoyoshi syndrome except premature ovarian failure which is quite unusual for this syndrome. Fewer than 60 cases have been reported worldwide so far making it a diagnostic challenge.

SUN-027 Androgen Induces Its Own Receptor Expression in Human Granulosa Cells

Androgens are very important in the regulation of normal ovarian function, as well as pathologies such as ovarian insufficiency and polycystic ovary syndrome. Androgen receptors are present in all major cell types of the ovarian follicle, and are most abundant in granulosa cells where they enhance proliferation during the follicular phase of the reproductive cycle. However, the molecular pathways mediating androgen actions in the ovary are not clearly delineated. We previously found that in human prostate cancer cells, androgen receptor (AR) is highly active in the extra-nuclear compartment, where it stimulates kinase signaling and enhances growth factor-induced cell proliferation. Further, these novel extra-nuclear actions of AR require paxillin, a cytoplasmic adapter protein that is implicated in cell growth, cytoskeletal remodeling and kinase signaling in many tissues. Here, we investigated the role of paxillin in androgen-regulated processes in human granulosa cells using the KGN cell line. We show that 25 nM dihydrotestosterone (DHT) treatment greatly induces the protein expression of AR, while its mRNA is unaffected. This induction is apparent as early as one hour after DHT treatment and the AR protein further accumulates with continued exposure. Importantly, near-depletion of paxillin with siRNA significantly blunted this induction, as well as basal AR protein expression. Our results indicate that androgens are capable of generating a positive feedback loop in granulosa cells at the level of their receptor, potentially greatly amplifying their actions. This occurs independently of AR gene transcription and requires paxillin, adding to the growing body of evidence that paxillin mediates extra-nuclear steroid signaling in various endocrine tissues. Our ongoing studies are aimed at identifying the mechanism of AR protein accumulation by DHT and clarifying paxillin involvement in this and other processes in the ovary.

Ovarian failure induced by 4-vinylcyclohexene diepoxide worsens the autonomic cardiovascular response to chronic unpredictable stress in rats

AimsAfter menopause, women are more responsive to stress and more prone to exhibit hypertension, which elevates the risk of cardiac diseases. This vulnerability is due, in part, to the decline of ovarian steroids plasma levels. The 4-vinylciclohexane diepoxide (VCD) causes a gradual depletion of ovarian follicles causing loss of the normal ovarian function and a hormonal profile comparable to menopause in humans. We aimed to verify whether the ovarian failure (OF) worsens the cardiovascular autonomic response to stress. Main methodsRats were treated with VCD (160 mg/kg) or oil for 15 days, exposed to chronic unpredictable stress (CUS) for 10 days and studied 80 and 180 days after VCD treatment. Key findings80 days after VCD-treatment, stressed rats showed increased sympathetic nerve activity, reduced parasympathetic activity and an increase in the overall spontaneous baroreflex sensitivity (BRS). 180 days after VCD treatment, BRS was impaired and the vascular sympathetic activity was increased, independently of stress exposure. SignificanceNeither 80 nor 180 days after the onset of VCD-treatment the hypertensive effects of stress were enhanced in rats. However, OF led to a worsening on different aspects of the cardiovascular response to stress, which can cause cardiovascular complications when associated with ovarian aging.

The Role of microRNAs in Ovarian Granulosa Cells in Health and Disease.

The granulosa cell (GC) is a critical somatic component of the ovary. It is essential for follicle development by supporting the developing oocyte, proliferating and producing sex steroids and disparate growth factors. Knowledge of the GC's function in normal ovarian development and function, and reproductive disorders, such as polycystic ovary syndrome (PCOS) and premature ovarian failure (POF), is largely acquired through clinical studies and preclinical animal models. Recently, microRNAs have been recognized to play important regulatory roles in GC pathophysiology. Here, we examine the recent findings on the role of miRNAs in the GC, including four related signaling pathways (Transforming growth factor-β pathway, Follicle-stimulating hormones pathway, hormone-related miRNAs, Apoptosis-related pathways) and relevant diseases. Therefore, miRNAs appear to be important regulators of GC function in both physiological and pathological conditions. We suggest that targeting specific microRNAs is a potential therapeutic option for treating ovary-related diseases, such as PCOS, POF, and GCT.

Proteomic analysis of mouse ovaries during the prepubertal stages

Postnatal folliculogenesis, primordial follicle activation and follicular development at early stage are important for normal ovarian function and fertility, and a comprehensive understanding of this process under physiological condition is necessary. To observe the regulation and mechanism of ovarian follicle development during the prepubertal stages, we collected the mouse ovaries from three time points, including 1 day, 7 days, and 4 weeks after birth. We then performed a proteomic analysis using tandem mass tags (TMT) labeling combined with a two-dimensional liquid chromatography-tandem mass spectrometry (2D LC-MS/MS) technique. A total of 706 proteins were determined to be significant differential abundance (P-SDA). Sixty upregulated proteins and 12 downregulated proteins that were P-SDA and 3 significant KEGG pathways (P < 0.05) were found at 7 days vs. 1 day after birth, while 237 upregulated proteins, 271 downregulated proteins and 42 significant KEGG pathways were found for 4 weeks vs. 7 days after birth. Some vital genes (Figla, Ooep, Padi6, Zp3, Hsd3b1, cyp11a1), key pathways (ECM-receptor interaction, focal adhesion, ovarian steroidogenesis, complement and coagulation cascades, PI3K/Akt/mTOR), and metabolic regulation (energy metabolism, lipid metabolism, metal ion metabolism) were found to be related to the postnatal folliculogenesis, primordial follicle activation and follicular development. Finally, qRT-PCR and western blotting verified some vital genes and further elucidated the developmental process of follicles, and the results may contribute to the understanding of the formation and activation of primordial follicle and follicular development.Significance: This study offers the first proteomic insights into mechanisms of follicle development under physiological condition during the prepubertal stages. By comparing P-SDA of mouse ovaries during various period of age, our data reveals that the regulation of primordial follicle formation and activation is significantly different from that of follicular development. These findings demonstrate that many unique molecular mechanisms underlie ovarian development could be used for ovarian disease research.

The incidence of unexpected poor ovarian response in Chinese young women.

Unexpected poor ovarian response (UPOR) is a problem for both clinicians and infertile couples, because our understanding of this situation is limited. This article investigated incidence of UPOR in women <35 years with normal ovarian reserve function with further analysis.This is a retrospective study, which included 567 women who accepted their first IVF-ET/ICSI. Based on the number of oocytes retrieved, clinic pregnancy rate of fresh cycle, and cycle cancellation rate of fresh cycle, the included cycles were divided into three groups namely unexpected poor ovarian response (UPOR) (n = 120), for which number of oocytes retrieved was not more than 6; normal ovarian response (NOR) (n = 223), for which number of oocytes retrieved was between 7 and 12; and unexpected high ovarian response (UHOR) (n = 224), for which the number of oocytes retrieved was 13 at least. The comparisons of clinical outcomes and correlated hormones among groups were carried out.The incidence of UPOR in Chinese women is 21.16%. Patient age (χ2 = 6.177, P = .0129), basic FSH (χ2 = 20.585, P < .0001), basic LH (χ2 = 11.689, P = .0006), and AFC (χ2 = 8.053, P = .0045) might be helpful for diagnosis of UPOR.The basic evaluation of ovarian function may no longer be simplified into normal and abnormal ovarian reserve function; rather, by using a detailed numerical analysis, such as basal FSH and LH levels, the ovarian response to ovulation induction may be predicted to some extent.

Abnormal expressions of ADAMTS-1, ADAMTS-9 and progesterone receptors are associated with lower oocyte maturation in women with polycystic ovary syndrome.

ADAMTS-1 and 9 play a crucial role in the ovulation and their altered levels may play a role in the pathogenesis of polycystic ovary syndrome (PCOS). The aim of this study was to assess ADAMTS-1 and 9 expression and their correlation with the oocyte quality and maturity in the cumulus cells (CCs) of PCOS patients and normovulatory women during an IVF procedure. Expression of ADAMTS-1 and 9 and progesterone receptors (PRs) in the CCs containing MII and germinal vesicle (GV) oocytes of 37 PCOS patients and 37 women with normal ovulatory function who underwent IVF treatment was evaluated using qRT-PCR. Moreover, correlation between ADAMTS-1 and 9 expression and oocyte quality were also investigated. mRNA expression levels of ADAMTS-1 and ADAMTS-9 were significantly reduced in the women with PCOS compared to the normovulatory women. ADAMTS-1 and ADAMTS-9 mRNA expression levels in the CCs showed a considerable correlation. Lower expression levels of ADAMTS-1 and ADAMTS-9 in PCOS patients were strongly correlated with diminished oocyte maturation. There was a remarkable association between ADAMTS-1 and ADAMTS-9 mRNA expression levels and oocyte quality. PRs (PRA and PRB) were dramatically decreased in PCOS patients when compared with the control group. The results of the present study indicated that ADAMTS-1 and ADAMTS-9 as well as PRs are downregulated in the human CCs in PCOS patients, which could be associated with impaired oocyte maturation and may result in a lower oocyte recovery and oocyte maturity rates, as well as lower fertilization rate.