Absence of mutations in the coding regions of follicle-stimulating hormone receptor gene in Singapore Chinese women with premature ovarian failure and polycystic ovary syndrome.

Normal gonadal function is critically dependent on the integrity of pituitary-gonadal axis, where follicle-stimulating hormone (FSH) plays a key role. In the female, FSH is required for follicular growth, estrogen production and oocyte maturation. Its function is mediated by its specific receptor (FSHR), and defective FSHR has been shown to affect folliculogenesis and ovarian function. In this study, we screened the entire coding region of FSHR gene for pathogenic mutations in women with premature ovarian failure (POF) (n = 16) and polycystic ovary syndrome (PCOS) (n = 124) and found no mutations in these patients. Two known polymorphisms, Thr307Ala and Ser680Asn showed similar distributions of the allelic variations and protein isoforms in PCOS and normal control subjects (n = 236). It appears from this study that mutations in the coding regions of FSHR gene are not a causative factor of the above clinical manifestations in Chinese Singapore women.     

Regulation of the genes for insulin-like growth factor (IGF) I and II and their receptors by steroids and gonadotropins in the ovary

Insulin-like growth factors (IGFs) I and II are two single-chain polypeptide hormones that are structurally related to each other and to proinsulin. Among the large number of growth factors involved in ovarian physiology, IGF-I and IGF-II are considered to be important progression factors for ovarian follicular development. To explore the ovarian expression of IGF-I, IGF-II and their receptor genes, a solution hybridization/RNase protection assay, was used. IGF-I mRNA was seen in the granulosa cells, and IGF-II mRNA in the theca-interstitial compartment. To study the hormonal regulation of the IGF-I and IGF-II gene, immature (21-day-old) hypohysectomized rats were treated with FSH (10 μg/day),GH (150 μg/day) and diethylstilbestrol (DES subcutaneous implant/5 days). Estrogen differentially regulated ovarian IGF-I and IGF-II gene expression. In concert with GH, estrogen up-regulated ovarian IGF-I mRNA, but significantly decreased hepatic IGF-I gene expression. Both IGF receptors (type I and type II) as well as the insulin receptor gene, were expressed in both ovarian cells. The expression of the type IIGF receptor gene (but not the type II IGF gene) was up-regulated by FSH and estrogen in vivo. In conclusion, these studies may serve to better understand the auto paracrine role of IGF, and their receptors in the pathophysiology of follicle recruitment, oocyte maturation and potentially embryo development.     

Ovarian maturation of Penaeus subtilis (Decapoda: Penaeidae): A new insight to describe oocyte development and somatic structures

We observed the presence of follicular cells (FC) in the ovaries of Penaeus subtilis (n = 1198), which led us to classify the development of germ cells into six phases: oogonia, previtellogenic oocytes, primary and secondary vitellogenic oocytes, mature oocytes and atretic oocytes. The FC changes their shape according to the development of germ cells and showed a different distribution along the ovary, which allowed differentiating vitellogenic oocytes into primary and secondary. We also observed that the postovulatory follicles (POF) are composed of follicular cells. The presence of POF in penaeids ovaries is rarely reported, but allows the differentiation between spent and resting stages, commonly grouped in reproductive biology research. Furthermore, observation of ovarian lining was useful to differentiate immature females from females that had spawned at least once. Thus, ovarian development was classified into six stages: immature, early developing, advanced developing, ripe, spent and resting. The distribution and shape variations of FC, ovarian lining features and presence of POF were considered crucial for the classification of ovarian maturation stages. The methods developed here may improve estimates of their reproductive cycle, size at first maturity and spawning season, which are important variables in future studies of the reproductive dynamics.     

卵巢早衰候选基因的突变研究进展

卵巢早衰病因复杂, 多数病例病因不明。在已知的病因中, 遗传因素非常重要。卵巢早衰不但可导致不孕症, 患者的低雌激素水平还增加了患骨质疏松症和冠心病的危险。目前临床上的主要治疗措施包括激素替代治疗和供卵治疗不孕症等, 但效果都不理想。鉴定卵巢早衰的致病基因是从根本上治疗和预防该病的基础。目前已发现在X染色体上和常染色体上多个基因与之相关, 文章综述了近年来对卵巢早衰候选基因的突变分析情况, 旨在从分子水平分析研究卵巢早衰的发病机制提供基础。     

Epigenetic control of the critical region for premature ovarian failure on autosomal genes translocated to the X chromosome: a hypothesis

Chromosomal rearrangements in Xq are frequently associated to premature ovarian failure (POF) and have contributed to define a POF “critical region” from Xq13.3 to Xq26. Search for X-linked genes responsible for the phenotype has been elusive as most rearrangements did not interrupt genes and many were mapped to gene deserts. We now report that ovary-expressed genes flanked autosomal breakpoints in four POF cases analyzed whose X chromosome breakpoints interrupted a gene poor region in Xq21, where no ovary-expressed candidate genes could be found. We also show that the global down regulation in the oocyte and up regulation in the ovary of X-linked genes compared to the autosomes is mainly due to genes in the POF “critical region”. We thus propose that POF, in X;autosome balanced translocations, may not only be caused by haploinsufficiency, but also by a oocyte-specific position effect on autosomal genes, dependent on dosage compensation mechanisms operating on the active X chromosome in mammals.     

Physical mapping of nine Xq translocation breakpoints and identification of XPNPEP2 as a premature ovarian failure candidate gene

Women with balanced translocations between the long arm of the X chromosome (Xq) and an autosome frequently suffer premature ovarian failure (POF). Two "critical regions" for POF which extend from Xq13-->q22 and from Xq22-->q26 have been identified using cytogenetics. To gain insight into the mechanism(s) responsible for ovarian failure in women with X;autosome translocations, we have molecularly characterized the translocation breakpoints of nine X chromosomes. We mapped the breakpoints using somatic cell hybrids retaining the derivative autosome and densely spaced markers from the X-chromosome physical map. One of the POF-associated breakpoints in a critical region (Xq25) mapped to a sequenced PAC clone. The translocation disrupts XPNPEP2, which encodes an Xaa-Pro aminopeptidase that hydrolyzes N-terminal Xaa-Pro bonds. XPNPEP2 mRNA was detected in fibroblasts that carry the translocation, suggesting that this gene at least partially escapes X inactivation. Although the physiologic substrates for the enzyme are not known, XPNPEP2 is a candidate gene for POF. Our breakpoint mapping data will help to identify additional candidate POF genes and to delineate the Xq POF critical region(s).     

卵巢早衰患者血清抑制素B、抗苗勒管激素及性激素水平与子宫动脉血流参数的相关性研究

目的:探讨卵巢早衰(POF)患者血清抑制素B(INHB)、抗苗勒管激素(AMH)及性激素水平与子宫动脉血流参数的相关性。方法:选择2018年5月至2020年5月期间我院收治的126例POF患者(POF组)和同期于我院进行体检的85例健康女性志愿者(对照组)。检测所有研究对象血清INHB、AMH以及促黄体生成素(LH)、促卵泡激素(FSH)、雌二醇(E2)水平,经阴道多普勒超声检测子宫动脉血流参数[收缩期峰值流速(PSV)、舒张末期流速(EDV),血流阻力指数(RI)、搏动指数(PI)]。Pearson相关性分析POF患者血清INHB、AMH、LH、FSH、E2水平与PSV、EDV、RI、PI的相关性。结果:POF组血清INHB、AMH、E2水平、PSV、EDV低于对照组(P<0.05),LH、FSH水平、RI、PI高于对照组(P<0.05)。Pearson相关性分析结果显示POF患者血清INHB、AMH、E2水平与PSV、EDV呈正相关(P<0.05),与RI、PI呈负相关(P<0.05),LH、FSH与PSV、EDV呈负相关(P<0.05),与RI、PI呈正相关(P<0.05)。结论:POF患者血清INHB、AMH、E2水平降低,LH、FSH水平升高,血清INHB、AMH和性激素与子宫动脉血流受限有关。     

A human homologue of the Drosophila melanogaster diaphanous gene is disrupted in a patient with premature ovarian failure: evidence for conserved function in oogenesis and implications for human sterility.

Premature ovarian failure (POF) is a defect of ovarian development and is characterized by primary or secondary amenorrhea, with elevated levels of serum gonadotropins, or by early menopause. The disorder has been attributed to various causes, including rearrangements of a large "critical region" in the long arm of the X chromosome. Here we report identification, in a family with POF, of a gene that is disrupted by a breakpoint. The gene is the human homologue of the Drosophila melanogaster diaphanous gene; mutated alleles of this gene affect spermatogenesis or oogenesis and lead to sterility. The protein (DIA) encoded by the human gene (DIA) is the first human member of the growing FH1/FH2 protein family. Members of this protein family affect cytokinesis and other actin-mediated morphogenetic processes that are required in early steps of development. We propose that the human DIA gene is one of the genes responsible for POF and that it affects the cell divisions that lead to ovarian follicle formation.     

Involvement of insulin and growth hormone (GH) during follicular development in the bovine ovary

Insulin and growth hormone (GH) play critical roles in the process of follicular development and maturation. However, the involvement of insulin receptor (IR) and GH receptor (GHR) during follicular development is not well understood. The aim of this study was to investigate the expression of IR and GHR mRNAs in the granulosa cells (GCs) and theca tissues (TCs) of the follicle at different developmental stages (preovulatory dominant follicles, POFs; estrogen-active dominant follicles, EADs; estrogen-inactive dominant follicles, EIDs; and small follicles, SFs), and second, to examine the effects of follicle-stimulating hormone (FSH) and estradiol (E2) on the expression of IR and GHR genes in cultured bovine GCs. Although the concentration of insulin in follicular fluid (FF) was constant at all developmental stages, the GH concentration in FF was significantly increased in the EAD and POF compared with the EID. IR mRNA in GCs and TCs was significantly increased in the POF compared with other follicles. Regarding GHR expression, significant increases of mRNA expression were observed in GCs of EAD compared to those of SF, EID and POF. GHR mRNA in TCs was significantly decreased in the SF compared with other follicles. In cultured GCs, FSH, but not E2, stimulated the expression of IR and GHR genes. Our results suggest that the increase in the expression of GHR may be a turning point for follicles to enter the ovulatory phase during final follicular development and that the insulin system may support the maturation of preovulatory follicles.     

Idiopathic premature ovarian failure in 63 young women

BACKGROUND: Premature ovarian failure (POF) in adolescents is defined as primary or secondary amenorrhea associated with high follicle-stimulating hormone (FSH) levels. In normal 46,XX patients, its etiology is most often unknown. We have evaluated the clinical, hormonal and ovarian phenotypes in patients with a normal karyotype who were diagnosed with POF before the age of 18. METHODS: Sixty-three patients were included in this retrospective study. RESULTS: The mean patient age was 20.4 years. The patients presented with three clinical patterns: lack of pubertal development (n = 23), primary amenorrhea with interrupted puberty (n = 18), and secondary amenorrhea with normal puberty (n = 22). Ten patients had a familial history of POF and 6 presented with hypothyroidism. The FSH, estradiol and inhibin B levels were not statistically different in the three clinical groups. Fifty percent of the patients presented small ovaries (length <2 cm) at ultrasonography. The presence of follicles was found at histology in only 7 of the 27 patients who underwent an ovarian biopsy. CONCLUSION: 46,XX patients presenting with early POF rarely presented a specific, identifiable disorder. We discuss the clinical management and different diagnosis strategies to improve our current knowledge of this syndrome.     

Molecular evidence that growth of dominant follicles involves a reduction in follicle-stimulating hormone dependence and an increase in luteinizing hormone dependence in cattle

The bovine dominant follicle (DF) model was used to identify molecular mechanisms potentially involved in initial growth of DF during the low FSH milieu of ovarian follicular waves. Follicular fluid and RNA from granulosa and theca cells were harvested from 10 individual DF obtained between 2 and 5.5 days after emergence of the first follicular wave of the estrous cycle. Follicular fluid was subjected to RIA to determine estradiol (E) and progesterone (P) concentrations and RNA to cDNA microarray analysis and (or) quantitative real-time PCR. Results showed that DF growth was associated with a decrease in intrafollicular E:P ratio and in mRNA for the FSH receptor, estrogen receptor 2 (ER beta), inhibin alpha, activin A receptor type I, and a proliferation (cyclin D2) and two proapoptotic factors (apoptosis regulatory protein Siva, Fas [TNFRSF6]-associated via death domain) in granulosa cells. In contrast, mRNAs for the LH receptor in granulosa cells and for two antiapoptotic factors (TGFB1-induced antiapoptotic factor 1, LAG1 longevity assurance homolog 4 [Saccharomyces cerevisiae]) and one proapoptotic factor (tumor necrosis factor [ligand] superfamily, member 8) were increased in theca cells. We conclude that the bovine DF provides a unique model to identify novel genes potentially involved in survival and apoptosis of follicular cells and, importantly, to determine the FSH-, estradiol-, and LH-target genes regulating its growth and function. Results provide new molecular evidence for the hypothesis that DF experience a reduction in FSH dependence but acquire increased LH dependence as they grow during the low FSH milieu of follicular waves.     

Blood Cell Mitochondrial DNA Content and Premature Ovarian Aging

Primary ovarian insufficiency (POI) is a critical fertility defect characterized by an anticipated and silent impairment of the follicular reserve, but its pathogenesis is largely unexplained. The frequent maternal inheritance of POI together with a remarkable dependence of ovarian folliculogenesis upon mitochondrial biogenesis and bioenergetics suggested the possible involvement of a generalized mitochondrial defect. Here, we verified the existence of a significant correlation between blood and ovarian mitochondrial DNA (mtDNA) content in a group of women undergoing ovarian hyperstimulation (OH), and then aimed to verify whether mtDNA content was significantly altered in the blood cells of POI women. We recruited 101 women with an impaired ovarian reserve: 59 women with premature ovarian failure (POF) and 42 poor responders (PR) to OH. A Taqman copy number assay revealed a significant mtDNA depletion (P<0.001) in both POF and PR women in comparison with 43 women of similar age and intact ovarian reserve, or 53 very old women with a previous physiological menopause. No pathogenic variations in the mitochondrial DNA polymerase γ (POLG) gene were detected in 57 POF or PR women with low blood mtDNA content. In conclusion, blood cell mtDNA depletion is a frequent finding among women with premature ovarian aging, suggesting that a still undetermined but generalized mitochondrial defect may frequently predispose to POI which could then be considered a form of anticipated aging in which the ovarian defect may represent the first manifestation. The determination of mtDNA content in blood may become an useful tool for the POI risk prediction.     

Human placenta‐derived mesenchymal stem cells inhibit apoptosis of granulosa cells induced by IRE1α pathway in autoimmune POF mice

Previous studies have shown that the ovarian failure in autoimmune‐induced premature ovarian failure (POF) mice could be improved by the transplantation of human placenta‐derived mesenchymal stem cells (hPMSCs); however, the protective mechanism of hPMSCs transplantation on ovarian dysfunction remains unclear. Ovarian dysfunction is closely related to the apoptosis of granulosa cells (GCs). To determine the effects of hPMSCs transplantation on GCs apoptosis, an autoimmune POF mice model was established with zona pellucida glycoprotein 3 (ZP3) peptide. It is reported that the inositol‐requiring enzyme 1α (IRE1α) and its downstream molecules play a central role in the endoplasmic reticulum (ER) stress‐induced apoptosis pathway. So the aim of this study is to investigate whether hPMSCs transplantation attenuated GCs apoptosis via inhibiting ER stress IRE1α signaling pathway. The ovarian dysfunction, follicular dysplasia, and GCs apoptosis were observed in the POF mice. And the IRE1α pathway was activated in ovaries of POF mice, as demonstrated by, increased X‐box binding protein 1 (XBP1), up‐regulated 78 kDa glucose‐regulated protein (GRP78) and caspase‐12. Following transplantation of hPMSCs, the ovarian structure and function were significantly improved in POF mice. In addition, the GCs apoptosis was obviously attenuated and IRE1α pathway was significantly inhibited. Transplantation of hPMSCs suppressed GCs apoptosis‐induced by ER stress IRE1α signaling pathway in POF mice, which might contribute to the hPMSCs transplantation‐mediating ovarian function recovery.     

Study of FMR1 gene association with ovarian dysfunction in a sample from the Basque Country

Premature ovarian failure (POF) is defined as cessation of menses before the age of 40. The most significant single gene associated with POF is the Fragile X Mental Retardation 1 gene (FMR1). In the present work we screened women with fertility problems from the Basque Country in order to determine, whether in these women, FMR1 CGG repeat size in the intermediate and premutation range was associated with their pathology, and whether intermediate and premutation carriers had endocrine signs of diminished ovarian function, using the most established measure of ovarian reserve, the gonadotropin FSH. A patient sample of 41 women with ovarian insufficiency and a control sample of 32 women with no fertility problems from the Basque Country were examined. The patient sample was classified into three categories according to the results of the retrospective assessment of their ovarian function. In group 2 of patients, women with irregular cycles, reduced fecundity and FSH levels ≥ 10 IU/l, there is a significant increase in the number of intermediate and premutation FMR1 alleles (35–54 CGG repeats). In group 3 of patients, women with amenorrhea for at least four consecutive months and FSH levels ≥ 10 IU/l, a significant increase in the number of intermediate FMR1 alleles (35–54 CGG repeats) was found in patients compared with controls. In this group all the patients had a serum concentration > 40 IU/l. The results suggest that in the analysed Basque sample the FMR1 gene has a role in the aetiology of POF. However, elevated FSH levels are more related to the menstrual cycle pattern than to the CGG repeat size.     

Role and gonadotrophic regulation of X-linked inhibitor of apoptosis protein expression during rat ovarian follicular development in vitro

Although FSH up-regulates follicular cell X-linked inhibitor of apoptosis protein (XIAP) expression and suppresses apoptosis in vivo, if these events are coincidental or causally related remains to be investigated. The present study examined the role and gonadotrophic regulation of XIAP expression during follicular development in vitro. Follicles (160-210 microm) cultured for 0-6 days with FSH (100 ng/ml) showed significant growth, as evidenced by increases in follicular size, cell number, and DNA contents. Follicular XIAP content was low in the absence of FSH but was increased by the addition of gonadotropin. Apoptosis was evident in follicles cultured without FSH but was suppressed in the presence of gonadotropin. At low FSH concentration (5 ng/ml), adenoviral XIAP sense cDNA expression increased XIAP and DNA contents, reduced apoptosis, and enhanced follicular growth. Infection of the FSH-stimulated follicles with XIAP antisense elicited opposite responses. In primary granulosa cell cultures, FSH significantly increased XIAP content, inhibited apoptosis, and decreased cell number, a response potentiated by XIAP sense expression. In conclusion, the present studies demonstrated, to our knowledge for the first time, that XIAP plays an important role in the regulation of ovarian follicular development. In addition, a follicle culture system coupled to an adenoviral gene-manipulation procedure has been established and may prove to be a useful approach in assessing the role of specific genes in follicular development and atresia.     

Autoimmune targeted disruption of the pituitary-ovarian axis causes premature ovarian failure

Premature ovarian failure (POF) is characterized by amenorrhea and high serum levels of follicle-stimulating hormone (FSH). POF causes female infertility and represents a substantial women's health risk affecting 1% of women by age 40. Although ovarian autoimmunity has been associated with POF, the identity of ovarian Ags recognized is unknown. In this study, we show that autoimmune-targeted disruption of the pituitary-ovarian axis leads to POF. Immunization of SWXJ female mice with the p215-234 peptide derived from mouse inhibin-alpha activates CD4(+) T cells and induces experimental autoimmune oophoritis with a unique biphasic phenotype characterized by an early stage of enhanced fertility followed by a delayed stage of POF. Affected mice show high serum levels of inhibin-alpha-neutralizing Abs that prevent inhibin-mediated down-regulation of activin-induced pituitary FSH release. The loss of activin/FSH down-regulation leads to prolonged metestrus-diestrus, superovulation, increased numbers of mature follicles, increased offspring, accelerated depletion of primordial follicles, and ultimately premature infertility. Thus, inhibin-alpha-targeted experimental autoimmune oophoritis is initiated by CD4(+) Th1 T cells that stimulate B cells to produce inhibin-alpha-neutralizing Abs directly capable of mediating POF and transferring disease into naive recipients. Our inhibin-alpha autoimmune model of POF shows how premature infertility may develop in the context of elevated FSH levels thereby closely mimicking the hallmark features of human POF.     

Characterization of goose SPMS: Molecular characterization and expression profiling of SPMS in the goose ovary

Spermine synthase (SPMS), which converts spermidine into spermine, is essential for normal cell growth and development processes in humans and other mammals, but the molecular characterization and expression profiling of the SPMS gene remain undetermined in goose tissues and ovarian follicles. In this study, the SPMS cDNA sequence of the Sichuan white goose was cloned and analysed, and SPMS mRNA expression was profiled in various tissues and ovarian follicles. The results showed that the open reading frame of the SPMS cDNA sequence was 1092?bp in length, encoding 363 amino acids with a molecular weight of 41?kDa. Among all the examined tissues, SPMS expression was highest in the spleen and cerebrum and lowest in the breast and thigh muscles. SPMS expression in the F1 follicle was significantly higher than that in the POF (except for POF2) (P?<?0.05). Our results indicate that SPMS might play an important role in follicular development and ovulation.