FSH Secretion Research Articles

Idiopathic central precocious puberty in a Klinefelter patient: highlights on gonadotropin levels and pathophysiology

BackgroundIdiopathic central precocious puberty (ICPP) is supposed to be non-existent in a context of testicular destruction that is typically present in Klinefelter syndrome (KS). Herein, we describe a rare case of ICPP in a Klinefelter patient (47,XXY) with 2 maternal X chromosomes. Moreover, we highlight the differences in gonadotropin levels in comparison to males with ICPP and a normal karyotype.Case presentationAn 8 years old boy with a history of cryptorchidism was evaluated for precocious puberty (Tanner staging: P2/G3). Both testes measured 25x35mm. His hormonal profile confirmed a central origin of precocious puberty with high serum testosterone (4.3 ng/ml), luteinizing hormone [LH (3.5 UI/l)] and follicle stimulating hormone [FSH (7.7 UI/l)] levels. Luteinizing hormone-releasing hormone (LHRH) test amplified LH and FSH secretion to 24 and 14 UI/l respectively. Brain magnetic resonance imaging (MRI) was normal. No MKRN3 mutation was detected. He was treated for ICPP for two years. During puberty, he suffered from hypergonadotropic hypogonadism leading to the diagnosis of KS (47,XXY karyotype). Chromosomal analysis by fluorescent multiplex polymerase chain reaction (PCR) using X chromosome microsatellite markers identified 2 maternal X chromosomes.Analysing 8 cases of KS developing ICPP (our reported case and 7 other published cases) revealed that these KS patients with ICPP have higher LH and FSH levels during ICPP episode than in ICPP patients with a normal karyotype (ICPP with KS vs ICPP with a normal karyotype: LH levels 9.4 ± 12 vs 1.1 ± 0.6 UI/l; FSH levels 23.1 ± 38.5 vs 2.7 ± 1.5 UI/l). Furthermore, their response to gonadotropin-releasing hormone (GnRH) stimulation is characterized by excessive LH and FSH secretion (LH levels post-GnRH: 58 ± 48 vs 15.5 ± 0.8 UI/l; FSH levels post-GnRH: 49.1 ± 62.1 vs 5.7 ± 3.9 UI/l).ConclusionsICPP in boys is extremely rare. The pathophysiology of ICPP in KS is unknown. However, maternal X supplementary chromosome and early testicular destruction may play a significant role in the initiation of ICPP, in part explaining the relative “overrepresentation of ICPP in KS. Thus, karyotype analysis could be considered for boys suffering from ICPP, especially if testicular size is smaller or gonadotropins are significantly elevated.

Using exosomal miRNAs extracted from porcine follicular fluid to investigate their role in oocyte development

sBackgroundFollicular development is crucial to normal oocyte maturation, with follicular size closely related to oocyte maturation. To better understand the molecular mechanisms behind porcine oocyte maturation, we obtained exosomal miRNA from porcine follicular fluid (PFF). These miRNA samples were then sequenced and analyzed regarding their different follicular sizes, as described in the methods section.ResultsFirst, these results showed that this process successfully isolated PFF exosomes. Nearly all valid reads from the PFF exosomal sequencing data were successfully mapped to the porcine genome database. Second, we used hierarchical clustering methods to determine that significantly expressed miRNAs were clustered into A, B, C, and D groups in our heatmap according to different follicle sizes. These results allowed for the targeting of potential mRNAs genes related to porcine oocyte development. Third, we chose ten, significantly expressed miRNAs and predicted their target genes for further GO analysis. These results showed that the expression levels of neurotransmitter secretion genes were greatly changed, as were many target genes involved in the regulation of FSH secretion. Notably, these are genes that are very closely related to oocyte maturation in growing follicles. We then used pathway analysis for these targeted genes based on the originally selected ten miRNAs. Results indicated that the pathways were mainly related to the biosynthesis of TGF-beta and its signaling pathway, which are very closely related to reproductive system functions.ConclusionsFinally, these exosomal miRNAs obtained from PFF may provide a valuable addition to our understanding of the mechanism of porcine oocyte maturation. It is also likely that these exosomal miRNAs could function as molecular biomarkers to choose high-quality oocytes and allow for in vitro porcine embryo production.

CircAkap17b acts as a miR-7 family molecular sponge to regulate FSH secretion in rat pituitary cells.

The pituitary gland functions as a prominent regulator of diverse physiologic processes by secreting multiple hormones. Circular RNAs (circRNAs) are an emerging novel type of endogenous noncoding RNA that have recently been recognized as powerful regulators participating in various biological processes. However, the physiological roles and molecular mechanisms of circRNAs in pituitary remain largely unclear. Herein, we concentrated on expounding the biological function and molecular mechanism of circRNA in rat pituitary. In this study, we identified a novel circRNA in pituitary tissue, circAkap17b, which was pituitary- and stage-specific. Then, we designed circAkap17b siRNA and constructed an overexpression plasmid to evaluate the effect of loss- and gain-of-circAkap17b function on FSH secretion. Interestingly, silencing circAkakp17b significantly inhibited FSH expression and secretion, while overexpression of circAkap17b enhanced FSH expression and secretion. Furthermore, dual luciferase reporter and RNA immunoprecipitation (RIP) assays confirmed that circAkap17b could serve as miR-7 sponge to regulate target genes. Additionally, miR-7b suppressed FSH expression and secretion by directly targeting Fshb through the dual luciferase reporter and RT-qPCR analysis. Additionally, rescue experiments showed that circAkap17b could regulate FSH secretion in pituitary cells through a circAkap17b-miR-7-Fshb axis. Collectively, we demonstrated that circAkap17b could act as a molecular sponge of miR-7 to upregulate expression of the target gene Fshb and facilitate FSH secretion. These findings provide evidence for a novel regulatory role of circRNAs in pituitary.

Molecular and hormonal changes caused by long-term use of high dose pregabalin on testicular tissue: the role of p38 MAPK, oxidative stress and apoptosis.

In 1990, pregabalin was introduced as a novel antiepileptic drug that acts by binding selectively to the alpha-2-delta subunits of voltage-gated calcium channels resulting in increasing neuronal GABA levels and inhibiting the release of exciting neurotransmitters. The aim of our study is to assess the hazardous effects of prolonged high-dose pregabalin (like that abused by addicts) on testes and to clarify the potential causative mechanisms. The current study was conducted on 70 adult male Wistar albino rats which were divided into 7 groups. In our study we evaluated the effect of pregabalin, at concentrations 150 and 300mg/kg/day for 90 days, on hormones; FSH, LH, testosterone and prolactin secretion. Our study also evaluated the expression of apoptosis-related genes BAX and BCL2 in testicular tissue in addition to the western blotted analysis of p38 Mitogen activated protein kinases (p38 MAPK). The levels of reduced glutathione, malondialdehyde and superoxide dismutase were also measured. Pregabalin decreased testosterone level while FSH, LH and prolactin showed a significant increase. It also produced genotoxicity through reversal of the BAX/BCL2 ratio; increased p38 MAPK level and induction of oxidative stress markers. The concomitant administration of vitamin E significantly reduced all the previously mentioned biochemical and hormonal adverse effects caused by pregabalin. Pregabalin can adversely affect male fertility particularly in addicts and patients who are being treated with it for long periods as those suffering from neuropathies and seizures. Antioxidants like vitamin E could have a role in amelioration.

Endocrineand follicular studies in Meishan pig

The enhanced early embryonic survival in Chinese Meishan compared with Large- White gilts may be due, in part, to differences in ovarian and endocrine function, particularly during the periovulatory period. The overall patterns of oestradiol, LH and FSH secretion were not different between Meishan and Large-White hybrid controls during this period, although circulating inhibin concentrations were higher in Meishan gilts. Thus, there appeared to be a decreased sensitivity to inhibin feedback on FSH secretion in Meishan gilts. Behavioural oestrus was exhibited earlier relative to the LH surge in Meishan gilts than in Large-White hybrid gilts, but the time interval from the oestradiol peak until the LH surge was similar in both breeds. This finding suggests that Meishan gilts are more sensitive in terms of initiating a behavioural response, but not in terms of positive feedback. Although preovulatory follicular characteristics were as variable in Meishan as in Large-White hybrid gilts, follicles from Meishan gilts were smaller, but contained a higher concentration of oestradiol in the follicular fluid. This was probably due to increased aromatase activity in both granulosa and theca cells of Meishan follicles. The enhanced maturation of the intrafollicular environment in Meishan gilts was reflected in the oocyte population which was at a more advanced stage of development in the period preceding ovulation. In addition to decreasing the time between onset of oestrus and ovulation, advancing the LH surge to coincide with onset of oestrus (via hCG administration) decreased embryo survival at day 30 of gestation. It is concluded that both endocrine and follicular mechanisms have a role in ensuring the prolificacy in the Meishan breed.

Apoptosis during folliculogenesis in pigs

The number of female germ cells in pig fetuses decreases by 70% between day 50 after mating and day 300 after birth. Approximately 55% of antral follicles undergo degeneration (atresia) except during the 3 days before oestrus, when only 15% of the follicles survive to ovulate. Apoptosis, a form of programmed cell death, is recognized as the mechanism of germ cell death and follicle atresia at all stages of folliculogenesis. The internucleosomal cleavage of genomic DNA caused by caspase-induced deoxyribonuclease activity was measured in pig granulosa cells by DNA fluorescence flow cytometry, densitometry of fluorescently labelled internucleosomal DNA fragments and immunohistochemical analysis of the 3' end labelling of deoxyribonuclease- nicked DNA on frozen tissue sections. Follicular atresia during the 3 days before oestrus is associated with a 60-70% decrease in the secretion of FSH. In granulosa cells, apoptosis is associated with decreased cell proliferation and reduced production of oestradiol and inhibin. In cultured pig granulosa cells, FSH and IGF-I are anti-apoptotic and a caspase inhibitor blocked apoptosis, thereby providing evidence of caspase activity. Oocytes in most follicles have resumed meiotic maturation; therefore, one role for apoptosis and follicle atresia may be to act as a barrier to ovulation of oocytes that have not remained in meiotic arrest.

Role of neuropeptides and amino acids in controlling secretion of hormones from the anterior pituitary gland in pigs

All reproductive processes involve one or more of the protein hormones secreted from the anterior pituitary gland: LH, FSH, prolactin, growth hormone, ACTH and thyroidstimulating hormone (TSH). Primary hormones of reproduction, such as LH and FSH, directly regulate a reproductive activity. For example, LH and FSH stimulate follicular growth and the associated secretion of oestradiol in sows. In contrast, secondary hormones of reproduction such as TSH are permissive and regulate other physiological systems that indirectly, but profoundly, influence reproduction. Reproduction in pigs can be enhanced by developing strategies to alter and control secretion of hormones from the anterior pituitary gland. However, the successful manipulation of adenohypophysial hormone secretion will require a sound understanding of the mechanisms controlling the function of the hypothalamic—pituitary axis. Hypothalamic hormones including GrtRH, dopamine, growth hormone-releasing hormone (GHRH), somatostatin, corticotrophin-releasing hormone (CRI4) and thyrotrophin-releasing hormone (TRI-I) are synthesized in perikarya that possess axons that terminate at the median eminence. These hormones are released into the hypothalamo—hypophysial portal vasculature, travel to the anterior pituitary gland and stimulate or inhibit secretion of adenohypophysial hormones. Secretion of hypothalamic hormones is ultimately controlled by a variety of neurotransmitters and neuropeptides, the most studied in swine being the endogenous opioid peptides (EOP) and more recently, the excitatory amino acids (ExAA). In general, EOP inhibit GrtRH and hence LH secretion, and this effect involves the central catecholaminergic system. A definitive role for EOP in the modulation of FSH release remains to be determined. EOP stimulate secretion of GHRH and thus growth hormone release, and depending on the animal model studied, EOP exert either stimulatory or inhibitory influences on prolactin secretion. ExAA, working via N-methyl-o-aspartate (NINIDA) receptors at the central nervous system, stimulate secretion of LH, FSH, growth hormone and prolactin in appropriate animal models. However, in certain situations, an inhibitory effect of ExAA on LH secretion has been demonstrated. The modulation of growth hormone and prolactin secretion by ExAA involves LOP. Research investigating the function of ExAA and EOP in the physiological control of swine reproduction warrants further scrutiny.

Expression profile of kisspeptin2 and gonadotropin-releasing hormone2 mRNA during photo-thermal and melatonin treatments in the female air-breathing catfish Heteropneustes fossilis.

In seasonally breeding vertebrates, extrinsic factors like photoperiod and temperature are major determinants, controlling the annual reproductive cycle. In teleosts, kisspeptin, which occurs in two molecular forms: kisspeptin1 (Kiss1) and kisspetin2 (Kiss2), has been reported to alter gonadotropin (Lh and Fsh) secretion but its effect on gonadotropin-releasing hormone (Gnrh) secretion is not unequivocally proved. In the catfish Heteropneustes fossilis, we isolated and characterized kiss2 and gnrh2 cDNAs and the present work reports effects of altered photo-thermal conditions and melatonin (MT, a pineal hormone) on their expressions in the brain. The exposure of the catfish to long photoperiod (LP, 16h light) at normal temperature (NT) or high temperature (HT, 28°C) at normal photoperiod (NP) for 14 or 28days stimulated both kiss2 and gnrh2 expression in both gonad resting and preparatory phases with the combination of LP + HT eliciting maximal effects. Short photoperiod (SP, 8h light) under NT or HT altered the gene expression according to the reproductive phase and temperature. MT that mediates photo-thermal signals to the brain inhibited brain kiss2 and gnrh2 gene expression in the NP + HT, LP + NT, and SP + NT groups. The altered photo-thermal conditions elicited changes in steroidogenic pathway as evident from changes in plasma E2, progesterone, and testosterone levels. The results show that brain kiss2-gnrh2 signaling is involved in photo-thermal-mediated mechanisms controlling reproduction.

How the therapy for endometriosis affects the hormonal status in female infertility

Aim. To determine changes in the values of pituitary and ovarian hormones in women with endometriosis-associated infertility, as well as to evaluate the impact of endometriosis therapy on hormonal parametersMaterials and Methods. Sixty-nine women with small-sized endometriomas with adenomyosis (target group) and 50 women with tubal infertility (controls) were examined. The values of pituitary (luteinizing and follicle-stimulating hormones, prolactin) and ovarian (estradiol, tubular antimullerian hormone) hormones were studied. The comparison of quantitative indicators between the studied groups of women was carried out using the Mann-Whitney U-test, the statistical significance of the results was assessed at a level of not less than 95% (risk of error not more than 5% - p <0.05).ResultsInfertile women with endometrioma and adenomyosis have lower AMH levels than infertile women with tubal factor. Among patients with endometriosis, there is an increase in serum FSH, LH, prolactin and estradiol compared with women without endometriosis.Dienogest moderately inhibits the secretion of FSH and LH; it has almost no effect on blood AMH and estradiol values. The use of dienogest can moderately increase the level of prolactin, It is noteworthy that these indicators are within the reference range. The use of diferelin for the treatment of endometriosis has a more active effect on the hormonal status of patients, given the decrease in such indicators as FSH and LH, AMH, estradiol and prolactinConclusions. Women with infertility against the background of endometriosis have minor changes in the hypothalamic-pituitary-ovarian system, which indicates the need to develop medical measures in order to correct them.The use of dienogest compared to diferelin affects the hormonal status of patients with endometriosis to a lesser extent, given the pronounced anti-proliferative effect.

Transcriptomic analysis of the porcine anterior pituitary gland during the peri‐implantation period

The peri-implantation period is controlled by signals originating from hypothalamic-pituitary-ovarian axis, uterus and developing embryos. The transcriptomic activity of the anterior pituitary gland may be important for the control of the peri-implantation period. The aim of this study was to determine the alternations in the transcriptomic profile of porcine anterior pituitary gland during the peri-implantation period (days 15-16 of pregnancy) in comparison with established for the respective days of the oestrous cycle. Analysis using a microarray approach indicated that the 651 genes (fold-change ˂1.2; p≤.05) were differentially expressed (DEGs) in the anterior pituitary of pigs during the peri-implantation period when compared to cyclic females. Of these DEGs, 404 were upregulated and 247 downregulated. Analysis of occurred relationships among DEGs revealed that some of them are involved in steroid-response and oestrogen synthesis, FSH secretion, immune response, PPAR signalling pathway and the potential for DNA methylation. In conclusion, the altered transcriptomic profile of the porcine pituitary gland in pigs during the peri-implantation period indicates the role of embryos presence in the creation of transcriptomic activity of the pituitary gland in pigs.

Augmentation of Nr4a3 and Suppression of Fshb Expression in the Pituitary Gland of Female Annexin A5 Null Mouse.

GnRH enhances the expression of annexin A5 (ANXA5) in pituitary gonadotropes, and ANXA5 enhances gonadotropin secretion. However, the impact of ANXA5 regulation on the expression of pituitary hormone genes remains unclear. Here, using quantitative PCR, we demonstrated that ANXA5 deficiency in female mice reduced the expression of Fshb and Gh in their pituitary glands. Transcriptome analysis confirmed a specific increase in Nr4a3 mRNA expression in addition to lower levels of Fshb expression in ANXA5-deficient female pituitary glands. This gene was then found to be a GnRH-inducible immediate early gene, and its increased expression caused protein to accumulate in the nucleus after administration of a GnRH agonist in LβT2 cells, which are an in vitro pituitary gonadotrope model. The increase in ANXA5 protein levels in LβT2 cells clearly suppressed Nr4a3 expression. siRNA-mediated inhibition of Nr4a3 expression increased Fshb expression. The results revealed that GnRH stimulates Nr4a3 and Anxa5 sequentially. NR4A3 suppression of Fshb may be necessary for later massive secretion of FSH by GnRH in gonadotropes, and Nr4a3 would be negatively regulated by ANXA5 to increase FSH secretion.

Kisspeptin and neurokinin B interactions in modulating gonadotropin secretion in women with polycystic ovary syndrome.

STUDY QUESTIONWhat is the role of the hypothalamic neuropeptide neurokinin B (NKB) and its interaction with kisspeptin on GnRH/LH secretion in women with polycystic ovary syndrome (PCOS)?SUMMARY ANSWERAdministration of neurokinin 3 receptor antagonist (NK3Ra) for 7 days reduced LH and FSH secretion and LH pulse frequency in women with PCOS, whilst the stimulatory LH response to kisspeptin-10 was maintained.WHAT IS KNOWN ALREADYPCOS is characterized by abnormal GnRH/LH secretion. NKB and kisspeptin are master regulators of GnRH/LH secretion, but their role in PCOS is unclear.STUDY DESIGN, SIZE, DURATIONThe NK3Ra MLE4901, 40 mg orally twice a day, was administered to women with PCOS for 7 days (n = 8) (vs no treatment, n = 7). On the last day of NK3Ra administration or the equivalent day in those not treated, women were randomized to 7-h kisspeptin-10 (4 µg/kg/h i.v.) or vehicle infusion. This was repeated with the alternate infusion in a subsequent cycle.PARTICIPANTS/MATERIALS, SETTING, METHODSSubjects were women with PCOS, studied in a Clinical Research Facility. Reproductive hormones were measured before and after NK3Ra administration. On the last day of NK3Ra administration (or the equivalent cycle day in untreated women), all women attended for an 8-h frequent blood sampling to allow analysis of the pulsatile LH secretion.MAIN RESULTS AND THE ROLE OF CHANCENK3Ra reduced LH secretion (4.0 ± 0.4 vs 6.5 ± 0.8 IU/l, P < 0.05) and pulse frequency (0.5 ± 0.1 vs 0.8 ± 0.1 pulses/h, P < 0.05); FSH secretion was also reduced (2.0 ± 0.3 vs 2.5 ± 0.4 IU/l, P < 0.05). Without NK3Ra pre-treatment, kisspeptin-10 increased LH secretion (5.2 ± 0.5 to 7.8 ± 1.0 IU/L, P < 0.05), with a positive relationship to oestradiol concentrations (r2 = 0.59, P < 0.05). After NK3Ra administration, the LH response to kisspeptin-10 was preserved (vehicle 3.5 ± 0.3 vs 9.0 ± 2.2 IU/l with kisspeptin-10, P < 0.05), but the positive correlation with oestradiol concentrations was abolished (r2 = 0.07, ns. after NK3Ra). FSH secretion was increased by kisspeptin-10 after NK3Ra treatment, but not without NK3Ra treatment.LIMITATIONS, REASONS FOR CAUTIONThe study did not explore the dose relationship of the effect of NK3R antagonism. The impact of obesity or other aspects of the variability of the PCOS phenotype was not studied due to the small number of subjects.WIDER IMPLICATIONS OF THE FINDINGSThese data demonstrate the interactive regulation of GnRH/LH secretion by NKB and kisspeptin in PCOS, and that the NKB system mediates aspects of oestrogenic feedback.STUDY FUNDING/COMPETING INTEREST(S)Wellcome Trust through Scottish Translational Medicine and Therapeutics Initiative (102419/Z/13/A) and MRC grants (G0701682 to R.P.M. and R.A.A.) and MR/N022556/1 to the MRC Centre for Reproductive Health. This work was performed within the Edinburgh Clinical Research Facility. J.T.G. has undertaken consultancy work for AstraZeneca and Takeda Pharmaceuticals and is an employee of Boehringer Ingelheim. R.P.M. has consulted for Ogeda and was CEO of Peptocrine. R.A.A. has undertaken consultancy work for Merck, Ferring, NeRRe Therapeutics and Sojournix Inc. J.D.V. and K.S. have nothing to disclose.TRIAL REGISTRATION NUMBERN/A.

MON-009 GLUT1-Mediated Glycolysis Facilitates GnRH-Induced Secretion of Luteinizing Hormone

Reproduction requires intensive energy expenditure, and energy availability impacts the function of the reproductive endocrine HPG-axis. Accordingly, the reproductive axis is suppressed during hypoglycemia. Circulating blood glucose can directly interact with gonadotropes within the highly vascular pituitary. Therefore, it is possible that gonadotropes may sense energy availability via the presence of glucose in the circulation and integrate this status with input from GnRH neurons to regulate hormone production. Gonadotropes dominantly express glucose transporter 1 (GLUT1) and increase glucose uptake in response to GnRH. Thus, we hypothesized that gonadotropes engage glycolysis in response to GnRH stimulation due to the high energy demand of protein synthesis required for LH production. We developed an approach to sort and successfully culture primary gonadotropes from wild type mice. Using this approach, we performed extracellular flux analysis and found that gonadotropes respond to GnRH by inducing GLUT1-mediated glycolysis that is independent of mitochondrial respiration. Knock-down of GLUT1 expression in the LβT2 gonadotrope cell line, glucose restriction, or treatment with the competitive inhibitor of glycolysis, 2-DG, diminished GnRH-induced LH secretion, indicating GLUT1 expression is necessary for maximal GnRH-induced LH secretion. We confirmed this observation in primary female mouse gonadotropes by limiting glucose availability which resulted in diminished basal LH and FSH secretion. Lastly, GLUT1 expression in the pituitary correlates with GnRH receptor expression and is increased during the LH surge in a mouse model. These results implicate glucose uptake through GLUT1 as permissive for gonadotrope secretion of LH and therefore reproductive function, especially the LH surge. We conclude that GLUT1-mediated glucose uptake is an important rate-limiting step in gonadotropin synthesis and operation of the HPG-axis.

SUN-032 Digenic Inheritance of PCSK1 and CHD7 Mutations in PAX4 Homozygous Diabetic Male with Normosmic Hypogonadotropic Hypogonadism

Background: Normosmic congenital hypogonadotropic hypogonadism denotes Kallmann syndrome not associated with anosmia or hyposmia. Over the past few years, the availability of next-generation sequencing has started to unravel the complex molecular basis of congenital hypogonadotrophic hypogonadism including digenic or oligogenic pathogenecity in addition to classic monogenic causality (1). Clinical Case: A 22-year-old male patient was referred to the endocrine clinic in 2018 with recent -onset hyperglycemia. His weight was 82.2 kg with a height of 180 cm (BMI of 25.3 kg/m2). Physical examination revealed small testes, micropenis, and no axillary and pubic terminal hair. His smell sense was intact. His hormonal test reveals low testosterone (0.10 ng/mL) and low free testosterone (0.65 pg/mL) levels with inappropriately low gonadotrophins levels. Secretion of LH and FSH increased 2-fold after GnRH stimulation. His bone age was 13-years 6-months old, and brain magnetic resonance imaging showed the presence of olfactory bulbs, and unremarkable findings except for small size of the pituitary gland. There were no signs associated with CHARGE syndrome (coloboma ocular, heart defects, atresia or stenosis of the choanae, retardation of growth and/or development, genitourinary anomalies, and ear abnormalities). Biochemical investigation demonstrated high serum glucose level and high HbA1c (13.8%). To identify variants to cause the phenotype of the proband, we adopted trio-based whole exome sequencing (WES) and candidate gene approach. Candidate genes was listed from the orphanet (https://www.orpha.net). WES of the proband revealed the presence of heterozygote missense mutations of the CHD7 gene (c.6107C>T, p.Pro2036Leu, rs369543203) and PCSK1 gene (c.239G>A, p.Arg80Gln, rs1799904). The missense variants were predicted to have a damaging effect on the encoded protein, by SIFT and PolyPhen-2 analyses. Genetic analyses of his family revealed that his father had the same heterozygote missense mutations of the CHD7 gene, but wild type of PCSK1. Proband’s mother had the same heterozygote missense mutations of PCSK1, but wild type of CHD7. Furthermore, the proband had homozygote missense mutation of PAX4 (c.575G>A, p.Arg192His, rs2233580) known as maturity-onset diabetes of the young (MODY) 9 gene. Both parents have the same but heterozygous mutation of PAX4 p.Arg192His, and pre-diabetic range of hyperglycemia. Conclusion: This is the first case demonstrating digenic inheritance of mutations in PCSK1 and CHD7 as a potential cause of normosmic hypogonadotrophic hypogonadism, interestingly in PAX4 homozygous diabetic male. Reference: (1) Maione L, et al. Genetic counseling for congenital hypogonadotropic hypogonadism and Kallmann syndrome: new challenges in the era of oligogenism and next-generation sequencing. Eur J Endocrinol. 2018;178(3):R55-R80.

Ovarian 25OH-vitamin D production in young women affected by polycystic ovary syndrome.

Vitamin D is involved in the regulatory mechanisms of ovarian function and is frequently low in PCOS patients. Since obesity and hyperinsulinemic state negatively influenced vitamin D levels, therefore, we evaluated the production of vitamin D at the ovarian level only in lean and normoinsulinemic PCOS subjects. Basal, GnRH analogue-induced ovarian production of 25OH-vitamin D (VitD) and a direct sampling at ovarian vein level were investigated. Basal and GnRH analogue-induced hormone levels were evaluated at peripheral level in 45 subjects, aged 18-39years, and in 22 healthy women with age- and BMI-matched as controls. In 12 PCOS patients, undergoing laparoscopy, a venous sampling at both peripheral and ovarian level was further done. All subjects presented low VitD levels, appropriate to the season and with no difference between PCOS and control subjects. GnRH analogue significantly stimulated plasma LH, FSH, 17-OHP and estradiol secretion (p from < 0.05 to < 0.001 vs basal levels), whereas no effect was observed on both serum AMH and VitD concentrations in all groups. A significant difference (p < 0.006), between peripheral and ovarian veins, was observed in both AMH and estradiol levels in PCOS subjects, while no gradient of VitD was detected. All patients presented with low VitD levels. The absence of any VitD variation, both at basal and after GnRH analogue administration, or at peripheral-ovarian vein gradient, suggests no pituitary-ovarian axis involvement in VitD production or its direct ovarian production in lean and normoinsulinemic PCOS subjects.

REGOLAZIONE DELLA FERTILITÀ: ARTE E NEUROSCIENZE

The term ‘fertility’ (from Latin language getting abundant harvests) leads back to several fields of the sciences; however, when it is linked to the physiology of reproduction it became, along with nutrition, as a fundamental behavior for the living species. The ‘mystery’ of human reproduction has been an attractive issue and extensively represented in figurative arts (paintings, sculptures, ect.) throughout the centuries and different cultures, being itself an ‘art’. Many scientists have been so fascinated by the art to have modified their personal and scientific lives. Art and sciences, in particular the neurosciences, share their origin during human history, being both the expression of human abilities to formulate an abstract thought to understand and describe the complexity of the world around us. For instance, paintings as the Cabanel’s Birth of Venus as well as the Primavera by Botticelli summarize several aspects of the sexual desire and reproduction; however, the neurosciences provide a more complex and artistic picture of the control of fertility. From the first descriptions, by Geoffrey Harris and Luciano Martini, of the control of the hormonal cascade governing reproductive functions by the hypothalamus, an important region of our brain, it came to the identification of the GnRH, the master regulator of hormonal reproductive axis. GnRH is produced by specific hypothalamic neurons and it is released in the blood vessels directed toward the pituitary gland in a pulsatile manner, finely tuned for rhythm and intensity, to regulate the secretion of pituitary gonadotropins LH and FSH. Released gonadotropin will exert trophic and regulatory effects on gonadal functions and on the production of sex hormones (testosterone and estrogens). A number of neuroendocrine signals control the development and the activity of GnRH neurons and whose functional alterations are responsible of several reproductive system diseases. According to the data so far available, it is fair to say that the development of such complex mechanisms, under the brain control, endowed with the extraordinary ability to create new individuals could be a real artwork.

Active immunization against AMH reveals its inhibitory role in the development of pre-ovulatory follicles in Zhedong White geese

The aim of this study was to investigate the effects of active immunization against recombinant Anti-Müllerian hormone (AMH) protein on the ovarian follicular development, egg production, and molecular regulatory mechanisms in broody-prone Zhedong White geese. For this, a recombinant goose AMH protein was expressed using a prokaryotic expression system. Fifty incubating geese from the same genetic background were selected and equally divided into two groups. The immunization group was actively immunized against the recombinant goose AMH protein, whereas the control group was immunized against bovine serum albumin (BSA). Immunization against AMH accelerated ovarian follicular development and increased clutch sizes by one to two eggs in two consecutive laying-incubation cycles. Furthermore, immunization against AMH upregulated the mRNA transcription levels of the FSH-beta gene in the pituitary gland, and FSHR, 3beta-HSD, and Smad4 genes in the granulosa layer of pre-ovulatory follicles; however, immunization downregulated the expression of the OCLN gene in the granulosa layer of pre-ovulatory follicles, and Smad5 and Smad9 genes in the granulosa layer of SYFs. These results suggest that AMH might hinder ovarian follicular development by decreasing both pituitary FSH secretion as well as ovarian follicular sensitivity to FSH. The latter molecular mechanism could be fulfilled by regulating Smad5 or Smad9 signals in SYFs, as well as the FSHR and Smad4 signals that affect progesterone synthesis and yolk deposition in the pre-ovulatory follicles.

Receptor Binding Inhibitor Suppresses Carcinogenesis of Cervical Cancer by Depressing Levels of FSHR and ERβ in Mice.

FSH Receptor Binding Inhibitor (FRBI) blocked the binding of FSH to FSHR. Our initial study revealed FRBI reduced the maturation rate, enhanced the apoptosis of sheep Cumulus-Oocyte Complex (COCs). Little is known about whether FRBI modulates ERβ and FSHR levels in the normal uterine and cancerous tissues. The present study aimed to evaluate the FRBI effects on the expressions of Estrogen Receptor-beta (ERβ) and FSH receptor (FSHR) in the uteri. 150 mice were assigned to FRBI+FSH (COM), FSH and control groups (CG). Mice of COM-1, COM-2 and COM-3 groups were simultaneously intramuscularly injected with 500, 750 and 1000 µg FRBI with 10 IU FSH, respectively for five days. Western blotting and qPCR were utilized to determine the expression of ERβ and FSHR. In comparison with FSH group, uterine lumen and glands of COM groups became narrow. The uterine wall and endometrial epithelium were thinned, and uterine lumen became narrow. Epithelial cells were decreased. Uterine wall thicknesses of COM-1, COM-2 and COM-3 groups were reduced by 6.49%, 14.89% and 15.69% on day 30 as compared with FSH group. Uterine perimetrium thicknesses of COM-1, COM-2 and COM-3 groups were reduced by 16.17%, 17.93% and 19.92% on day 20 in comparison with FSH group. Levels of FSHR mRNAs and proteins of COM-1, COM-2 and COM-3 groups were less than FSH group on days 20 and 30 (P<0.05). ERβ protein of COM-3 group was less than FSH group. Serum estradiol (E2) and FSH concentrations of COM-2 and COM-3 were lower than that of FSH group on day 30. FRBI could decrease UWT and UPT, also block the uterine development, decline expression levels of ERβ and FSHR protein. Additionally, FRBI reduced the secretion of secretion of FSH and E2. Downregulating expression of FSHR and ERβ may be a potential treatment regimen for cervical cancer patients.

Divergent expression patterns of pituitary gonadotropin subunit and GnRH receptor genes to continuous GnRH in vitro and in vivo

Continuous, as opposed to pulsatile, delivery of hypothalamic gonadotropin-releasing hormone (GnRH) leads to a marked decrease in secretion of pituitary gonadotropins LH and FSH and impairment of reproductive function. Here we studied the expression profile of gonadotropin subunit and GnRH receptor genes in rat pituitary in vitro and in vivo to clarify their expression profiles in the absence and continuous presence of GnRH. Culturing of pituitary cells in GnRH-free conditions downregulated Fshb, Cga, and Gnrhr expression, whereas continuous treatment with GnRH agonists upregulated Cga expression progressively and Gnrhr and Fshb expression transiently, accompanied by a prolonged blockade of Fshb but not Gnrhr expression. In contrast, Lhb expression was relatively insensitive to loss of endogenous GnRH and continuous treatment with GnRH, probably reflecting the status of Egr1 and Nr5a1 expression. Similar patterns of responses were observed in vivo after administration of a GnRH agonist. However, continuous treatment with GnRH stimulated LH secretion in vitro and in vivo, leading to decrease in LH cell content despite high basal Lhb expression. These data suggest that blockade of Fshb expression and depletion of the LH secretory pool are two major factors accounting for weakening of the gonadotroph secretory function during continuous GnRH treatment.

Remaja pria 18 tahun dengan hipogonadotropik-hipogonadisme dan postur tubuh pendek: sebuah laporan kasus

Background: Hypogonadotropic hypogonadism (HH) is congenital or acquired disease affects hypothalamus or pituitary, so the secretion of GnRH, FSH, LH, and testosterone are inadequate. Clinical manifestations both physical and psychological impair patient’s quality of life, so authors try to report this case.Case description: An 18 years old man complained about a small penis. Small penis cause patient had introvert personality and problems in social relations. On anthropometric status, body weight was below 3rd percentile, height was below 3rd percentile, and body mass index (BMI) was thin. The genital examination was 2nd-grade tanner, penis with 3 cm length and 3 cm circumference, minimal little pubic hair, and testicular volume &lt;2 cc. Ultrasound examination showed no presence of testes in the right or left scrotum fossa, inguinal, and lumbar region. Chromosome analysis showed 46 XY, and hormones analysis found FSH, LH, and testosterone below the normal limit. The patient was given 2000 unit HCG injection and after that FSH, LH, and testosterone hormone increase compared before HCG injections.Conclusion: HH can impair physical and psychological patient’s quality of life through delays in sexual maturation and bone maturation. Strategies to recognize symptoms in early childhood and puberty induction on time can significantly improve the sexual and reproductive function of HH patients.