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PCOS, the most common female endocrine disorder of unknown etiology is characterized by reproductive abnormalities and associated metabolic conditions comprising insulin resistance, Type 2 diabetes mellitus and dyslipidemia. We previously reported that transgenic overexpression of NGF, a marker of sympathetic hyperactivity, directed to the ovary by the mouse 17alpha-hydroxylase promoter (17NF mice), results in ovarian abnormalities similar to those seen in PCOS women (1). To investigate whether ovarian overproduction of NGF also induces common metabolic alterations of PCOS we assessed glucose homeostasis by glucose tolerance test, plasma insulin levels and body composition by Dexa scan in young female 17NF mice compared with WT mice. 17NF mice exhibited increased body weight and alterations in body fat distribution with a greater accumulation of visceral fat compared to subcutaneous fat (p<0.01). 17NF mice also displayed glucose intolerance (p<0.01) and hyperinsulinemia (p<0.05), which, similar to PCOS patients, occurred independently of body weight. Since we have previously established that 17NF mice exhibit increased extrinsic sympathetic input to the ovary (1) we went on to determine whether overexpression of ovarian NGF is capable of altering systemic sympathetic outflow by measuring interscapular brown adipose tissue temperature (iBAT-T) and UCP1 expression. We found that 24h iBAT-T was higher in 17NF mice than WT mice and was significantly different when just the dark period (19:00-07:00) was considered (p<0.05). Interestingly, 17NF mice also exhibited cardiac dysfunction similar to what has been reported in chronic hypertensive subjects. Echocardiography revealed left ventricular enlargement (p<0.01),) and left ventricular posterior wall thinning (p<0.01). Moreover, the internal diameter of the right common carotid artery (RCCA) was increased during systole and diastole (p<0.001). Systolic-diastolic changes in the diameter of the RCCA and area of the left ven

Polycystic ovarian syndrome (PCOS), the most common female endocrine disorder of unknown etiology, is characterized by reproductive abnormalities and associated metabolic conditions comprising insulin resistance, type 2 diabetes mellitus, and dyslipidemia. We previously reported that transgenic overexpression of nerve growth factor (NGF), a marker of sympathetic hyperactivity, directed to the ovary by the mouse 17alpha-hydroxylase/C17-20 lyase promoter (17NF mice), results in ovarian abnormalities similar to those seen in PCOS women. To investigate whether ovarian overproduction of NGF also induces common metabolic alterations of PCOS, we assessed glucose homeostasis by glucose tolerance test, plasma insulin levels, and body composition by dual-energy x-ray absorptiometry scan in young female 17NF mice and wild-type mice. 17NF mice exhibited increased body weight and alterations in body fat distribution with a greater accumulation of visceral fat compared with sc fat (P < .01). 17NF mice also displayed glucose intolerance (P < .01), decreased insulin-mediated glucose disposal (P < .01), and hyperinsulinemia (P < .05), which, similar to PCOS patients, occurred independently of body weight. Additionally, 17NF mice exhibited increased sympathetic outflow observed as increased interscapular brown adipose tissue temperature. This change was evident during the dark period (7 PM to 7 AM) and occurred concomitant with increased interscapular brown adipose tissue uncoupling protein 1 expression. These findings suggest that overexpression of NGF in the ovary may suffice to cause both reproductive and metabolic alterations characteristic of PCOS and support the hypothesis that sympathetic hyperactivity may contribute to the development and/or progression of PCOS.Copyright © 2014 by the Endocrine Society.

Neurotrophins (NTs), once believed to be neural-specific trophic factors, are now known to also provide developmental cuestonon-neural cells. Inthe ovary, NTs contribute to both the formation and development of follicles. Here we show that oocyte-specific deletion of the Ntrk2 gene that encodes the NTRK2 receptor (NTRK2) for neurotrophin-4/5 and brain-derived neurotrophic factor (BDNF) results in post-pubertal oocyte death, loss of follicular organization, and early adulthood infertility. Oocytes lacking NTRK2 do not respond to gonadotropins with activation of phospha-tidylinositol 3-kinase (PI3K)-AKT-mediated signaling. Before puberty, oocytes only express a truncated NTRK2 form (NTRK2. T1), but at puberty full-length (NTRK2. FL) receptors are rapidly induced by the preovulatory gonadotropin surge. A cell line expressing both NTRK2. T1 and the kisspeptin receptor (KISS1R) responds to BDNF stimulation with activation of Ntrk2 expression only if kiss-peptin is present. This suggests that BDNF and kisspeptin that are produced by granulosa cells (GCs) of periovulatory follicles act in concert to mediate the effect of gonadotropins on Ntrk2 expression in oocytes. In keeping with this finding, the oocytes of NTRK2-intact mice fail to respond to go-nadotropins with increased Ntrk2 expression in the absence of KISS1R. Our results demonstrate that the preovulatory gonadotropin surge promotes oocyte survival at the onset of reproductive cyclicity by inducing oocyte expression of NTRK2. FL receptors that set in motion an AKT-mediated survival pathway. They also suggest that gonadotropins activate NTRK2. FL expression via a dual communication pathway involving BDNF and kisspeptin produced in GCs and their respective receptors NTRK2. T1 and KISS1R expressed in oocytes. Copyright © 2014 by the Endocrine Society.

Background: PCOS, the most common female endocrine disorder of unknown etiology is characterized by reproductive abnormalities and associated metabolic conditions comprising insulin resistance, Type 2 diabetes mellitus and dyslipidemia. We previously reported that transgenic overexpression of NGF, a marker of sympathetic hyperactivity, directed to the ovary by the mouse 17alpha-hydroxylase promoter (17NF mice), results in ovarian abnormalities similar to that seen in PCOS women (Dissen et al, 2009). Objective and hypotheses: In the present study we sought to investigate the metabolic profile exhibited by these transgenic mice. Method(s): We studied 17NF mice and its wild type littermate at 10, 15 and 20 weeks of age. We determined glucose homeostasis by glucose tolerance test (GTT) and body composition and bone mineral density (BMD) by Dexa scan. We also measured interscapular brown adipose tissue temperature (iBAT-T) and UCP1 expression as a marker of sympathetic nerve activity Results: We found that at 10 weeks of age 17NF mice display glucose intolerance (p< 0.01). This occurred despite no difference in body weight or total body fat, although 17NF mice showed an increase in visceral fat (p< 0.05). This impairment in glucose tolerance was maintained to 15 and 20 weeks of age when 17NF mice also displayed significantly increased body weight, body fat and visceral fat compared with WT mice. Interestingly, 17NF mice also exhibited an increased BMD from 15 weeks of age compared with WT (p< 0.05). Twenty four hours iBAT-T was higher in 17NF mice than WT at 10 and 20 weeks of age and was significantly different when just the dark period was considered (p< 0.05) . Conclusion(s): These findings suggest an overexpression of NGF in the ovary may be sufficient to cause both reproductive and metabolic alterations that are characteristic of PCOS. Most importantly, this new animal model could therefore allow for the exploration of new treatments of PCOS.

[Abstract] Lin28 and Lin28b are related RNA-binding proteins that inhibit the maturation of miRNAs of the let-7 family and participate in the control of cellular stemness and early embryonic development. Considerable interest has arisen recently concerning other physiological roles of the Lin28/let-7 axis, including its potential involvement in the control of puberty, as suggested by genome-wide association studies and functional genomics. We report herein the expression profiles of Lin28 and let-7 members in the rat hypothalamus during postnatal maturation and in selected models of altered puberty. The expression patterns of c-Myc (upstream positive regulator of Lin28), mir-145 (negative regulator of c-Myc), and mir-132 and mir-9 (putative miRNA repressors of Lin28, predicted by bioinformatic algorithms) were also explored. In male and female rats, Lin28, Lin28b, and c-Myc mRNAs displayed very high hypothalamic expression during the neonatal period, markedly decreased during the infantile-to-juvenile transition and reached minimal levels before/around puberty. A similar puberty-related decline was observed for Lin28b in monkey hypothalamus but not in the rat cortex, suggesting species conservation and tissue specificity. Conversely, let-7a, let-7b, mir-132, and mir-145, but not mir-9, showed opposite expression profiles. Perturbation of brain sex differentiation and puberty, by neonatal treatment with estrogen or androgen, altered the expression ratios of Lin28/let-7 at the time of puberty. Changes in the c-Myc/Lin28b/let-7 pathway were also detected in models of delayed puberty linked to early photoperiod manipulation and, to a lesser extent, postnatal underfeeding or chronic subnutrition. Altogether, our data are the first to document dramatic changes in the expression of the Lin28/let-7 axis in the rat hypothalamus during the postnatal maturation and after different manipulations that disturb puberty, thus suggesting the potential involvement of developmental ch

Excessive nerve growth factor (NGF) production by the ovary, achieved via a transgenic approach, results in arrested antral follicle growth, reduced ovulatory capacity, and a predisposition to cyst formation in response to mildly elevated LH levels. Two salient features in these mutant mice (termed 17NF) are an elevated production of 17alpha-hydroxyprogesterone (17-OHP4), testosterone, and estradiol (E2) in response to gonadotropins, and an increased frequency of granulosa cell (GC) apoptosis. In this study, we show that the increase in steroidal response is associated with enhanced expression of Cyp17a1, Hsd17b, and Cyp19a1, which encode the enzymes catalyzing the synthesis of 17-OHP4, testosterone, and E2 respectively. Using a proteomic approach, we identified stathmin (STMN1), as a protein that is overproduced in 17NF ovaries. In its phosphorylated state, STMN1 mediates a cell death signal initiated by tumor necrosis factor alpha (TNF). STMN1 is expressed in GCs and excessive NGF increases its abundance as well as that of its forms phosphorylated at serine (Ser) 16, 25, and 38. TNF synthesis is also increased in 17NF ovaries, and this change is abolished by blocking neurotrophic tyrosine kinase receptors. Inhibiting TNF actions in vivo by administering a soluble TNF receptor prevented the increase in total and phosphorylated STMN1 production, as well as GC apoptosis in NGF-overproducing ovaries. These results indicate that an excess of NGF in the ovary promotes steroidogenesis by enhancing the expression of enzyme genes involved in 17-OHP4, testosterone, and E2 synthesis, and causes GC apoptosis by activating a TNF/ STMN1-mediated cell death pathway. © 2011 Society for Reproduction and Fertility.

Current evidence suggests that the acquisition of female reproductive capacity and the maintenance of mature reproductive function are related processes transcriptionally regulated by gene networks operating within the neuroendocrine brain. One of these genes, termed enhanced at puberty1(EAP1), encodes an upstream regulator of these processes. Selective inhibition of EAP1 expression in discrete regions of the rat and nonhuman primate (NHP) hypothalamus, via targeted delivery of RNA interference, either disrupts (rats) or abolishes (monkeys) reproductive cycles. The striking loss of menstrual cyclicity resulting from knocking down hypothalamic EAP1 expression suggests that diminished EAP1 function may contribute to disorders of the menstrual cycle of neuroendocrine origin. Here we show that a single-nucleotide polymorphism in the 5'-flanking region of EAP1 gene is associated with increased incidence of amenorrhea/oligomenorrhea in NHP. In the presence of the risk allele, binding of the transcription factor mothers against decapentaplegic homolog 3 (SMAD3) to its recognition site contained within the polymorphic sequence in the monkey EAP1 promoter is reduced. The risk allele also diminishes the increase in EAP1 promoter activity elicited by TGFbeta1, a peptide that activates a SMAD3/4-mediated signaling pathway to regulate gene transcription. These findings indicate that common genetic variation in the EAP1 locus increases the susceptibility of NHP to loss/disruption of menstrual cyclicity. They also raise the possibility that polymorphisms in EAP1 may increase the risk of functional hypothalamic amenorrhea in humans. Copyright © 2012 by The Endocrine Society.

Tropomyosin-related kinase (TRK) receptor B (TRKB) mediates the supportive actions of neurotrophin 4/5 and brain-derived neurotrophic factor on early ovarian follicle development. Absence of TRKB receptors reduces granulosa cell (GC) proliferation and delays follicle growth. In the present study, we offer mechanistic insights into this phenomenon. DNA array and quantitative PCR analysis of ovaries from TrkB-null mice revealed that by the end of the first week of postnatal life, Jagged1, Hes1, and Hey2 mRNA abundance is reduced in the absence of TRKB receptors. Although Jagged1 encodes a NOTCH receptor ligand, Hes1 and Hey2 are downstream targets of the JAGGED1-NOTCH2 signaling system. Jagged1 is predominantly expressed in oocytes, and the abundance of JAGGED1 is decreased in TrkB-/- oocytes. Lack of TRKB receptors also resulted in reduced expression of c-Myc, a NOTCH target gene that promotes entry into the cell cycle, but did not alter the expression of genes encoding core regulators of cell-cycle progression. Selective restoration of JAGGED1 synthesis in oocytes of TrkB-/- ovaries via lentiviral-mediated transfer of the Jagged1 gene under the control of the growth differentiation factor 9 (Gdf9) promoter rescued c-Myc expression, GC proliferation, and follicle growth. These results suggest that neurotrophins acting via TRKB receptors facilitate early follicle growth by supporting a JAGGED1-NOTCH2 oocyte-to-GC communication pathway, which promotes GC proliferation via a c-MYC-dependent mechanism. Copyright © 2011 by The Endocrine Society.

Serono Symposia International convened an expert panel to review the impact of environmental influences on the regulation of pubertal onset and progression while identifying critical data gaps and future research priorities. An expert panel reviewed the literature on endocrine-disrupting chemicals, body size, and puberty. The panel concluded that available experimental animal and human data support a possible role of endocrine-disrupting chemicals and body size in relation to alterations in pubertal onset and progression in boys and girls. Critical data gaps prioritized for future research initiatives include (1) etiologic research that focus on environmentally relevant levels of endocrine-disrupting chemicals and body size in relation to normal puberty as well as its variants, (2) exposure assessment of relevant endocrine-disrupting chemicals during critical windows of human development, and (3) basic research to identify the primary signal(s) for the onset of gonadotropin-releasing hormone-dependent/central puberty and gonadotropin-releasing hormone-independent/peripheral puberty. Prospective studies of couples who are planning pregnancies or pregnant women are needed to capture the continuum of exposures at critical windows while assessing a spectrum of pubertal markers as outcomes. Coupled with comparative species studies, such research may provide insight regarding the causal ordering of events that underlie pubertal onset and progression and their role in the pathway of adult-onset disease Udgivelsesdato: 2008/2