Bisphenol A (BPA) causes reproductive toxicities, but the mechanisms are still unclear. In the present study, we sought to clarify these mechanisms in comparison with those of 17beta-estradiol (E2). Prepubertal Wistar/ST male rats (4 weeks old) were subcutaneously administered BPA (0, 20, 100 and 200 mg/kg/day) or E2 (10 and 100 microg/kg/day) for 6 weeks. Both BPA and E2 treatments decreased plasma and testicular testosterone levels, and plasma luteinizing hormone (LH), but not E2 and follicle-stimulating hormone levels, though E2 treatment increased its plasma level. In relation to the decreased testosterone levels, BPA and E2 decreased expressions of steroidogenic enzymes and cholesterol carrier protein in Leydig cells. Thus, decreased testosterone levels in plasma might have resulted from decreased expressions of these enzymes and protein as well as from decreased plasma LH levels. Interestingly, the changes in steroidogenic enzymes and carrier protein were observed at lower levels of exposure to BPA or E2 than those inhibiting plasma LH levels. Microscopically, 200 mg/kg BPA and 100 microg/kg E2 significantly decreased Leydig cell numbers in the testis. In addition, BPA and E2 also decreased expression of estrogen receptor alpha-mRNA, which might be related to the decreased numbers of Leydig cells. Thus, BPA directly affects not only the Leydig cells but also the pituitary gland, but the former may be impaired at lower exposure concentrations than the latter.

Sex hormones regulate cholangiocyte hyperplasia in bile duct-ligated (BDL) rats. We studied whether follicle-stimulating hormone (FSH) regulates cholangiocyte proliferation. FSH receptor (FSHR) and FSH expression was evaluated in liver sections, purified cholangiocytes, and cholangiocyte cultures (NRICC). In vivo, normal female and male rats were treated with FSH or immediately after BDL with antide (a gonadotropin-releasing hormone antagonist blocking FSH secretion) or a neutralizing FSH antibody for 1 wk. We evaluated 1) cholangiocyte proliferation in sections and cholangiocytes and 2) changes in secretin-stimulated cAMP (functional index of cholangiocyte growth) levels, and ERK1/2 and Elk-1 phosphorylation. NRICC were stimulated with FSH before evaluation of proliferation, cAMP/IP(3) levels, and ERK1/2 and Elk-1 phosphorylation. To determine whether FSH regulates cholangiocyte proliferation by an autocrine mechanism, we evaluated the effects of 1) cholangiocyte supernatant (containing FSH) on NRICC proliferation and 2) FSH silencing in NRICC before measuring proliferation and ERK1/2 and Elk-1 phosphorylation. Cholangiocytes and NRICC express FSHR and FSH and secrete FSH. In vivo administration of FSH to normal rats increased, whereas administration of antide and anti-FSH antibody to BDL rats decreased 1) ductal mass and 2) secretin-stimulated cAMP levels, proliferation, and ERK1/2 and Elk-1 phosphorylation in cholangiocytes compared with controls. In NRICC, FSH increased cholangiocyte proliferation, cAMP levels, and ERK1/2 and Elk-1 phosphorylation. The supernatant of cholangiocytes increased NRICC proliferation, inhibited by preincubation with anti-FSH antibody. Silencing of FSH gene decreases cholangiocyte proliferation and ERK1/2 and Elk-1 phosphorylation. Modulation of cholangiocyte FSH expression may be important for the management of cholangiopathies.

Japanese flounder (Paralichthys olivaceus), a teleost fish that has XX (female)/XY (male) sex determination system, exhibits temperature-dependent sex determination. We have previously shown that high water temperature or an aromatase inhibitor treatment causes the sex-reversal from genetic females to phenotypic males and suppression of mRNA expression of ovary-type P450 aromatase (cyp19a1), a steroidogenic enzyme responsible for the conversion of androgens to estrogens, in Japanese flounder. In the present study, we demonstrate that high water temperature treatment suppresses specifically mRNA expression of the forkhead transcription factor gene foxl2, and follicle-stimulating hormone receptor (fshr) in gonads during early sex differentiation. Moreover, transient transfection assay shows that the flounder Foxl2 and cAMP analog can activate the cyp19a1 gene transcription in vitro. These results strongly suggest that FSH signaling and Foxl2 are involved in the transcriptional regulation of cyp19a1 gene during gonadal sex differentiation in Japanese flounder with temperature-dependent sex determination.

We have investigated the biological characteristics of an immortalized granulosa cell line (COV434), which may be used to study follicular and oocyte maturation in vitro. Granulosa cell function was defined as consisting of three distinct properties: (i) production of 17beta-oestradiol in response to follicle stimulating hormone (FSH); (ii) presence of specific molecular markers of apoptosis enabling the induction of follicular atresia; and (iii) capacity to form intercellular connections with cells surrounding an oocyte. The addition of FSH to the culture medium supplemented with 10% fetal calf serum and 4-androstene-3,17-dione resulted in proliferation of the COV434 granulosa cells and in an increased synthesis of 17beta-oestradiol, indicating the presence of the FSH receptor and cytochrome P450 aromatase in these cells. The receptor for luteinizing hormone (LH) was undetectable. Similar expression of various apoptosis-associated genes was found in COV434 granulosa cells and in granulosa cells of patients stimulated with gonadotrophins for in-vitro fertilization, thus indicating that the immortalized COV434 granulosa cells were able to sustain apoptosis. Multiple intercellular connections were formed during co-culture of COV434 granulosa cells with cumulus cells containing an immature oocyte but not with cumulus cells devoid of an oocyte. Detailed morphological analysis of the intercellular connections with scanning electron microscopy and confocal light microscopy demonstrated the presence of long slender structures. It is concluded that the immortalized human granulosa cell line COV434 may be useful for experimental studies on follicular development.

OBJECTIVE

We sought to establish normative data for spontaneous and gonadotropin-releasing hormone (GnRH)-stimulated serum luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels measured by new immunochemiluminometric assays (ICMA) in children and adolescents.

METHODS

Random serum samples were obtained from 375 normal subjects (0.1 to 17.7 years, 230 female subjects). Intravenous GnRH stimulation tests were performed in 41 normal subjects (4.8 to 18 years, 20 female subjects). Normal ranges were calculated by age and Tanner stage. Immunochemiluminometric assays of LH and FSH concentrations were compared with levels obtained by a sensitive immunofluorometric assay and a less sensitive radioimmunoassay.

RESULTS

Random gonadotropin concentrations in normal children followed the pattern of transient elevation in infancy, low but measurable prepubertal levels, and markedly increased values at puberty. Spontaneous LH levels were higher in male infants but were not statistically different in boys and girls after infancy. Mean prepubertal LH was 0.04 +/- 0.04 IU/L (n = 66), rising 100-fold during puberty. Spontaneous FSH levels were much higher than LH values, were higher in female infants, and rose threefold at puberty. Peak GnRH-stimulated LH was identical in prepubertal boys and girls (1.8 +/- 1.3 IU/L, n = 17) and increased 20-fold at puberty. Mean peak GnRH-stimulated FSH was highest in prepubertal female subjects. Luteinizing hormone values measured by ICMA and immunofluorometric assay were highly correlated, but radioimmunoassay levels diverged markedly from ICMA levels at lower concentrations. Because absolute levels were higher, FSH values correlated adequately in the three assays throughout the normal physiologic range.

CONCLUSIONS

Measurement of LH by ICMA is much more sensitive than older assay methods. Spontaneous LH can be accurately measured by ICMA to the very low levels present in normal prepubertal children, providing a potentially important biochemical discriminator of pubertal status. An ICMA GnRH-stimulated LH level greater than 5 IU/L is suggestive of maturing gonadotropin secretion. The ICMA LH assays provide significant enhancement in sensitivity; these assays should be used when levels may be low, and by their accuracy may reduce the time and expense of testing procedures.

OBJECTIVE

Smoking is associated with increased follicle-stimulating hormone levels and early menopause. Smoking may directly accelerate ovarian follicular depletion or may act indirectly by increasing the pituitary production of follicle-stimulating hormone. Antimüllerian hormone (AMH), produced by ovarian follicles, is a more direct measure of ovarian reserve. The objective of our study was to determine the extent to which smoking influences ovarian reserve, as measured by AMH levels.

METHODS

A community sample of 284 women aged 38 to 50 years completed a self-administered questionnaire including a detailed smoking history. Serum AMH levels were measured on day 2, 3, or 4 of the menstrual cycle. The association between AMH and smoking was analyzed using linear regression, adjusting for age and body mass index.

RESULTS

Participants aged 38 to 42, 43 to 45, and 46 to 50 years had geometric mean AMH values of 6.7 pM (95% CI, 5.2-8.7 pM), 2.7 pM (95% CI, 1.9-3.8 pM), and 1.3 pM (95% CI, 1.0-1.7 pM), respectively. Current smokers, but not past smokers, had 44% lower AMH values than did the reference group (participants with neither active nor former or passive smoke exposure; P = 0.04). Passive smoking had no effect on AMH values when compared with the reference group (P = 0.55). The impact of smoking on AMH values was not dose dependent based on cigarettes per day (P = 0.08) or pack-years (P = 0.22). Finally, prenatal exposure to smoking (either maternal or paternal) had no impact on AMH levels (P = 0.47 and P = 0.89, respectively).

CONCLUSIONS

Active smoking, but not former smoking, is associated with decreased AMH values in late-reproductive-age and perimenopausal women, suggesting a possible direct effect of smoking on the depletion of the antral but not primordial follicles. The direct impact of active smoking on AMH levels in younger women requires further investigation.

To assess adult stem cell differentiation in the testis, we injected bone marrow cells from adult green fluorescent protein (GFP) transgenic mice into the seminiferous tubules and the testicular interstitium of busulfan-treated wild-type or c-kit mutant (W/W(v)) mice. Ten to 12 weeks after transplantation, we examined the fate of the transplanted bone marrow cells and found that they survived in recipient testes. In both the busulfan-treated and W/W(v) mice, some of the GFP-positive donor cells had a Sertoli cell appearance and expressed follicle-stimulating hormone receptor within the seminiferous tubules. In addition, GFP-positive donor cells were found in the interstitium of recipient testes, and they expressed the cytochrome P450 side chain cleavage enzyme (P450scc). In the seminiferous tubules of busulfan-treated mice, GFP-positive donor cells had the appearance of spermatogonia or spermatocytes and expressed VASA. However, this was not found in the seminiferous tubules of W/W(v) mice. We conclude that adult bone marrow cells, in a favorable testicular environment, differentiate into somatic and germ cell lineages. The resident neighboring cells in the recipient testis may control site-appropriate stem cell differentiation. This clinically relevant finding raises the possibility for treatment of male infertility and testosterone deficiency through the therapeutic use of stem cells.

The hypothalamic hormone gonadotropin-releasing hormone (GnRH) is released in a pulsatile fashion, with its frequency varying throughout the reproductive cycle. Varying pulse frequencies and amplitudes differentially regulate the biosynthesis and secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) by pituitary gonadotropes. The mechanism by which this occurs remains a major question in reproductive physiology. Previous studies have been limited by lack of available cell lines that express the LH and FSH subunit genes and respond to GnRH. We have overcome this limitation by transfecting the rat pituitary GH3 cell line with rat GnRH receptor (GnRHR) cDNA driven by a heterologous promoter. These cells, when cotransfected with regulatory regions of the common alpha, LH beta, or FSH beta subunit gene fused to a luciferase reporter gene, respond to GnRH with an increase in luciferase activity. Using this model, we demonstrate that different cell surface densities of the GnRHR result in the differential regulation of LH and FSH subunit gene expression by GnRH. This suggests that the differential regulation of gonadotropin subunit gene expression by GnRH observed in vivo in rats may, in turn, be mediated by varying gonadotrope cell surface GnRHR concentrations. This provides a physiologic mechanism by which a single ligand can act through a single receptor to regulate differentially the production of two hormones in the same cell.

Gonadal quiescence prior to puberty in primates results from a diminished secretion of the pituitary gonadotropic hormones, follicle-stimulating hormone and luteinizing hormone, which, in turn, is occasioned by an interruption of pulsatile release of gonadotropin-releasing hormone (GnRH) from the hypothalamus during this phase of development. A discharge of GnRH may be provoked from the hypothalamus of prepubertal monkeys, however, by an i.v. injection of N-methyl-D-aspartate (NMDA), an analog of the putative excitatory neurotransmitter, aspartate. Since this action of NMDA is blocked by the specific NMDA receptor antagonist, DL-2-amino-5-phosphonopentanoic acid, the release of GnRH is likely mediated by NMDA receptors located either on the GnRH neurons themselves or on afferents to the GnRH cells. We report here that prolonged intermittent NMDA stimulation of GnRH neurons within the hypothalamus of the juvenile monkey for 16-30 wk results, with surprising ease, in the onset of precocious puberty with full activation of the hypothalamic-pituitary-Leydig cell axis and initiation of spermatogenesis. These findings demonstrate that, in primates, the network of hypothalamic GnRH neurons, which in adulthood provides the drive to the gonadotropin-secreting cells of the anterior pituitary gland, must now be viewed together with the pituitary and gonads as a nonlimiting component of the control system that governs the onset of puberty in these species.

OBJECTIVE

To investigate whether anti-Müllerian hormone (AMH) is better than antral follicle count (AFC) in predicting oocyte yield and embryo quality after controlled ovarian hyperstimulation for in vitro fertilization (IVF).

METHODS

This is a prospective observational study involving 162 women (<40 years old) undergoing their first IVF cycle at an IVF unit within a university hospital. AMH and AFC measurements were made on day 3 of the cycle within 3 months of starting ovarian stimulation. A standard long down-regulation protocol using gonadotrophin releasing hormone agonist and recombinant follicle stimulating hormone was used. A maximum of two embryos were transferred on day 2 or 3 following oocyte retrieval. The primary outcome was the number of good quality embryos available for transfer and freezing. Embryos were graded according to the number of blastomeres, the difference in blastomere size and the degree of fragmentation, into grades 1-4. Secondary outcomes included the number of oocytes retrieved and fertilized and the live birth rate. Correlation between different parameters was calculated using Spearman's correlation coefficient. Receiver operating characteristic (ROC) curves were generated for AMH and AFC to compare ability of parameters to predict top quality or frozen embryos and the occurrence of a live birth.

RESULTS

Of the 137 women who had fresh embryo transfer, 52 became pregnant (32.1% pregnancy rate per cycle started) and 38 had a live birth (23.5% live birth rate per cycle started). Both AMH and AFC had highly significant correlations with the number of oocytes retrieved and the number of oocytes fertilized (P<0.001). The two markers were also significantly associated with the number of top quality embryos available for transfer and the number of embryos frozen (P<0.01). With regard to live birth, AMH performed better than AFC (P<0.01 and P<0.05, respectively), but both markers were more valuable in predicting the absence rather than the occurrence of live birth (negative predictive value 84%).

CONCLUSIONS

AMH and AFC are comparable predictors of oocytes retrieved and of the number of good quality embryos available for transfer and freezing. Prediction of live birth may help clinicians selecting patients suitable for single embryo transfer.

As both gonadotropins, LH and FSH, are required for normal spermatogenesis, patients with pituitary insufficiency need hCG plus human menopausal gonadotropin therapy to induce spermatogenesis and establish fertility. In a patient hypophysectomized because of a pituitary tumor, who, despite undetectable serum gonadotropin levels, had normal testis volume and semen parameters and fathered three children under testosterone substitution alone, we hypothesized an activating mutation of the FSH receptor. Exon 10 of the FSH receptor gene was amplified from genomic DNA by PCR, screened by single stranded conformation polymorphism gel electrophoresis, and sequenced. We identified a heterozygous A->>G base change at nucleotide position 1700, leading to an Asp->>Gly transition in codon 567 in the third intracytoplasmatic loop. COS-7 cells transiently transfected with the mutated receptor displayed a 1.5-fold increase in basal cAMP production compared to wild-type receptor, indicating that this mutation leads to ligand-independent constitutive activation of the FSH receptor. We conclude that this activating mutation of the FSH receptor, the first ever described, autonomously sustains spermatogenesis in the absence of gonadotropins.

A variety of hypotheses have been proposed to explain the premenstrual syndromes. These hypotheses serve as rationales for an equally diverse range of proposed treatments. To investigate these hypotheses, we obtained multiple blood samples across the menstrual cycle in women with well-characterized menstrually related mood disorder and in control subjects. No diagnosis-related differences were observed in the levels or patterns of secretion of progesterone, estradiol, follicle-stimulating hormone, luteinizing hormone, testosterone-estradiol-binding globulin, dehydroepiandrosterone sulfate, dihydrotestosterone, prolactin, or cortisol. Our data suggest that premenstrual syndrome does not represent a simple hormonal deficiency and that the cited rationales for several of the proposed treatments are of questionable merit.

FSH promoted the rapid phosphorylation of the nuclear protein histone H3 in immature rat ovarian granulosa cells under experimental conditions that lead to cellular differentiation and not proliferation. FSH-stimulated histone H3 phosphorylation correlated with cAMP-dependent protein kinase A (PKA) activation and translocation of the PKA catalytic subunit to a nuclear-enriched fraction and was inhibited by the PKA inhibitor H89, and histone H3 phosphorylation was stimulated in cells treated with agents that raise intracellular cAMP levels such as forskolin and 8-bromo-cAMP. FSH-stimulated histone H3 phosphorylation in granulosa cells mapped to ser-10, a site previously identified as the PKA phosphorylation site in various mitotically active cells as the mitosis-specific phosphorylation site. Injection of the FSH analog PMSG to immature rats, which is known to stimulate granulosa cell proliferation as well as differentiation, also promoted histone H3 phosphorylation on ser-10 in granulosa cells. These results establish that FSH-stimulated histone H3 phosphorylation in granulosa cells is linked not only to granulosa cell mitosis but also to granulosa cell differentiation and that FSH-stimulated histone H3 phosphorylation on ser-10 in isolated granulosa cells is mediated by PKA. These results also identify the PKA-dependent histone H3 phosphorylation as an early nuclear protein marker for FSH-stimulated differentiation of granulosa cells. Based on the recently described function of histone H3 as a coactivator of transcription, these results are consistent with the hypothesis that phosphorylated histone H3 may facilitate PKA-dependent gene transcription in granulosa cells leading to the preovulatory phenotype.

OBJECTIVE

The recovery of immature oocytes followed by in-vitro maturation (IVM) and in-vitro fertilization is an attractive alternative to conventional in-vitro fertilization treatment in which controlled ovarian stimulation with gonadotropins is used to increase the number of available oocytes and embryos. Significant progress has been made to improve pregnancy and implantation rates from in-vitro matured oocytes. This review summarizes current knowledge and achievements in human oocyte in-vitro maturation for clinical application, and will highlight recent advances reported in in-vitro maturation treatment.

RESULTS

It has been demonstrated that priming of ovarian immature oocytes with follicle-stimulating hormone or human chorionic gonadotropin prior to immature oocyte retrieval improves oocyte maturation rates and embryo quality as well as pregnancy rates in infertile women with polycystic ovaries or polycystic ovary syndrome. The size of follicles may be important for the subsequent embryonic development, but the developmental competence of oocytes derived from the small antral follicles is not adversely affected by the presence of a dominant follicle. However oocyte maturation in vitro is profoundly affected by culture conditions. Currently more than 300 healthy infants have been born following immature oocyte retrieval and in-vitro maturation. In general, the clinical pregnancy and implantation rates have reached 30-35% and 10-15% respectively in infertile women with polycystic ovaries or polycystic ovary syndrome.

CONCLUSIONS

In-vitro maturation treatment can now be offered as a successful option to infertile women with polycystic ovaries or polycystic ovary syndrome. It is possible to combine natural cycle in-vitro fertilization with immature oocyte retrieval followed by in-vitro maturation, and thus offer women with various causes of infertility reasonable pregnancy and implantation rates without recourse to ovarian stimulation. Further research remains to be done to address the mechanism of oocyte maturation in order to refine culture conditions and improve the implantation rate of oocytes matured in vitro.

Studies performed in vitro suggest that a novel 12 amino acid RF amide peptide, isolated from the quail hypothalamus, is a gonadotrophin inhibitory hormone (GnIH). The aim of the present study was to investigate this hypothesis in the domestic chicken. Injections of GnIH into nest-deprived incubating hens failed to depress the concentration of plasma luteinizing hormone (LH). Addition of GnIH to short-term (120 min) cultures of diced pituitary glands from adult cockerels depressed follicle-stimulating hormone (FSH) and LH release and depressed common alpha and FSHbeta gonadotrophin subunit mRNAs, with no effect on LHbeta subunit mRNA. Hypothalamic GnIH mRNA was higher in incubating (out-of-lay) than in laying hens, but there was no significant difference in the amount of hypothalamic GnIH mRNA in out-of-lay and laying broiler breeder hens at the end of a laying year. It is concluded that avian GnIH may play a role in controlling gonadotrophin synthesis and associated constitutive release in the domestic chicken.

To examine whether the concomitant administration of a gonadotrophin-releasing hormone agonist (GnRHa) during combination chemotherapy to young women with lymphoma may facilitate preservation of gonadal function, a prospective clinical protocol was undertaken in 18 cycling women with lymphoma, aged 15-40 years. Thirteen patients suffered from Hodgkin disease (HD) and 5 from non-Hodgkin lymphoma. After informed consent a monthly injection of depot D-TRP6-GnRHa was administered for a maximum of 6 months starting prior to chemotherapy. Most of these patients (15/18) were treated with the MOPP/ABV(D) combination chemotherapy followed by mantle field irradiation in 10 patients. Hormonal profile [luteinizing hormone (LH), follicle stimulating hormone (FSH), oestradiol, testosterone, progesterone, insulin-like growth factor (IGF)-1, prolactin] was taken before the GnRHa/chemotherapy co-treatment, and monthly thereafter until resuming spontaneous ovulation and menses. This group of prospectively treated lymphoma patients was compared to a matched control group of 18 women (aged 17-40 years) who have been treated with chemotherapy, mostly MOPP/ABV (14/18), with (11) or without (7) mantle field radiotherapy. Fourteen had Hodgkin's and four non-Hodgkin's lymphoma. Gonadal function was determined clinically, hormonally (LH, FSH, oestradiol, progesterone), and sonographically. Two of the patients in each group died from refractory disease. Of the remaining 16 patients, 15 (93.7%) resumed spontaneous ovulation and menses within 3-8 months of termination of the combined chemotherapy/GnRHa co-treatment. In contrast, only seven (39%) of the 18 similarly treated patients in the control group (chemotherapy without GnRHa) resumed ovarian cyclic activity (regular menses). The other 11 experienced premature ovarian failure (POF) (61%). Out preliminary data suggest a possible significant protective effect of GnRHa co-treatment with chemotherapy from irreversible ovarian damage (POF).

OBJECTIVE

Initial results of a randomized trial comparing carboplatin with radiotherapy (RT) as adjuvant treatment for stage I seminoma found carboplatin had a noninferior relapse-free rate (RFR) and had reduced contralateral germ cell tumors (GCTs) in the short-term. Updated results with a median follow-up of 6.5 years are now reported.

METHODS

Random assignment was between RT and one infusion of carboplatin dosed at 7 × (glomerular filtration rate + 25) on the basis of EDTA (n = 357) and 90% of this dose if determined on the basis of creatinine clearance (n = 202). The trial was powered to exclude a doubling in RFRs assuming a 96-97% 2-year RFR after radiotherapy (hazard ratio [HR], approximately 2.0).

RESULTS

Overall, 1,447 patients were randomly assigned in a 3-to-5 ratio (carboplatin, n = 573; RT, n = 904). RFRs at 5 years were 94.7% for carboplatin and 96.0% for RT (RT-C 90% CI, 0.7% to 3.5%; HR, 1.25; 90% CI, 0.83 to 1.89). One death as a result of seminoma (in RT arm) occurred. Patients receiving at least 99% of the 7 × AUC dose had a 5-year RFR of 96.1% (95% CI, 93.4% to 97.7%) compared with 92.6% (95% CI, 88.0% to 95.5%) in those who received lower doses (HR, 0.51; 95% CI, 0.24 to 1.07; P = .08). There was a clear reduction in the rate of contralateral GCTs (carboplatin, n = 2; RT, n = 15; HR, 0.22; 95% CI, 0.05 to 0.95; P = .03), and elevated pretreatment follicle-stimulating hormone (FSH) levels >> 12 IU/L) was a strong predictor (HR, 8.57; 95% CI, 1.82 to 40.38).

CONCLUSIONS

These updated results confirm the noninferiority of single dose carboplatin (at 7 × AUC dose) versus RT in terms of RFR and establish a statistically significant reduction in the medium term of risk of second GCT produced by this treatment.

OBJECTIVE

Telomeres are DNA repeats which cap and protect chromosome ends, facilitate homologue pairing and chiasmata formation during early meiosis, and shorten with cell division and exposure to reactive oxygen to mediate aging. Early germ cells contain telomerase, a reverse transcriptase which adds telomeres to 3-prime DNA ends, but telomerase activity declines in oocytes, fixing telomere length earlier during development. Experimentally induced telomere shortening in mice disrupts meiosis, impairs chiasmata formation, halts embryonic cell cycles, and promotes apoptosis in embryos, a phenotype which mimics reproductive senescence in women. Ethical constraints limit study of human embryos to nondestructive assays, such as morphologic evaluation under transmission optics, but cytoplasmic fragmentation is a reliable marker of apoptosis.

METHODS

Study design consisted of observational study of effect of telomere length in human eggs on cytoplasmic fragmentation, and on other morphologic features of preimplantation embryos. To test the hypothesis that telomere shortening triggers apoptosis in human embryos, we evaluated telomere length as a predictor of cytoplasmic fragmentation in embryos from women undergoing in vitro fertilization.

RESULTS

Telomere length negatively predicted fragmentation in day 3 preimplantation embryos, after controlling for patient age and basal follicle stimulating hormone level. Telomere length did not predict other features of preimplantation embryo morphology.

CONCLUSIONS

The finding that telomere length in human eggs predicts cytoplasmic fragmentation in embryos provides evidence that telomere shortening induces apoptosis in human preimplantation embryos, consistent with a telomere theory of reproductive senescence in women.

Activin was initially described as a protein that stimulates release of follicle stimulating hormone from the pituitary, and it is well known for its important roles in different reproductive functions. In recent years, this multifunctional factor has attracted the attention of researchers in other fields, as new functions of activin in angiogenesis, inflammation, immunity, fibrosis and cancer have been discovered. Studies from our laboratory have identified activin as a crucial regulator of wound healing and skin carcinogenesis. On the one hand, it strongly accelerates the healing process of skin wounds but, on the other hand, it enhances scar formation and the susceptibility to skin tumorigenesis. Finally, results from several laboratories have revealed that activin enhances tumour formation and/or progression in some other organs, in particular through its effect on the tumour microenvironment, and that it also promotes cancer-induced bone disruption and muscle wasting. These findings provide the basis for the use of activin or its downstream targets for the improvement of impaired wound healing, and of activin antagonists for the prevention and treatment of fibrosis and of malignant tumours that overexpress activin. Here, we summarize the previously described roles of activin in wound healing and scar formation and discuss functional studies that revealed different functions of activin in the pathogenesis of cancer. The relevance of these findings for clinical applications will be highlighted.

The hormonal responses of nine male, strength athletes to strength exercise were examined. The athletes performed one moderate- and one high-intensity strength exercise workout. In the high-intensity workout, the load was 100% of each subject's three-repetition maximum (3-RM) for squats and front squats, and 100% of each subject's six-repetition maximum (6-RM) for leg extensions. In the moderate-intensity workout, the load was 70% of the high-intensity protocol. Rest periods between sets were 4-6 min for both workouts. Blood samples were taken before, 30 min into, and every 15 min for the 1st h after exercise, and then 3, 7, 11, 22 and 33 h after exercise, thus allowing examination of both the acute and prolonged hormonal responses. Blood samples were analyzed for testosterone, luteinizing hormone (LH), follicle stimulating hormone (FSH), cortisol, adrenocorticotrophic hormone (ACTH), growth hormone (GH), insulin-like growth factor (IGF-1), insulin, sex hormone binding globulin, creatine kinase, total protein, glucose and lactate. The acute responses of testosterone and cortisol were greater during the high-intensity protocol as compared to the moderate-intensity protocol. The cortisol response was associated with an increase in ACTH concentration. LH and FSH showed no response to either protocol. Acute GH responses were not different between protocols. There were great inter-individual differences in acute GH responses to both protocols. There were no significant differences between protocols with regard to prolonged responses for any hormone. In both trials, IGF-1 concentrations were significantly lower at 0800 hours the morning after exercise as compared to concentrations found at 0800 hours the morning before exercise. The mechanisms responsible for reducing IGF-1 concentration in these trials are unclear, and it is not known if this reduction observed 22 hours after exercise is of physiological significance.

BACKGROUND

In late reproductive-aged breast cancer survivors, there is a need for real-time biomarkers of postchemotherapy ovarian function. The objective was to determine whether antimullerian hormone (AMH) and inhibin B are such biomarkers. The authors tested whether AMH and inhibin B were impacted by breast cancer treatment by comparing cancer survivors to age-matched control women and determined the association between these hormones and postchemotherapy menstrual pattern.

METHODS

Breast cancer patients (n = 127) with American Joint Committee on Cancer stage I to III disease who were premenopausal at diagnosis were enrolled postchemotherapy and observed. The primary endpoint was chemotherapy-related amenorrhea (CRA) >> or = 12 months of amenorrhea after chemotherapy). Matched pair analyses compared AMH, inhibin B, and follicle-stimulating hormone (FSH) levels between cancer and age-matched control subjects. Associations between hormones, CRA status, and change in CRA status over time were assessed.

RESULTS

The median age of the patients at chemotherapy was 43.2 years (range, 26.7-57.8 years). At enrollment, median follow-up since chemotherapy was 2.1 years, and 55% of subjects had CRA. Compared with age-matched controls, cancer subjects had significantly lower AMH (P = .004) and inhibin B (P < .001) and higher FSH (P < .001). AMH (P = .002) and inhibin B (P = .001) were found to be significantly associated with risk of CRA, even after controlling for FSH. AMH was significantly lower (P = .03) and FSH was significantly higher (P = .04) in menstruating subjects who developed subsequent CRA.

CONCLUSIONS

AMH and inhibin B are 2 additional measures of postchemotherapy ovarian function in late reproductive-aged breast cancer survivors. With further research and validation, these hormones may supplement limited current tools for assessing and predicting postchemotherapy ovarian function.

OBJECTIVE

We establish whether a subset of infertile men have decreased serum testosterone-to-estradiol ratios and whether this condition can be corrected with an oral aromatase inhibitor.

METHODS

The serum testosterone-to-estradiol ratios of 63 men with severe male factor infertility or hypergonadotropic hypogonadism (mean follicle-stimulating hormone 21.2 +/- 1.8) were compared with those of an age matched, fertile, control reference group. Of the 63 men 43 were azoospermic with biopsy proved severe male infertility and 20 were oligospermic. The men with the lowest ratios (less than 20th percentile) were treated with 50 to 100 mg of the aromatase inhibitor testolactone orally twice daily. Testosterone-to-estradiol ratios and semen analyses were evaluated during testolactone therapy.

RESULTS

Men with severe male infertility had significantly lower testosterone (328 versus 543 ng/dl, p <0.01) and higher estradiol (58.4 versus 43.5 ng/l, p = 0.01) than fertile control reference subjects, resulting in a decreased testosterone-to-estradiol ratio (x10(-1) = 6.9 +/- 0.6 versus 14.5 +/- 1.2, respectively, p <0.01). Of the 45 men treated with testolactone a correction of these abnormalities was seen and ratios (x10(-1)) increased into the normal range (5.0 +/- 0.3 to 12.7 +/- 1.2, p <0.01). Semen analyses were considered evaluable only in men with sperm in the ejaculate before aromatase inhibitor treatment. Semen analyses before and during testolactone treatment revealed significant increases in sperm concentration (16.1 to 28.9 million sperm per ml, p = 0.03) and motility (27.1% to 45.3%, p <0.01) in 12 oligospermic men.

CONCLUSIONS

We identified an endocrinopathy in men with severe male factor infertility that is characterized by a decreased serum testosterone-to-estradiol ratio. This ratio can be corrected by aromatase inhibition, resulting in a significant improvement in semen parameters in oligospermic patients.

Excitatory amino acid neurotransmission is an essential component of the neuroendocrine transmission line that regulates anterior pituitary luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion. Excitatory amino acids (EAAs), such as glutamate and aspartate, are found in large concentrations in presynaptic boutons of a variety of important hypothalamic nuclei, including the arcuate nucleus, the suprachiasmatic nucleus, the supraoptic nucleus, the paraventricular nucleus, and the preoptic area. EAA receptors can be divided into two broad groups, namely, ionotropic and metabotropic receptors. Ionotropic receptors are subdivided into NMDA (N-methyl-D-aspartate), kainate, and AMPA (DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors. Their main mode of action is by the modulation of Na+, K+, and Ca2+ ion channels. Metabotropic receptors, on the other hand, act by a G-protein-stimulated release of intracellular Ca2+ or modulation of adenylate cyclase activity. The different EAA receptor subtypes are found in a variety of areas of the hypothalamus and the brain. In a variety of species, the administration of glutamate, NMDA, or kainate leads to LH release mediated through the stimulation of hypothalamic gonadotropin hormone-releasing hormone (GnRH) release. The major site of NMDA action appears to be the preoptic area--where GnRH cell bodies reside. AMPA and kainate appear to act primarily at the arcuate nucleus/median eminence, the site of GnRH nerve terminals. NMDA may also act upon noradrenergic neurons in the locus coeruleus to influence hypothalamic GnRH release. The steroid-induced LH surge in ovariectomized animals and the preovulatory surge of LH in cycling animals and in pregnant mare's serum gonadotropic-primed animals are blocked by the NMDA antagonist MK801 and the AMPA/kainate antagonist DNQX. MK801 also suppressed FSH surges in most instances, whereas DNQX had no effect on FSH surges. In the ovariectomized female rat, both the NMDA antagonist AP5 and the AMPA/kainate antagonist DNQX, lowered mean LH levels, LH pulse amplitude, and LH pulse frequency. Activation of NMDA receptors advanced the time of vaginal opening in the immature female rat, while kainate and DNQX were without effect. Gonadal steroid removal (castration) did not alter NMDA receptor levels or affinity in the hypothalamus of female or male rats. Likewise, steroid replacement to castrate rats did not affect hypothalamic NMDA receptor levels or NMDA R1 mRNA levels. Similarly, NMDA and kainate receptor levels in the hypothalamus did not change during the time of puberty in the female rat. In contrast, AMPA receptor (GluR1) immunoreactive levels in the magnocellular preoptic area (mPOA), the arcuate nucleus (ARC), and the suprachiasmatic nucleus (SCN) were found to be markedly elevated during the time of the LH surge in estradiol-progesterone-treated castrate rats compared to those of the vehicle-only-treated castrate rat. The release rates of glutamate and aspartate in the POA were found to be significantly elevated during the steroid-induced LH surge in the ovariectomized adult rat.(ABSTRACT TRUNCATED AT 400 WORDS)

OBJECTIVE

To determine whether or not hysterectomy leads to an earlier onset of the menopause.

METHODS

A prospective cohort study.

METHODS

Gynaecology service of large urban hospital.

METHODS

Premenopausal women with and without hysterectomy.

METHODS

Multivariate survival analysis techniques were used to adjust for differences in initial follicle stimulating hormone (FSH) levels, body mass index, smoking and unilateral oophorectomy between the groups.

METHODS

FSH levels were measured for five years following hysterectomy and compared with the comparison group. Menopause was defined as a single FSH measurement of at least 40 IU/L.

RESULTS

Two hundred and fifty-seven women undergoing hysterectomy were compared with 259 women who had not undergone a hysterectomy. Fifty-three women (20.6%) in the hysterectomy group and 19 women (7.3%) in the comparison group reached menopause over the five years of the study. Women in the hysterectomy group with a pre-operative FSH <10 IU/L reached menopause 3.7 years (95% CI 1.5-6.0 years) earlier than women in the comparison group independent of BMI, smoking and unilateral oophorectomy. Twenty-eight women in the hysterectomy group had unilateral oophorectomy and 10 (35.7%) of these women reached menopause over the five years of follow up. Women in the hysterectomy group with unilateral oophorectomy reached menopause 4.4 years (95% CI 0.6, 7.9 years) earlier than women with both ovaries in the hysterectomy group independent of baseline FSH, BMI and smoking.

CONCLUSIONS

Hysterectomy is associated with an earlier onset of menopause. Hysterectomy with unilateral oophorectomy is associated with an even earlier onset of the menopause in this study.