Bisphenol A Deranges the Endocannabinoid System of Primary Sertoli Cells with an Impact on Inhibin B Production.

Bisphenol A (BPA) is an endocrine disruptor that negatively affects spermatogenesis, a process where Sertoli cells play a central role. Thus, in the present study we sought to ascertain whether BPA could modulate the endocannabinoid (eCB) system in exposed mouse primary Sertoli cells. Under our experimental conditions, BPA turned out to be cytotoxic to Sertoli cells with an half-maximal inhibitory concentration (IC50) of ~6.0 µM. Exposure to a non-cytotoxic dose of BPA (i.e., 0.5 µM for 48 h) increased the expression levels of specific components of the eCB system, namely: type-1 cannabinoid (CB1) receptor and diacylglycerol lipase-α (DAGL-α), at mRNA level, type-2 cannabinoid (CB2) receptor, transient receptor potential vanilloid 1 (TRPV1) receptors, and DAGL-ß, at protein level. Interestingly, BPA also increased the production of inhibin B, but not that of transferrin, and blockade of either CB2 receptor or TRPV1 receptor further enhanced the BPA effect. Altogether, our study provides unprecedented evidence that BPA deranges the eCB system of Sertoli cells towards CB2- and TRPV1-dependent signal transduction, both receptors being engaged in modulating BPA effects on inhibin B production. These findings add CB2 and TRPV1 receptors, and hence the eCB signaling, to the other molecular targets of BPA already known in mammalian cells.

Deletion of Spata2 by CRISPR/Cas9n causes increased inhibin alpha expression and attenuated fertility in male mice.

As somatic cells in the testis seminiferous tubule, Sertoli cells provide the medium for spermatogenesis. One of the important functions of Sertoli cells is synthesizing and secreting cell factors to affect the production of sperm; however, much of those molecular regulation mechanisms remain unknown. Here, we confirm the localization of protein SPATA2 (spermatogenesis-associated protein 2), which had previously been shown to be highly expressed in Sertoli cells of the adult mouse testis. To further conduct a functional study, we generated SPATA2 global knockout mice via use of the CRISPR/Cas9n gene editing technology. The 120-day-old knockout mice testes showed almost a 40% decrease in size and weight and variations in the histomorphology of the seminiferous epithelium, with a 40% decrease in sperm count. Further examination revealed that the proliferation of germ cells in the seminiferous tubules was attenuated by 28%. In addition, we found that SPATA2 deletion led to an approximately 70% increase in the inhibin alpha-subunit mRNA and protein level in the testes compared to that of wild-type mice. Our data revealed the impact of SPATA2 on male fertility and suggested that SPATA2 ensures the normal secretory function of Sertoli cells.

Role of FSH glycan structure in the regulation of Sertoli cell inhibin production.

Variations in follicle-stimulating hormone (FSH) carbohydrate composition and structure are associated with important structural and functional changes in Sertoli cells (SCs) during sexual maturation. The aim of the present study was to investigate the impact of FSH oligosaccharide structure and its interaction with gonadal factors on the regulation of monomeric and dimeric inhibin production at different maturation stages of the SC. Recombinant human FSH (rhFSH) glycosylation variants were isolated according to their sialylation degree (AC and BA) and complexity of oligosaccharides (CO and HY). Native rhFSH stimulated inhibin α-subunit (Pro-αC) but did not show any effect on inhibin B (INHB) production in immature SCs isolated from 8-day-old rats. Activin A stimulated INHB and had a synergistic effect on FSH to stimulate Pro-αC. The less acidic/sialylated rhFSH charge analogues, BA, were the only charge analogue mix that stimulated INHB as well as the most potent stimulus for Pro-αC production. Native rhFSH stimulated both Pro-αC and INHB in SCs at a more advanced maturation stage, isolated from 20-day-old rats. In these cells, all rhFSH glycosylation variants increased INHB and Pro-αC production, even in the presence of growth factors. The BA preparation exerted a more marked stimulatory effect on INHB and Pro-αC than the AC. Glycoforms bearing high mannose and hybrid-type oligosaccharides, HY, stimulated INHB and Pro-αC more effectively than those bearing complex oligosaccharides, CO, even in the presence of gonadal growth factors. These findings demonstrate the modulatory effect of FSH oligosaccharide structure on the regulation of inhibin production in the male gonad.

Regulation of spermatogenesis: an evolutionary biologist's perspective.

This review describes the regulation of spermatogenesis taking into consideration the hypothalamic-pituitary gonadal axis, the male reproductive organs and the endocrine and paracrine factors involved in the control of sperm production and the release of androgens. Instead of detailed descriptions of many hormones and growth factors, we attempt to provide an integrative and evolutionary view by comparing different species and considering their specific needs for successful male reproduction. The review focuses on species specific differences in the structural organization of spermatogenesis and indicates that the crucial regulatory mechanisms controlling sperm output are targeted toward differentiating spermatogonia when they initiate clonal expansion. We argue that the further differentiation of germ cells is following a highly coordinated and strictly predetermined morphogenetic cascade widely independent of hormonal control. We propose a hypothetical "ancient" model. Spermatogenesis and steroidogenesis are controlled by a master switch (GnRH pulse generator) under whose control two separate feedback systems provide independent control of androgen (LH-testosterone) and sperm production (FSH-inhibin). This scenario offers high flexibility and has seen uncountable adaptions to optimize the specific needs of different species. Models for the hormonal regulation in hamsters, laboratory rodents and primates are presented to illustrate the species specific diversity.

Germline mutations of inhibins in early-onset ovarian epithelial tumors.

To identify novel genetic bases of early-onset epithelial ovarian tumors, we used the trio exome sequencing strategy in a patient without familial history of cancer who presented metastatic serous ovarian adenocarcinomas at 21 years of age. We identified a single de novo mutation (c.1157A>G/p.Asn386Ser) within the INHBA gene encoding the ßA-subunit of inhibins/activins, which play a key role in ovarian development. In vitro, this mutation alters the ratio of secreted activins and inhibins. In a second patient with early-onset serous borderline papillary cystadenoma, we identified an unreported germline mutation (c.179G>T/p.Arg60Leu) of the INHA gene encoding the α-subunit, the partner of the ßA-subunit. This mutation also alters the secreted activin/inhibin ratio, by disrupting both inhibin A and inhibin B biosynthesis. In a cohort of 62 cases, we detected an additional unreported germline mutation of the INHBA gene (c.839G>A/p.Gly280Glu). Our results strongly suggest that inhibin mutations contribute to the genetic determinism of epithelial ovarian tumors.

Expression of endoplasmic reticulum stress proteins is a candidate marker of brain metastasis in both ErbB-2+ and ErbB-2- primary breast tumors.

The increasing incidence of breast cancer brain metastasis in patients with otherwise well-controlled systemic cancer is a key challenge in cancer research. It is necessary to understand the properties of brain-tropic tumor cells to identify patients at risk for brain metastasis. Here we attempt to identify functional phenotypes that might enhance brain metastasis. To obtain an accurate classification of brain metastasis proteins, we mapped organ-specific brain metastasis gene expression signatures onto an experimental protein-protein interaction network based on brain metastatic cells. Thirty-seven proteins were differentially expressed between brain metastases and non-brain metastases. Analysis of metastatic tissues, the use of bioinformatic approaches, and the characterization of protein expression in tumors with or without metastasis identified candidate markers. A multivariate analysis based on stepwise logistic regression revealed GRP94, FN14, and inhibin as the best combination to discriminate between brain and non-brain metastases (ROC AUC = 0.85, 95% CI = 0.73 to 0.96 for the combination of the three proteins). These markers substantially improve the discrimination of brain metastasis compared with ErbB-2 alone (AUC = 0.76, 95% CI = 0.60 to 0.93). Furthermore, GRP94 was a better negative marker (LR = 0.16) than ErbB-2 (LR = 0.42). We conclude that, in breast carcinomas, certain proteins associated with the endoplasmic reticulum stress phenotype are candidate markers of brain metastasis.

Human testis steroidogenesis is inhibited by phthalates.

BACKGROUND: Phthalic acid esters are widely used in the manufacture of plastics. Numerous studies have shown that these phthalates impair testicular testosterone production in the rat. However, the scarce and contradictory data concerning humans have cast doubt over whether these compounds are also anti-androgenic in man. We therefore investigated the direct effects of di-(2-ethylhexyl) phthalate (DEHP) and mono-(2-ethylhexyl) phthalate (MEHP) on organo-cultured adult human testis and a human cell line. METHODS: Adult human testis explants or NCI-H295R adrenocortical human cells were cultured with DEHP or MEHP. The effects of ketoconazole, used as a reference molecule, were also assessed. RESULTS: In both models, DEHP and MEHP significantly inhibited testosterone production. The effects of both phthalates appeared to be specific for steroidogenesis, as INSL3 production by Leydig cells was not altered. Furthermore, the phthalates of interest had no effect on inhibin B production by Sertoli cells or on germ cell apoptosis. As only a small fraction of the phthalates added was found in the testis explants, and as these compounds were found to be metabolized, we estimate that the anti-androgenic effects observed occurred at concentrations of phthalates that are of the same order of magnitude as exposures reported in the literature for men. CONCLUSIONS: We provide the first evidence that DEHP and MEHP can inhibit testosterone production in the adult human testis. This is consistent with recent epidemiological findings of an inverse correlation between exposure to MEHP and testosterone concentrations.

A mathematical model of the bovine oestrous cycle: simulating outcomes of dietary and pharmacological interventions.

A mathematical model was constructed to simulate the bovine oestrous cycle by using nonlinear differential equations to describe the biological mechanisms which regulate the cycle. The model predicts circulating concentrations of gonadotrophin-releasing hormone, follicle-stimulating hormone, luteinizing hormone, oestradiol, inhibin and progesterone. These hormones collectively provide control and feedback mechanisms between the hypothalamus, pituitary gland and ovaries, which regulate ovarian follicular dynamics, corpus luteum function and ovulation. When follicular growth parameters are altered, the model predicts that cows will exhibit either two or three follicular waves per cycle, as seen in practice. Changes in other parameters allow the model to simulate: effects of nutrition on follicle recruitment and size of the ovulatory follicle; effects of negative energy balance on postpartum anoestrus; and effects of pharmacological intervention on hormone profiles and timing of ovulation. It is concluded that this model provides a sound basis for exploring factors that influence the bovine oestrous cycle in order to test hypotheses about nutritional and hormonal influences which, with further validation, should help to design dietary or pharmacological strategies for improving reproductive performance in cattle.

Human INHBB Gene Variant (c.1079T>C:p.Met360Thr) Alters Testis Germ Cell Content, but Does Not Impact Fertility in Mice.

Testicular-derived inhibin B (α/ß B dimers) acts in an endocrine manner to suppress pituitary production of follicle-stimulating hormone (FSH), by blocking the actions of activins (ß A/B/ß A/B dimers). Previously, we identified a homozygous genetic variant (c.1079T>C:p.Met360Thr) arising from uniparental disomy of chromosome 2 in the INHBB gene (ß B-subunit of inhibin B and activin B) in a man suffering from infertility (azoospermia). In this study, we aimed to test the causality of the p.Met360Thr variant in INHBB and testis function. Here, we used CRISPR/Cas9 technology to generate InhbbM364T/M364T mice, where mouse INHBB p.Met364 corresponds with human p.Met360. Surprisingly, we found that the testes of male InhbbM364T/M364T mutant mice were significantly larger compared with those of aged-matched wildtype littermates at 12 and 24 weeks of age. This was attributed to a significant increase in Sertoli cell and round spermatid number and, consequently, seminiferous tubule area in InhbbM364T/M364T males compared to wildtype males. Despite this testis phenotype, male InhbbM364T/M364T mutant mice retained normal fertility. Serum hormone analyses, however, indicated that the InhbbM364T variant resulted in reduced circulating levels of activin B but did not affect FSH production. We also examined the effect of this p.Met360Thr and an additional INHBB variant (c.314C>T: p.Thr105Met) found in another infertile man on inhibin B and activin B in vitro biosynthesis. We found that both INHBB variants resulted in a significant disruption to activin B in vitro biosynthesis. Together, this analysis supports that INHBB variants that limit activin B production have consequences for testis composition in males.

Involvement of activin signaling in abnormalities of mouse vagina exposed neonatally to diethylstilbestrol.

Perinatal exposure to a synthetic estrogen, diethylstilbestrol (DES), causes cervicovaginal adenosis and permanent hyperplastic cornified vaginal epithelium with keratinization in mice. To investigate the mechanisms of the induction of vaginal abnormalities by DES, we have focused on activin A signaling. We have found that the ßA-subunit mRNA is mainly expressed in the neonatal vaginal stroma, whereas activin A receptor type IB is localized in the neonatal vaginal epithelium. SMAD2, the intracellular signaling protein, is phosphorylated in the neonatal vagina. Cell proliferation in the vaginal epithelium grown in vitro is reduced by DES treatment or by activin signaling suppression through inhibin treatment. Thus, activin A (a homodimer of the ßA-subunit) in the stroma stimulates epithelial cell proliferation in the neonatal vagina. DES treatment decreases the expression of the ßA-subunit and activin receptor IIB but increases the expression of the ßB-subunit and inhibin receptor. Neonatal DES treatment inhibits the phosphorylation of SMAD2 in the vaginal epithelium, indicating the inhibition of activin A signaling in the vaginal epithelium by neonatal DES treatment. Treatment with DES or inhibin, a native antagonist of activin, induces adenosis-like structures and keratinization in the vagina grown in vitro. These data suggest that the suppression of activin A signaling by DES is involved in the induction of cervicovaginal adenosis and keratinization in the neonatal mouse vaginal epithelium.

Induction of activin A is essential for the neuroprotective action of basic fibroblast growth factor in vivo.

Exogenous application of neurotrophic growth factors has emerged as a new and particularly promising approach not only to promote functional recovery after acute brain injury but also to protect neurons against the immediate effect of the injury. Among the various growth factors and cytokines studied so far, the neuroprotective and neurotrophic profile of basic fibroblast growth factor (bFGF) is the best documented. Using an animal model of acute excitotoxic brain injury, we report here that the neuroprotective action of bFGF, which is now being tested in stroke patients, depends on the induction of activin A, a member of the transforming growth factor-beta superfamily. Our evidence for this previously unknown mechanism of action of bFGF is that bFGF strongly enhanced lesion-associated induction of activin A; in the presence of the activin-neutralizing protein follistatin, bFGF was no longer capable of rescuing neurons from excitotoxic death; and recombinant activin A exerted a neuroprotective effect by itself. Our data indicate that the development of substances influencing activin expression or receptor binding should offer new ways to fight neuronal loss in ischemic and traumatic brain injury.

Inhibin-positive hepatic carcinoma: proposal for a solid-tubulocystic variant of intrahepatic cholangiocarcinoma.

Inhibin-positive hepatic carcinoma is a rare primary liver neoplasm that resembles sex cord-stromal tumor and thyroid follicular tumors. The term "cholangioblastic variant of intrahepatic cholangiocarcinoma" has been proposed. This study describes the clinicopathologic, immunophenotypic, and molecular features of a small series (n = 6) of this rare tumor. Albumin in situ hybridization (ISH) and capture-based next-generation sequencing (NGS) were also performed. All tumors occurred in young women (mean age 32.5 years, range 19-44 years) as a solitary large mass (mean 15.8 cm, range 6.9-23.5 cm). All tumors showed a highly distinctive morphology with sheets and large nests of tumor cells alternating with tubular and cystic areas imparting a sex cord-like or thyroid follicle-like morphology. Cytologic atypia was mild, and mitotic activity was low. All cases were positive for inhibin, as well as pancytokeratin, CK7, CK19, and albumin ISH. Synaptophysin and chromogranin showed focal or patchy staining, whereas INSM1 was negative. Markers for hepatocellular differentiation, thyroid origin, and sex cord-stromal tumor were negative. There were no recurrent genomic changes based on capture-based NGS of ∼500 cancer genes. Recurrence and/or metastasis was seen in three (50%) cases (follow-up time range for all cases: 5 months to 2 years). In conclusion, this series describes the distinctive morphology, immunophenotypic features, and diffuse albumin staining in six cases of a rare inhibin-positive primary liver carcinoma that runs an aggressive course similar to intrahepatic cholangiocarcinoma. Genomic changes typical of cholangiocarcinoma or hepatocellular carcinoma were not identified, and there were no recurrent genetic abnormalities. We propose the term "solid-tubulocystic variant of intrahepatic cholangiocarcinoma" to reflect the spectrum of morphologic patterns observed in this tumor.

Activin A/erythroid differentiation factor is induced during human monocyte activation.

Activin A/erythroid differentiation factor (EDF), a dimeric polypeptide hormone composed of two beta A subunits, regulates growth and erythroid differentiation of human hematopoietic progenitor and erythroleukemia cells. We have identified activated human peripheral blood monocytes as a natural source of activin A/EDF. In these cells, lipopolysaccharide (LPS) induced rapidly the expression of the beta A subunit mRNAs through protein kinase C-dependent transcriptional regulation. The beta A subunit mRNA expression was also increased by 1,25-dihydroxyvitamin D3, an inducer of macrophage maturation of monocytes. Western analysis with an anti-beta A antibody and an erythroid differentiation bioassay confirmed that the conditioned media of LPS-activated monocytes contained the activin A/EDF protein. We suggest that monocyte/macrophage-derived activin A/EDF may not only modulate hematopoiesis but may also act as a mediator molecule in the diverse physiologic and pathogenetic events in which these cells are involved.

Spinal Versus Intracranial Meningioma: Aberrant Expression of CD10 and Inhibin with Relation to Clinicopathological Features and Prognosis.

CD10 and inhibin are used mainly in CNS pathology to distinguish hemangioblastoma from metastatic clear cell renal cell carcinoma. Some meningiomas can mimic both tumors and so we aimed at this study to investigate the expression of both markers in a large number of meningioma cases. One hundred thirty-four meningioma samples were collected, 14 of them were spinal and 120 were intracranial. Manual TMA blocks were constructed using modified mechanical pencil tip method and immunohistochemistry for CD10 and inhibin was done. Intracranial meningioma occurred in significantly younger age than spinal ones. Most of spinal meningiomas were of transitional histology. CD10 was expressed in 14% of cases with significant positivity in spinal rather than intracranial cases. Transitional meningiomas showed the highest positivity for CD10 expression, while the least positive was the meningiotheliomatous type. Inhibin was expressed in 6% of cases with no significant relation to clinicopathological and histological features. There was no significant relationship between the expression of CD10 and inhibin expression in meningiomas. In conclusion, spinal meningiomas differ than intracranial ones in many clinicopathological and biological aspects. Among these differences is CD10 expression being more expressed in spinal meningiomas. However CD10 and inhibin are aberrantly expressed in a proportion of meningiomas, both have no relations to poor prognostic factors but more caution should be exerted during usage of these markers in diagnosis of hemangioblastoma and metastatic RCC. Further studies are suggested for exploring more biological differences between spinal and intracranial meningiomas.

Associations between andrological measures, hormones and semen quality in fertile Australian men: inverse relationship between obesity and sperm output.

BACKGROUND: The World Health Organization developed a time to pregnancy (TTP) study (number of menstrual cycles taken to conceive) to determine whether the average TTP is increasing and semen quality decreasing with time. The present study describes clinical, semen and hormone characteristics obtained from male partners of pregnant women in Melbourne, Australia, and examines the associations between these characteristics. METHODS: Male partners (n = 225) of pregnant women (16-32 weeks) who conceived naturally had physical examination, health and lifestyle questionnaires, semen and hormone (FSH, LH, sex hormone-binding globulin, testosterone and Inhibin B) analyses. RESULTS: Previously known associations between semen, hormone and clinical variables were confirmed as significant: sperm numbers (concentration and total sperm count) correlated positively with Inhibin B and inversely with FSH and left varicocele, while total testicular volume correlated positively with sperm numbers and Inhibin B and inversely with FSH. However, only abstinence, total testicular volume, varicocele grade and obesity (BMI > 30 kg/m2) were independently significantly related to total sperm count. Compared with those with BMI < 30 (n = 188), obese subjects (n = 35) had significantly lower total sperm count (mean 324 versus 231 million, P = 0.013) and Inhibin B (187 versus 140 pg/ml, P < 0.001) but not FSH (3.4 versus 4.0 IU/l, P = 0.6). CONCLUSIONS: Obese fertile men appear to have reduced testicular function. Whether this is cause or effect, i.e. adiposity impairing spermatogenesis or reduced testicular function promoting fat deposition, remains to be determined.

Requirement for activin A and transforming growth factor--beta 1 pro-regions in homodimer assembly.

Many proteins are initially synthesized as part of a large precursor. The role of the pro-region in the biosynthesis of transforming growth factor--beta 1 (TGF-beta 1) and activin A, two structurally related disulfide-linked homodimers synthesized as large precursors, was studied. Vectors that expressed either the pro-region or the mature regions of these molecules were used in complementation experiments, only when the pro-region was coexpressed with the mature region did intracellular dimerization and secretion of biologically active homodimers occur. The pro-regions of activin A and TGF-beta 1, therefore, aid the folding, disulfide bond formation, and export of their respective homodimers.

Ovarian involvement by desmoplastic small round cell tumor with leydig cell hyperplasia showing an unusual immunophenotype (cytokeratin negative, calretinin and inhibin positive) mimicking poorly differentiated sertoli leydig cell tumor.

Desmoplastic small round cell tumor (DSRCT) is a rare aggressive tumor primarily involving serosal surfaces in adolescents and young men. Diagnosis is based on specific clinicomorphologic, immunohistochemical, and genetic features. We report here a variant of DSRCT involving the ovaries that mimics the Sertoli-Leydig cell tumor in a 21-year-old woman complaining of abdominal pain. Abdominal ultrasonography and computed tomography showed a right adnexal mass. She had a slightly raised serum CA-125 level. Frozen section examination identified the right ovarian mass as a poorly differentiated Sertoli-Leydig cell tumor. The surgically resected tumor and left ovary and omentum implants found during laparoscopy were diagnosed as DSRCT with Leydig cell hyperplasia. Immunohistochemically, the tumor cells were negative for epithelial markers but were positive for calretinin and inhibin. The patient is still undergoing chemotherapy at 8 months after initial presentation with partial response. This case showed that DSRCT with unusual immunohistochemical profiles and Leydig cells hyperplasia pose a diagnostic challenge. Molecular genetic techniques may help in these cases.

Cytokeratin 7, inhibin, and p63 in testicular germ cell tumor: superior markers of choriocarcinoma compared to ß-human chorionic gonadotropin.

Choriocarcinoma can be difficult to differentiate from other subtypes of testicular germ cell tumor and can occur unexpectedly in a distant, late metastasis. The aim of this investigation was to identify a marker superior to ß-human chorionic gonadotropin (ß-hCG) for choriocarcinoma. Sixty-two primary and metastatic testicular germ cell tumors (27 choriocarcinomas, 19 yolk sac tumors, 29 embryonal carcinomas, 28 seminomas, 22 teratomas, 3 epithelioid trophoblastic tumors [ETTs]) were analyzed for immunohistochemical expression of cytokeratin 7 (CK7), inhibin, p63, and ß-hCG. All choriocarcinomas and ETTs were strongly positive for CK7, whereas seminomas were negative and 52% of embryonal carcinomas had weak reactivity. Eighty-four percent of yolk sac tumors and 59% of teratomas were CK7 positive. Eighty-nine percent of choriocarcinomas and 100% of ETTs were positive for inhibin, with reactivity highlighting syncytiotrophoblasts, whereas seminomas, embryonal carcinomas, yolk sac tumors, and teratomas were negative. Eighty-five percent of choriocarcinomas expressed p63, with staining mostly in mononucleated trophoblasts, whereas seminomas, embryonal carcinomas, and yolk sac tumors were negative. Teratomas expressed p63 in 32% of cases. ß-hCG was reactive in 96% of choriocarcinomas, 33% of ETTs, 46% of seminomas, 54% of embryonal carcinomas, 47% of yolk sac tumors, and 32% of teratomas. ß-hCG staining within other subtypes was more likely if choriocarcinoma was present elsewhere in the tumor (P = .0002). CK7 is a highly sensitive marker for choriocarcinoma and differentiates choriocarcinoma from seminoma and embryonal carcinoma. Inhibin and p63 are sensitive and specific for choriocarcinoma versus seminoma, embryonal carcinoma, and yolk sac tumor. To identify choriocarcinoma, CK7, inhibin, and p63 are superior to ß-hCG.

Inhibin, activin, follistatin and FSH serum levels and testicular production are highly modulated during the first spermatogenic wave in mice.

Testicular development is governed by the combined influence of hormones and proteins, including FSH, inhibins, activins and follistatin (FST). This study documents the expression of these proteins and their corresponding mRNAs, in testes and serum from mice aged 0 through 91 days post partum (dpp), using real-time PCR, in situ hybridisation, immunohistochemistry, ELISA and RIA. Serum immunoactive total inhibin and FSH levels were negatively correlated during development, with FSH levels rising and inhibin levels falling. Activin A production changed significantly during development, with subunit mRNA and protein levels declining rapidly after 4 dpp, while simultaneously levels of the activin antagonists, FST and inhibin/activin beta(C), increased. Inhibin/activin beta(A) and beta(B) subunit mRNAs were detected in Sertoli, germ and Leydig cells throughout testis development, with the beta(A) subunit also detected in peritubular myoid cells. The alpha, beta(A), beta(B) and beta(C) subunit proteins were detected in Sertoli and Leydig cells of developing and adult mouse testes. While beta(A) and beta(B) subunit proteins were observed in spermatogonia and spermatocytes in immature testes, beta(C) was localised to leptotene and zygotene spermatocytes in immature and adult testes. Nuclear beta(A) subunit protein was observed in primary spermatocytes and nuclear beta(C) subunit in gonocytes and round spermatids. The changing spatial and temporal distributions of inhibins and activins indicate that their modulated synthesis and action are important during onset of murine spermatogenesis. This study provides a foundation for evaluation of these proteins in mice with disturbed testicular development, enabling their role in normal and perturbed spermatogenesis to be more fully understood.

Inhibin A: marker for diagnosis of ectopic and early abnormal pregnancies.

A prospective case-control study was performed to determine whether inhibin A concentration is a clinically useful marker of ectopic pregnancy (EP). Inhibin A concentration in patients diagnosed with EP by laparoscopic and pathological findings (n = 17) was compared with that in missed miscarriage (n = 35), incomplete miscarriage (n = 14), spontaneous miscarriage (n = 5), threatened miscarriage (n = 6), normal pregnancy (n = 24) and non-pregnant controls (n = 20). The data were analysed using the Mann-Whitney U-test. EP yielded significantly lower inhibin A concentrations compared with normal pregnancy, 12.7 +/- 11.7 versus 237.3 +/- 125.9 pg/ml (P < 0.0002), and similar concentrations to non-pregnant controls (13.3 +/- 14.3 pg/ml). Inhibin A concentrations in abnormal pregnancies were significantly lower than in the normal pregnancy group: missed miscarriage 42.4 +/- 54.9 pg/ml (P < 0.0002); spontaneous miscarriage 47.5 +/- 55.6 pg/ml (P < 0.0002); and incomplete miscarriage 12.2 +/- 10.5 pg/ml (P < 0.0002). Threatened miscarriage was not statistically different to normal pregnancy (183.1 +/- 119.4 pg/ml). Human chorionic gonadotrophin concentrations in EP were not statistically significantly different compared with missed miscarriage and incomplete miscarriage. In conclusion, serum inhibin A concentration may be a reliable marker of EP.