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Direct luteinizing hormone action triggers adrenocortical tumorigenesis in castrated mice transgenic for the murine inhibin alpha-subunit promoter/simian virus 40 T-antigen fusion gene.

Transgenic (TG) mice, expressing the Simian Virus 40 T-antigen (Tag) under a 6-kb fragment of the murine inhibin alpha-subunit promoter (inh alpha p), develop gonadal tumors of granulosa/theca or Leydig cell origin. We showed previously that adrenocortical tumors develop if the TG mice are gonadectomized but never develop in intact animals. However, if functional gonadectomy was induced by GnRH antagonist treatment or by cross-breeding the TG mice into the hypogonadotropic hpg genetic background, neither gonadal nor adrenal tumors appeared. Since the most obvious difference between the gonadectomized and GnRH-antagonist-treated or Tag/hpg double mutant mice is the elevated gonadotropin secretion in the first group, we examined whether the adrenal tumorigenesis would be gonadotropin-dependent. Surprisingly, both the adrenal tumors and a cell line (C alpha 1) derived from one of them expressed highly functional LH receptors (LHR), as assessed by Northern hybridization, immunocytochemistry, ligand binding, and human CG (hCG)-stimulated cAMP and steroid production. No FSH receptor expression was found in the adrenal tumors by RT-PCR. hCG treatment of the C alpha 1 cells stimulated their proliferation, as measured by [3H]thymidine incorporation. This effect was related to hCG-stimulated steroidogenesis since progesterone, testosterone, and estradiol, at physiological concentrations, also stimulated the C alpha 1 cell proliferation. Different adrenocortical cells expressed initially LHR and Tag, whereas both were highly expressed in the tumor cells. In conclusion, the high level of functional LHR in the adrenal tumors indicates that this receptor can function as tumor promoter when ectopically expressed and stimulated by the ligand hormone.

Adrenal Cortex Neoplasms↗

[Pure gonadal dysgenesis. Case report with unusual anatomical and endocrine findings].

The syndrome of pure gonadal dysgenesis (PGD) cannot always easily be distinguished from other disorders of gonadal development. Relations are evident with Turner's syndrome, females with hypoplastic ovaries, male pseudohermaphroditism, mixed gonadal dysgenesis and the vanishing testes syndrome. The case is reported of a 40 year old female with primary amenorrhea, alopecia, eunuchoid features, XY karyotype with normal breast development and sexual hair after estrogen therapy. On laparotomy streak ovaries were found at ovarian site. Pathohistological examination revealed on the left side wolffian duct remnants such as ductuli deferentes and epididymis besides sparse Leydig-(hilus-)cells and on the right side only a rudimentary fallopian tube with subendothelial accumulation of hyperplastic Leydig-(hilus-)cells. Serum-testosterone elevation above the normal female range (630 ng/dl) persisted following gonadectomy (151 ng/dl). Ectopic Leydig-(hilus-)cells were regarded responsible for the continuing testosterone production. The present case lies on borderline between PGD and mixed gonadal dysgenesis because remnants of wolffian duct derivatives suggest unilateral fetal testicular activity; classification as PGD however was justified in purely female body features and lacking evidence of testicular tissue.

Adult↗

Molecular cloning of a molluscan gonadotropin-releasing hormone receptor orthologue specifically expressed in the gonad.

Despite their economic importance, only very little information is available regarding (neuro)endocrine mechanisms of reproduction in bivalve molluscs. To gain insights into the molecular control of gonadic development of these animals, G protein-coupled receptors (GPCR) expressed in the gonad of the pacific oyster Crassostrea gigas were investigated. One such receptor was cloned by RT-PCR using oligonucleotide primers derived from consensus sequences of various vertebrate (neuro)peptide receptors. This receptor named Cg-GnRH-related receptor (Cg GnRH-R) exhibits a high degree of amino acid sequence identity with both vertebrate GnRH receptors and insect AKH receptors. Quantitative RT-PCR shows a specific expression of Cg-GnRH-R in both male and female gonads during the reproductive cycle. This demonstrates for the first time the plausible involvement of a GnRH receptor orthologue in the control of reproduction in a protostomian invertebrate.

5' Untranslated Regions↗

Enhancing spawning in the grey mullet (Mugil cephalus) by removal of dopaminergic inhibition.

A dot-blot immunoassay for the detection of vitellogenin (Vtg) in plasma of adult grey mullet (Mugil cephalus) was developed. The assay identified the sex of the tested fish prior to detectable gonadal development, enabling the establishment of broodstock at the desired ratio of 7:4 females to males. This broodstock was maintained under natural photoperiod, and used to study the relative effect of gonadotropin-releasing hormone (GnRH) and dopamine antagonists on oocyte maturation and ovulation, as well as the effect of 17alpha-methyltestosterone (MT) on spermiation. Three groups of females were treated with: (i) a single injection of dopamine antagonist, domperidone (Dom), (ii) GnRH analog (GnRHa) administered via ethylene-vinyl acetate copolymer (EVAc) slow-release implants or (iii) a combination of both Dom and GnRHa. Males were treated with MT, administered via EVAc slow-release implants. An additional group of untreated fish was used as a control. The Dom treatment proved to be more potent than the GnRHa treatment, and did not differ significantly from the combined treatment. The Dom and Dom+GnRHa treatments accelerated oocyte development and increased plasma estradiol levels equally, whereas the GnRHa treatment did not vary significantly from the control. MT was found to be a potent spermiating agent, which enhanced steady milt production in all treated males. In contrast, no spontaneous spermiation occurred in untreated males. Plasma 11-ketotestosterone (11-KT) levels were significantly higher in MT-treated males than in the controls. Interestingly, MT-treated males held with the GnRHa+Dom-treated females showed higher levels of plasma 11-KT than those held with GnRHa-treated females, indicating an additive effect which is probably attributable to female pheromones. Fully mature females were induced to spawn by injecting GnRHa alone or coupled with metaclopramide (a dopamine D2 receptors antagonist). The combined treatment, which included a dopamine antagonist, was found to be more potent in inducing ovulation and spawning as compared to GnRHa alone. In conclusion, our data suggest that dopaminergic inhibition is a major barrier along the reproductive axis that arrests spontaneous spawning in captive mullets.

Animals↗

A proposed multigeneration protocol for Japanese medaka (Oryzias latipes) to evaluate effects of endocrine disruptors.

Definitive data on reproductive impairment of chronically exposed populations may be required to assess the appropriateness of the existing test methods for hazard identification and prioritization of endocrine modulators. Multigeneration toxicity testing protocols for wildlife receptors are lacking. To help address this gap we describe a multigeneration fish assay using the freshwater fish, Japanese medake (Oryzias latipes). This test species has been used for the evaluation of carcinogenic, teratogenic and reproductive effects and is sensitive to estrogen exposure producing ovo-testis, altered biochemical parameters and phenotypic characteristics. Due to the short life cycle, a multigeneration test with medaka can be conducted in 1 year. Endpoints evaluated include: survival, growth, sex ratio, fecundity, embryonic lesion occurrence, embryonic stage development, gonadal and hepatic somatic indices, histopathology and biochemical parameters. As new endpoints are developed they can be incorporated into the protocol. Results of a positive control (17 beta-estradiol) study are presented to give an indication of the baseline associated with various test endpoints and to highlight the importance of nutrition in the experimental design. 17 beta-Estradiol treatment induced vitellogenin production in male and female medaka, feminized males, and disrupted egg production. The proposed protocol provides researchers with an effective multigeneration fish test that can be used to examine potential effects of stressors at the population, individual, cellular and subcellular level.

Animals↗

Effect of pinealectomy on testosterone, estradiol-17 beta, esterone, and 17 alpha-hydroxyprogesterone levels during the annual reproductive cycle in the freshwater catfish, Clarias batrachus.

The present study examines the relationship of the pineal gland to gonadal activity during the annual reproductive cycle in the freshwater catfish Clarias batrachus. The hormonal profiles of testosterone, estradiol-17 beta, estrone, and 17 alpha-hydroxyprogesterone along with the gonadosomatic index (GSI) were monitored to assess the effect of pinealectomy. Pinealectomy appeared to accelerate the gonadal development from January to May. Removal of pineal prior to the start of gonadal recrudescence (January and February), induced initiation of ovarian recrudescence earlier than scheduled in nature. On the other hand pinealectomy during the vitellogenic phase (May to June) reduced the gonadal activity, suggesting the stimulatory role of pineal on gonads. During the remaining period of July to December (postvitellogenic and early resting phase) pinealectomy had no effect on gonadal activity. Thus, in this species the pineal gland during the annual reproductive cycle appears to have either inhibitory, stimulatory, or no effect on gonadal physiology depending upon the sexual conditions.

17-alpha-Hydroxyprogesterone↗

Induction and maintenance of gonadotropin and lactogen receptors in hypoprolactinemic rats.

The effect of 2-bromo-alpha-ergocriptine (BR)-induced hypoprolactinemia on the induction and maintenance of testicular gonadotropin and lactogen receptors was studied in 60-day-old rats after receptor regulation by a gonadotropin-releasing hormone agonist analog [[D-Ser-(tBu)6]des-Gly10-GnRH N-ethylamide (GnRH-A)] and in young animals during sexual maturation. In adult animals, BR treatment delayed the reappearance of LH binding in the testis after GnRH-A injection, but had no effect on the recovery of lactogen binding. BR treatment also inhibited the increase of LH binding that occurred in control animals during the experimental period, but did not affect lactogen binding. Furthermore, BR potentiated the effect of GnRH-A on the decrease of Leydig cell testosterone synthesis observed 2 days later in vitro. Eight days after GnRH-A injection, concomitant BR treatment significantly inhibited the recovery of Leydig cell cAMP production. In peripubertal (25- to 46-day-old) animals, BR diminished the normal rise in testicular LH receptors, but did not affect the increase in lactogen receptors. Serum testosterone levels and other features of pubertal development, such as balano-preputial separation and spermatogenesis, were unaffected by hypoprolactinemia. In neonatal female animals, significant lactogen binding was detected at 3 days of age, whereas hCG binding was not demonstrable until 9 days after birth. These findings indicate that the expression of lactogen receptors precedes that of LH receptors in the developing gonad, and that the increase of LH binding in the testis during pubertal development requires normal circulating PRL levels. In adult animals, hypoprolactinemia potentiates GnRH-A-induced desensitization of steroidogenesis and cAMP formation, as well as LH receptor down-regulation, delaying the recovery of these phenomena. Although decreased serum PRL levels were associated with a marked reduction in testicular LH receptors both during development and in adult life, the absence of changes in lactogen receptors indicates that the latter sites are largely independent of the circulating PRL concentration.

Animals↗

Human testicular development and the role of the mesonephros in the origin of a dual Sertoli cell system.

Some aspects of the development of the human testis (and overy) are discussed and the main theories regarding gonadal differentiation summarized. The major part of this review deals with the origin and differentiation of the three groups of somatic cellular content: Sertoli cells, Leydig cells and peritubular cells. The most important role of the mesonephros in gonadal development is described. Under the influence of the mesonephros, a second type of meiosis-inducing Sertoli cell differentiates and becomes the opponent of a meiosis-preventing type of Sertoli cell which derives from the coelomic epithelium. All somatic cells are pooled in the central gonadal blastema which is part of the medulla. They migrate via the rete blastema to the sites of their final differentiation. Included are the precursors of the Leydig cells and the peritubular cells.

Cell Differentiation↗

Toxic substances and reproductive disorders in Baltic fish and crustaceans.

In the Baltic Sea ecosystem reproductive disorders have occurred in top consumers such as seals and some fish-eating birds, due to biomagnification of toxic substances, e.g. DDT and PCBs. Reproductive disturbances have also affected fish during the last 25 years. However, there is no strong evidence that toxic substances have caused these problems. Rather, the disorders seem to result from a combination of two or more biotic or abiotic factors. The M74 syndrome, which kills fry of salmon and sea trout, is characterized by a deficiency of thiamine (vitamin B1). Several factors may contribute to the thiamine deficiency, including the diet of salmon in the sea and halogenated organic compounds. Cod do not develop M74, and poor cod recruitment is mainly due to poor oxygen conditions in the spawning areas in combination with overfishing. Toxic compounds in pulp-mill effluents retard gonadal development in perch, but the mechanisms and the active substances have not been identified. Recruitment problems in perch in the coastal waters outside some pulp mills may also be explained by a lack of food items for juvenile fish, rather than reproductive failure. There are very limited data on reproductive disorders in crustaceans from the Baltic Sea. Most data come from studies of the benthic amphipod Monoporeia affinis, which has been used in monitoring programs. Several signs of reproductive disorder have been reported in this amphipod, e.g. malformation and death of embryos, and asynchronous maturation of males and females.

Animals↗

The formation of the gonadal ridge in Xenopus laevis. II. A scanning electron microscope study.

This paper studies the surface morphology of the developing gonadal ridge in X. laevis between stages 44 and 49 (Nieuwkoop & Faber, 1956). During this period the primordial germ cells (PGCs) move laterally from the dorsal mesentery of the gut to the position of the presumptive gonadal ridge. As they do so the coelomic lining cells lateral to the mesentery differentiate into a specialized, longitudinally orientated band, stretching nearly the full length of the dorsal mesentery on each side. The PGCs migrate beneath this band of cells, which thus becomes the germinal epithelium of the gonadal ridge. We have demonstrated by irradiation experiments that this specialized band of cells can differentiate independently, in the absence of the PGCs.

Animals↗

Multiple alternative splicing of mouse Dmrt1 during gonadal differentiation.

Evolutionarily conserved Dmrt1 encodes a transcriptional regulator that is expressed exclusively in the gonads and is required for testis differentiation. Here we report that four transcripts of the mouse Dmrt1 were generated in developing gonads and adult testis by alternative splicing. Dmrt1 a encodes the known protein with 374 amino acids. Dmrt1 b, Dmrt1 c, and Dmrt1 d encode predicted proteins with 212, 257, and 194 amino acids, respectively. Dmrt1 a2 and Dmrt1 a3 have anterior alternative polyadenylation signals in 3'UTR than the known Dmrt1 transcript (Dmrt1 a1). Dmrt1 a2 lacks 18 nucleic acids in 3'UTR. Interestingly, Dmrt1 b lacks exon5, Dmrt1 c lacks exon3, and Dmrt1 d lacks both exon1 and exon2 which encode DM domain. RT-PCR showed that all of Dmrt1 transcripts were only detectable in adult testis. However, during gonadogenesis, the multiple alternatively spliced transcripts all had gonad-specific and sexually dimorphic expression profiles. Northern blot and real time fluorescent quantitative RT-PCR further indicated that the expression of Dmrt1 a was dominantly higher than those of Dmrt1 b, c, and d, although they showed a similar up and down pattern of expression during embryo development. The expression of Dmrt1 transcripts climbed up to a climax at 13.5dpc in both male and female gonad. Afterwards, expression in male gonad decreased to a low level and maintained, whereas female one reduced rapidly until undetectable in adult ovary. These results provide new insight into roles of regulation at level of splicing of the Dmrt1 in governing sex differentiation.

Alternative Splicing↗

The use of cytogenetic techniques for the diagnosis of endocrine diseases.

Chromosome analysis is the genetic technique useful in evaluating endocrine disease. Indications for cytogenetic studies include suspected sex chromosomal aneuploidies as well as repeated fetal wastage. In patients with ambiguous genitalia, cytogenetic studies are mandatory, not only to provide information for assigning sex but, more importantly, for identifying individuals who are at risk for developing gonadal tumors because of the combination of a Y chromosome and dysgenetic gonads.

Abortion, Spontaneous↗

Germ cell migration and early development of the gonads in the trisomy 16 mouse--an animal model for Down's syndrome.

The aneuploid condition of patients with Down's syndrome (trisomy 21) frequently leads to a sub- or infertility of these individuals. Gonads from adults and fetuses with trisomy 21 demonstrated histologically a remarkable reduction in germ cells. Disorders in the germ cell migration, the early development of the gonads as well as meiotic defects are thought to contribute to this pathomorphology. To gain information about premeiotic defects, investigations on the trisomy 16 mouse, an animal model for Down's syndrome, were carried out. By means of morphometric studies a delay in migration and a reduction in primordial germ cells was evaluated in trisomic mice of embryonic day 11 (E11). At day E13 a generalized growth retardation of the developing gonads was obvious in trisomic animals. Additionally performed electron microscopic examinations revealed signs of germ cell demise in trisomy 16 mice. Thus, the mechanisms of a diminished proliferation capacity, impaired migration and premature death of germ cells represent premeiotic disorders that presumably contribute to the pathomorphology observed in the gonads of individuals with Down's syndrome.

Alkaline Phosphatase↗

Identification and comparative expression analysis of a second wt1 gene in zebrafish.

The Wilms' tumor suppressor gene wt1 encodes a zinc-finger transcription factor that plays an important role in the development of the mammalian genitourinary system. Mutations in WT1 in humans lead to anomalies of kidney and gonad development and cause Wilms' tumor, a pediatric kidney cancer. The inactivation of both wt1 alleles in mice gives rise to multiple organ defects, among them agenesis of kidney, spleen, and gonads. In zebrafish, an ortholog of wt1 has been described that is expressed in the pronephric field and is later restricted to the podocytes. Here, we report the existence of a second wt1 gene in zebrafish, which we have named wt1b (we named the initial gene wt1a). The overall sequence identity of the two Wt1 proteins is 70% and 92% between the zinc-finger regions, respectively. In contrast to wt1a, wt1b is expressed from the earliest stages of development onward, albeit at low levels. Both wt1a and wt1b are expressed in the intermediate mesoderm, with wt1b being restricted to a smaller area lying at the caudal end of the wt1a expression domain. In adult fish, high expression levels for both genes can be found in gonads, kidney, heart, spleen, and muscle.

Amino Acid Sequence↗

Hypoxia alters testis development in neonatal rats.

OBJECTIVE: To investigate the effects of continued hypoxia on postnatal development of the rat testis. DESIGN: Wistar rats were exposed to simulated hypoxia from birth to postnatal day (PND) 45. Testosterone (T) in the plasma and the testis was measured in rats at PND 3, 7, 14, 21, 28, 35 and 45 respectively. Testis histology and cellular ultrastructure were studied. RESULTS: a) Hypoxia induced a significant arrest of testes weight gain after PND 28; b) T release was significantly attenuated after PND 21; c) Alterations in histology and cellular ultrastructure were found in the testis, showing the swelling of testis interstitium and the enlargement of mitochondria in Leydig cells. CONCLUSION: Postnatal hypoxia stress alters testis development both in terms of function and structure, especially at the critical age of gonadal development.

Altitude↗

Noninvasive characterization of gonad maturation and determination of the sex of Pacific oysters by MRI.

The aim of this study was to test the ability of magnetic resonance imaging (MRI) technique to characterize gonad development and to determine the sex of live Pacific oysters through their shells. A preliminary nuclear magnetic resonance (NMR) relaxometry study was conducted to characterize T1 and T2 NMR relaxation parameters for the main oyster organs. This showed that T1-weighted MRI sequences were most appropriate to optimize contrasts between tissues in images. The results showed that gray levels of gonads in images acquired with gradient-echo sequence were variably affected by T2* weighting effect. However, the ovaries systematically gave a hypersignal in spin-echo T1-weighted images, and stack histograms of female oysters showed a peak well separated from that of male oysters. An automated method is proposed to quantify the development of oysters and their gonad maturation and to identify their sex.

Animals↗

Gene expression during gonadogenesis in the chicken embryo.

Genes implicated in vertebrate sex determination and differentiation were studied in embryonic chicken gonads using reverse transcription and the polymerase chain reaction (RT-PCR). Expression profiles were obtained during gonadal sex differentiation for AMH, SOX9, SOX3, the Wilm's Tumour gene, WT1, and the orphan nuclear receptor genes, SF1 and DAX1. Some of these genes showed sexually dimorphic expression profiles during gonadal development, whereas others were expressed at similar levels in both sexes. The gene encoding Anti-Müllerian hormone (AMH) was expressed in both sexes prior to and during sexual differentiation of the gonads, with levels of expression consistently higher in males than in females. SOX9 expression was male-specific, and was up-regulated after the detection of AMH transcripts. SOX3 expression was observed prior to clear SOX9 expression and was up-regulated in both sexes at the onset of gonadal sex differentiation (but declined later in development). The WT1 gene was highly expressed in both sexes, whereas SF1 expression was clearly higher in developing ovaries compared to testes. DAX1 transcripts were observed in both sexes at all stages examined, but expression appeared somewhat higher in developing ovaries. These expression profiles are analysed in terms of current theories of vertebrate sex determination.

Actins↗