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Production of estradiol by the fetal rat testis.

Testes from 17- to 20-d-old rat fetuses were cultured in vitro for various time intervals in Medium 199 alone or with added gonadotrophins. Estradiol released into the culture media was determined by radioimmunoassay. A basal estradiol secretion rate was inexistant or undetectable at all stages studied. In early stages (17 and 18 d) there was no difference in the stimulatory effect of FSH or LH at the same concentration. At 19 d, the prevalence of FSH became apparent. At 20 d, a significant action of FSH was noted after only 3 h culture time. 1-Methyl-1, 4-androstadiene-3,17-dione, an aromatase inhibitor, markedly depressed FSH-stimulated estradiol secretion. In the 20-d old testis, stimulation of estradiol production by FSH was more rapid and reached a higher level than by (Bu)2cAMP. It is suggested that the difference in the action of LH and FSH reflects the difference in the time of appearance of the corresponding receptors in the developing fetal testis.

Animals↗

The human fetal testis is a site of expression of neurotrophins and their receptors: regulation of the germ cell and peritubular cell population.

In the fetal testis, organization of the tissue into two compartments consisting of cords containing Sertoli and germ cells surrounded by peritubular cells and of other cells within the interstitium is essential for subsequent function. Neurotrophins (NTs) act as survival and differentiation factors in the nervous system and have been detected in the developing rodent testis. Expression of mRNA for nerve growth factor; NTs 3 and 4 and brain-derived neurotrophic factor; the high-affinity receptors TrkA, TrkB, and TrkC; and the low-affinity p75 receptor were detected in the human testis between 14 and 19 wk gestation. NT4 mRNA and protein were predominantly localized to the peritubular cells. These cells were also the site of expression of p75. By contrast, nerve growth factor and NT3 were mainly expressed in Sertoli and interstitial cells. Treatment of testis organ cultures with the Trk-specific kinase inhibitor K252a resulted in a marked decrease in both gonocyte and peritubular cell number and proliferation with little effect on Sertoli cells. These data demonstrate the expression of NTs and their receptors in the human fetal testis during the second trimester and indicate possible roles in the regulation of proliferation and survival of germ cells and peritubular cells.

Adult↗

A dose-finding study of in utero and lactational exposure to diethylstilboestrol and flutamide in 129/Sv mice.

The purpose of the study was to test the experimental conditions and find the appropriate dose range for further investigations of the disruptive effects of oestrogens and antiandrogens on the development of testis. Groups of four 129/Sv mice were exposed from day 6 after mating until weaning to 1, 10 or 100 microg/kg/day of the non-steroidal oestrogen diethylstilboestrol and to 0.1, 1 or 10 mg/kg/day of the non-steroidal antiandrogen flutamide. The number of implantation sites, the litter size and reproductive parameters were examined and nipple development, anogenital distance and testicular morphology were investigated in the offspring. Dose-related post-implantation loss was seen in both diethylstilboestrol- and flutamide-treated dams and the mean litter size was smaller in the groups given the high dosages of diethylstilboestrol and flutamide. Disturbance of testicular development was seen in males exposed to diethylstilboestrol. Because of the small data material, no statistical analyses were performed. Our findings indicate that very high doses of both diethylstilboestrol and flutamide given at early stages of gestation result in a high post-implantation loss and should be avoided in further experiments using this strain which is known to have a poor reproductive performance. Exposure of the dams before mating may better reflect human exposure, but will presumably require even lower dose levels.

Androgen Antagonists↗

Both L-Lactyl and D-Lactyl Enantiomers Modify Histones in Mouse Testis.

Dynamic histone posttranslational modifications are crucial to precisely orchestrate gene expression programs. The recently discovered histone lysine lactylation has already been explored in various pathological contexts, but less in normal tissues. This modification exists as two enantiomers, L- and D-lactylation; the former may more likely modify histones due to abundant L-lactate produced by glycolysis. Here, we report the identification by proteomics of L- and D-lactylation on lysines of histones H3 and H4 in mouse testis. We developed a targeted proteomic analysis of histone peptides using synthetic sequences modified by L- or D-lactyl, to acquire reliable identification and quantification data. Some histone peptides bearing either enantiomer are separated by reversed-phase chromatography. Interestingly, despite the fact that L-lactate is much more abundant than D-lactate in mouse testis, we estimated abundance ratios of L-over D-lactylation to lie between 0.4 and 1.6 on seven residues of histones H3 and H4. Next, targeted proteomic analyses were performed on histones extracted from meiotic and postmeiotic male germ cells (spermatocytes and round spermatids, respectively), which are known to use L-lactate as a main source of energy. Nonetheless, residues 18 and 23 of histone H3 (H3K18 and H3K23) were reliably quantified and shown to harbor balanced amounts of both enantiomers. The stoichiometry of lactylation is low over the whole sequence of H3 and H4, representing about 0.01 to 0.44%: this contrasts with acetylation which exists at up to 25 to 35% relative abundances on some N-terminal lysines. Yet, lactylation appears to be more abundant than acetylation on the C-terminal half of H3 and H4, where the latter modification is scarce. Collectively, our results suggest a mechanism producing a mixture of the two enantiomers of lactate, or of a more direct substrate for lactylation, that leads to the modification of histones by L- and D-lactylation.

Animals↗

Exposure with the environmental estrogen bisphenol A disrupts the male reproductive tract in young mice.

Environmental estrogens (endocrine disruptive chemicals) have been shown to affect reproduction in wild life and it has been reported that maternal exposure with those chemicals have adverse effects on the male reproductive tract. However, little is known about the potential effects of prepubertal or pubertal exposure with environmental estrogens on the male reproductive tract. Here we examine plasma hormone levels and histology in the testis of mice following either 4- or 8-week oral administration of bisphenol A. Plasma free testosterone levels were dramatically decreased following 8 weeks of bisphenol A treatment compared with control group and morphologically multinucleated giant cells having greater than three nuclei were found in seminiferous tubules in the testis following the 8-week bisphenol A treatment. No differences in plasma corticosterone and luteinizing hormone levels were seen between bisphenol A and control groups. Thus, exposure with bisphenol A around pubertal period may directly disrupt the male reproductive tract. These facts suggest that more detailed studies will warrant the assessment of the risk to the developing human testis from exposure to bisphenol A and other environmental estrogens in prepubertal and pubertal period.

Administration, Oral↗

Subunit structure of casein kinase II from bovine testis. Demonstration that the alpha and alpha' subunits are distinct polypeptides.

The relationship between the alpha and alpha' subunits of casein kinase II was studied. For this study, a rapid scheme for the purification of the enzyme from bovine testis was developed. Using a combination of chromatography on DEAE-cellulose, phosphocellulose, hydroxylapatite, gel filtration on Sephacryl S-300 and heparin-agarose, the enzyme was purified approximately 7,000-fold. The purification scheme was completed within 48 h and resulted in the purification of milligram quantities of casein kinase II from 1 kg of fresh bovine testis. The purified enzyme had high specific activity (3,000-5,000 nmol of phosphate transferred per min/mg protein) when assayed at 30 degrees C with ATP and the synthetic peptide RRRDDDSDDD as substrates. The isolated enzyme was a phosphoprotein with an alkali-labile phosphate content exceeding 2 mol/mol protein. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis three polypeptides were apparent: alpha (Mr 45,000), alpha' (Mr 40,000), and beta (Mr 26,000). Several lines of evidence conclusively demonstrated that the alpha and alpha' subunits are distinct polypeptides. Two-dimensional maps of 125I-tryptic peptides derived from the two proteins were related, but distinct. An antipeptide antibody was raised in rabbits which reacted only with the alpha subunit on immunoblots and failed to react with either the alpha' or beta subunits. Direct comparison of peptide sequences obtained from the alpha and alpha' subunits revealed differences between the two polypeptides. The results of this study clearly demonstrate that the alpha and alpha' subunits of casein kinase II are not related by post-translational modification and are probably encoded by different genes.

Amino Acid Sequence↗

Cytologic diagnosis of rhabdomyosarcoma in a patient with germ cell tumor. A case report.

The development of sarcomas in patients with germ cell tumors is rare but has been reported previously. Theories about its pathogenesis include derivation of the tumor cells from pluripotential germ cells and malignant transformation from teratomatous elements. This report describes the occurrence of rhabdomyosarcoma in a patient with a history of mixed germ cell tumor of the testis who developed a malignant pleural effusion. Cytologic evaluation of the fluid revealed single malignant cells with high nuclear/cytoplasmic ratios and prominent nucleoli. While these features initially suggested a diagnosis of metastatic seminoma, careful cytologic and immunohistochemical examination revealed features consistent with metastatic rhabdomyosarcoma. The exfoliative cytologic findings of other germ cell tumors are also discussed.

Adult↗

The immunohistochemical localization of transforming growth factor-beta 2 in the fetal and neonatal rat testis.

The localization of transforming growth factor beta-2 (TGF beta 2) in the fetal and neonatal testis (from day 13.5 of fetal life to postnatal day 9) was investigated by an immunohistochemical staining method employing a specific polyclonal antibody. Immunostaining appeared on fetal day 13.5 in primitive Sertoli cells as they begin to come in contact with each other and surround the germ cells to form the seminiferous cords. Staining in Sertoli cells was still clearly observed until fetal day 16.5 and became faint or undetectable from fetal day 18.5 onwards. In fetal-type Leydig cells, a positive reaction for TGF beta 2 appeared on day 16.5 and became very intense from day 18.5 onwards. In the germ cells, immunoreactivity for TGF beta 2 appeared on fetal day 20.5, rose to a maximum on postnatal day 4 and decreased thereafter. On postnatal day 9, staining was still present in type A spermatogonia and absent in type B spermatogonia. No immunoreactivity was detected in peritubular cells on any day studied. In conclusion, our results are in favour of an autocrine/paracrine role of TGF beta 2 in the differentiation of the testis during the perinatal period. It may be involved in the organization of the seminiferous cords, the regulation of testosterone production and the regulation of the number of germ cells. When compared with the immunolocalization of TGF beta 1 that we have previously reported [1], the present study suggests that the roles of TGF beta 2 in the developing rat testis can be specific but also overlap from those of TGF beta 1.

Animals↗

Pattern of gonadotropin secretion and ultrasonographic evaluation of developmental changes in the testis of early and late maturing bull calves.

This was a study that retrospectively analyzed serum gonadotropin secretion and the ultrasonographic appearance of the testis during development in prepubertal bull calves to determine whether there were differences between early and late maturing bulls. Blood samples were taken every other week from 2 wk of age until puberty. Samples were also taken at 12 minute intervals for 12 hours at 4, 10, 20, 25, 30, 35, 40 and 45 wk of age. The GnRH treatment was administered 10 hours after the start of each period of frequent blood sampling. Bull calves fell into two distinctive groups, with one group maturing between 36.6 and 44.2 wk (n = 12) and the other between 46.4 and 48.9 wk of age (n = 8). In samples taken every other week mean serum LH concentrations were greater in early maturing bulls than in late maturing bulls at 12, 14 and 16 wk of age (P<0.05). In blood samples taken every 12 minutes for 10 hours early maturing bull calves had higher mean serum LH concentrations at 4 and 10 wk of age (P<0.05) and higher LH pulse frequency at 10 and 20 wk of age (P<0.05). Mean serum LH concentrations at 4, 10 and 40 wk of age and LH pulse frequency at 10 and 20 wk of age were negatively correlated with age at puberty in bull calves. Mean pixel units of the right and left testis were higher from 34 to 40 wk of age in early maturing than in late maturing animals (P<0.05). It seems possible that hormone measurements and ultrasonographic characteristics of the testes could be developed into powerful tools for studies on the regulation of reproductive development and may aid in the prediction of reproductive potential.

Animals↗

[Histological characteristics of the testis and epididymis of calves during postnatal development].

A histologic investigation was carried out of the testis and epididymis of a total of 42 calves of the Black-and-white grouped in 14 test groups. The animals were currently castrated after the age of 2 months and were raised in the open. Material for investigation was taken on the 1st and 15th day, and once in every following month up to the completion of 12 months of age. It was found that up to the 3rd month there was no lumen in the testicular tubules, but only differentiating Sertoli cells and gonocytes were seen. Spermatogonia and spermatocytes were first observed at the age of 4 months, and prespermatids and sporadic spermatids--in the 8th month. Single spermatozoa in the phase of final shaping appeared in the 10th month, and completed spermatogenesis--in the 11th month. The histogenesis of the wall of the tubules of the epididymis completed earlier as compared to the histogenesis process in the testis--by the age of 8 months. Later on there were morphologically in the epididymis some proliferation and folding of the tubules with widening of the lumen, whereupon the amount of the surrounding connective tissue relatively decreased.

Animals↗

Cancer of the undescended or maldescended testis.

An analysis of 45 cryptorchids (by history or examination) with a testicular cancer treated at Memorial Hospital, between 1934 and 1973, is presented. Twenty-five patients had the cryptorchid state repaired at ages four to 27 years, either spontaneously or by orchiopexy or hormonal therapy. Ipsilateral (24) or contralateral (one) intrascrotal testis tumors developed four to 47 years later. Twenty cryptorchid patients presented with ipsilateral inguinal (eleven), abdominal (seven), or contralateral intrascrotal (two) tumors. There were 18 pure seminomas, 17 embryonal carcinomas, nine teratocarcinomas, and one reticulum cell sarcoma. Five year survival rates as estimated by the product-limit method were 60% for the unrepaired cases and 41% for the repaired cases. The survival seems to follow histologic type and anatomical stage, whether the testis is within the scrotum or not. Five year survival similarly estimated was 78% in the seminomas and 29% in the other tumors. Twelve of thirteen survivors (including nine with seminoma) received postoperative irradiation to the regional lymphatics and eleven were without recurrent tumor for periods ranging from six to 28 years.

Adolescent↗

Newborn and immature rat testes contain gonadotropin-releasing hormone (GnRH) receptors, and their testosterone production is stimulated by a GnRH agonist in vitro.

The possibility of direct gonadal effects of GnRH in the newborn and immature rat testis was studied by two approaches: the presence of specific high-affinity receptors for a GnRH agonist analogue (Buserelin) was studied in animals between the ages of 1 and 60 days; and the effect of Buserelin on testicular testosterone production was studied in vitro. GnRH receptors were found in the testis tissue at all ages studied, and there was a gradual increase in the number of binding sites per testis with advancing age. The highest concentration of GnRH receptors was found in the rat testis tissue on day 1 of life, 3.2 +/- 0.4 fmoles/g tissue (mean +/- SE, n = 5). The binding declined to a nadir, 1.5 +/- 0.5 fmoles/g tissue (in = 10), on day 15 post-partum (p less than 0.05), and thereafter increased again gradually to a level of 2.7 +/- 0.3 fmoles/g (n = 5) on day 60 of life (p less than 0.05). The biphasic concentration curve of GnRH receptors followed closely the volume density of Leydig cells in the developing rat testis. Buserelin at a concentration of 10(-7) M was able to stimulate significantly (p less than 0.01), by 98-146%, the testosterone production of decapsulated testes at all ages studied between 1 and 60 days of life. These results indicate the presence of functional GnRH receptors in the testis tissue of newborn and immature rats.

Animals↗

Suppressive effect of perinatal testes on the differentiation of fetal ovaries transplanted into adult males in the rat.

A 14 d ovarian primordium was transplanted with a fetal testis (13-18 d and 21 d of gestation) or a neonatal testis (15, 20, 30 and 45 d after birth) into the renal subcapsular position of an adult male rat. Two weeks after transplantation, transplants were examined as to the degree of ovarian and testicular differentiation. In the combination of a 14 d ovary and a 13 d testis, there were 3 types of result: either the ovary or the testis alone developed or both gonads developed well. Ovaries transplanted in union with 15-18 d testes did not develop, although the testes developed normally. Some ovaries in union with 21 d testes developed normally. In combination with infantile testes, the incidence of developed ovaries increased as the age of testes advanced. These results suggest that the 13 d fetal testes begin to suppress the development of cotransplanted 14 d ovaries, that 14-18 d fetal testes maintain such suppressive effects and that this effect gradually diminishes in infantile testes as they progress toward 45 d after birth.

Animals↗

The prepubertal testis: a quiescent or a silently active organ?

The development of the testis is characterised by dramatic changes between birth and adulthood. The most conspicuous changes take place during puberty, when the seminiferous tubule diameter increases significantly owing to an important proliferation of germ cells, giving rise to spermatozoa, and to the development of a tubular lumen; in the interstitial tissue characteristic Leydig cells appear and secrete high levels of testosterone. These pubertal changes of the testis can be detected clinically since they result in testicular volume and serum androgen level increments. The prepubertal testis has classically been defined as a quiescent organ. However, since adequate stereological methods for microscopic analysis are available, it has been shown that the male gonad triplicates its volume between birth and the onset of puberty. Sertoli cells and spermatogonia proliferate intensely; this is critical for the development of quantitatively normal adult spermatogenesis. Seminiferous tubule volume increases owing to an increment in tubular length, not diameter. Sertoli cells are also functionally active during childhood: they produce high amounts of anti-Müllerian hormone during the whole prepubertal period, and inhibin B until the age of 2-4 years. Anti-Müllerian hormone and inhibin seem to play a role as modulators of the proliferation and differentiation of Leydig cell precursors.

Adult↗

Immunocytochemical localization of bioregulatory peptides in marmoset testes.

Immunocytochemical localization of neuropeptides (beta-endorphin, substance P, arginine vasopressin, oxytocin), pituitary hormones (adrenocorticotropin, prolactin, growth hormone, follicle stimulating hormone (FSH), gonadal inhibin, gastrin, and human chorionic gonadotrophin (hCG)) was carried out in marmoset testis during development. Both intensity of immunostaining and distribution of these peptides in testicular compartments viz. seminiferous tubules and Leydig cells changed dramatically during development. In vitro biosynthesis of inhibin and FSH was increased by hCG, whereas prolactin (5 micrograms) and prostatic inhibin peptide suppressed the synthesis of these hormones.

Aging↗

Related function of mouse SOX3, SOX9, and SRY HMG domains assayed by male sex determination.

Sox genes encode proteins related to each other, and to the sex determining gene Sry, by the presence of a DNA binding motif known as the HMG domain. Although HMG domains can bind to related DNA sequences, Sox gene products may achieve target gene specificity by binding to preferred target sequences or by interacting with specific partner proteins. To assess their functional similarities, we replaced the HMG box of Sry with the HMG box of Sox3 or Sox9 and tested whether these constructs caused sex reversal in XX mice. Our results indicate that such chimeric transgenes can functionally replace Sry and elicit development of testis cords, male patterns of gene expression, and elaboration of male secondary sexual characteristics. This implies that chimeric SRY proteins with SOX HMG domains can bind to and regulate SRY target genes and that potential SRY partner factor interactions are not disrupted by HMG domain substitutions. genesis 28:111-124, 2000.

Amino Acid Sequence↗

Structure of the right testis of sexually mature genetically female fowl experimentally masculinized during embryonic life and submitted to a posthatching left castration.

Posthatching left castration of genetically female fowl, Gallus domesticus, preceded, during embryonic life, by a masculinizing treatment associating a testis graft and an antiestrogen resulted in the development of the right rudimentary gonad into a testis. Examined after the sexual maturity, the right testis of most treated animals was entirely composed of seminiferous tubules possessing a spermatogenic cell complement. Spermiogenesis proceeded to the stage of spermatozoon in 4 out of 17 treated animals and was almost as well organized as in a normal cock testis in 3 of them. Testis development appeared then as clearly improved, compared to that described previously in only left-castrated, with or without treatment with an antiestrogen, or only sex-reversed female fowl. The possible mechanism of this improvement is discussed.

Animals↗

A vector-primer-cloner-sequencer plasmid for the construction of cDNA libraries: evidence for a rat glyceraldehyde-3-phosphate dehydrogenase-like mRNA and a ferritin mRNA within testis.

We have developed a vector-primer-cloner-sequencer (VPCS), based on the pUC plasmids, which is easy to prepare. Stable cDNA libraries have been generated from human, rat, and bull testis. The inserts are of various size classes, including high-molecular-weight reverse transcripts which can be easily sequenced. The utility of this VPCS has been demonstrated by isolating a rat ferritin and glyceraldehyde-3-phosphate dehydrogenase-like clone.

Animals↗