Transforming growth factor-β (TGF-β) and bone morphogenetic protein (BMP) signaling has fundamental roles in the regulation of the stem cell niche for both embryonic and adult stem cells. In zebrafish, male germ stem cell niche is regulated by follicle-stimulating hormone (Fsh) through different members of the TGF-β superfamily. On the other hand, the specific roles of TGF-β and BMP signaling pathways are unknown in the zebrafish male germ stem cell niche. Considering this lack of information, the present study aimed to investigate the pharmacological inhibition of TGF-β (A83-01) and BMP (DMH1) signaling pathways in the presence of recombinant zebrafish Fsh using testicular explants. We also reanalyzed single cell-RNA sequencing (sc-RNA-seq) dataset from adult zebrafish testes to identify the testicular cellular sites of smad expression, and to understand the physiological significance of the changes in smad transcript levels after inhibition of TGF-β or BMP pathways. Our results showed that A83-01 potentiated the pro-stimulatory effects of Fsh on spermatogonial differentiation leading to an increase in the proportion area occupied by differentiated spermatogonia with concomitant reduction of type A undifferentiated (Aund) spermatogonia. In agreement, expression analysis showed lower mRNA levels for the pluripotency gene pou5f3, and increased expression of dazl (marker of type B spermatogonia and spermatocyte) and igf3 (pro-stimulatory growth factor) following the co-treatment with TGF-β inhibitor and Fsh. Contrariwise, the inhibition of BMP signaling nullified the pro-stimulatory effects of Fsh, resulting in a reduction of differentiated spermatogonia and increased proportion area occupied by type Aund spermatogonia. Supporting this evidence, BMP signaling inhibition increased the mRNA levels of pluripotency genes nanog and pou5f3, and decreased dazl levels when compared to control. The sc-RNA-seq data unveiled a distinctive pattern of smad expression among testicular cells, primarily observed in spermatogonia (smad 2, 3a, 3b, 8), spermatocytes (smad 2, 3a, 8), Sertoli cells (smad 1, 3a, 3b), and Leydig cells (smad 1, 2). This finding supports the notion that inhibition of TGF-β and BMP signaling pathways may predominantly impact cellular components within the spermatogonial niche, namely spermatogonia, Sertoli, and Leydig cells. In conclusion, our study demonstrated that TGF-β and BMP signaling pathways exert antagonistic roles in the zebrafish germ stem cell niche. The members of the TGF-β subfamily are mainly involved in maintaining the undifferentiated state of spermatogonia, while the BMP subfamily promotes spermatogonial differentiation. Therefore, in the complex regulation of the germ stem cell niche by Fsh, members of the BMP subfamily (pro-differentiation) should be more predominant in the niche than those belonging to the TGF-β (anti-differentiation). Overall, these findings are not only relevant for understanding the regulation of germ stem cell niche but may also be useful for expanding in vitro the number of undifferentiated spermatogonia more efficiently than using recombinant hormones or growth factors.
- MeSH
- buněčná diferenciace genetika MeSH
- dánio pruhované * genetika MeSH
- folikuly stimulující hormon farmakologie metabolismus MeSH
- messenger RNA genetika MeSH
- pyrazoly * MeSH
- spermatogeneze genetika MeSH
- spermatogonie * metabolismus MeSH
- testis metabolismus MeSH
- thiosemikarbazony * MeSH
- transformující růstový faktor beta metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Bone morphogenetic proteins (BMPs) and receptors (BMPR-1A, BMPR-1B, BMPR-2) have been shown to be vital for female reproduction, while their roles in males are poorly described. Our study was undertaken to specify the function of BMPR-1B in steroidogenic enzyme gene expression, testosterone production and reproductive development in male mice, given that Bmpr1b mRNA is expressed in mouse testis and Bmpr1b knockout results in compromised fertility. Male mice were passively immunized for 6 days with anti-BMPR-1B in the presence or absence of exogenous gonadotrophins. We then measured the effects of anti-BMPR-1B on testicular hydroxysteroid dehydrogenase isoforms (Hsd3b1, Hsd3b6, and Hsd17b3) and aromatase (Cyp19) mRNA expression, testicular and serum testosterone levels, and testis and seminal vesicle weight. In vitro testosterone production in response to anti-BMPR-1B was determined using testicular culture, and Leydig cell culture in the presence or absence of gonadotrophins. In Leydig cell culture the contribution of seminiferous tubules and Leydig cells were examined by preconditioning the media with these testicular constituents. In adult mice, anti-BMPR-1B increased testosterone and Hsd3b1 but decreased Hsd3b6 and Cyp19 mRNA. In adult testicular culture and seminiferous tubule conditioned Leydig cell culture, anti-BMPR-1B reduced testosterone, while in normal and Leydig cell conditioned Leydig cell culture it increased testosterone levels. In pubertal mice, anti-BMPR-1B reduced gonadotrophin stimulated seminal vesicle growth. In conclusion, BMPR-1B has specific developmental functions in the autocrine and paracrine regulation of testicular steroidogenic enzyme gene expression and testosterone production in adults and in the development of seminal vesicles during puberty.
- MeSH
- aromatasa metabolismus MeSH
- exprese genu MeSH
- kostní morfogenetické proteiny metabolismus MeSH
- messenger RNA genetika metabolismus MeSH
- myši MeSH
- pohlavní dospělost MeSH
- receptory kostního morfogenetického proteinu metabolismus MeSH
- testis * metabolismus MeSH
- testosteron * MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The mycotoxin zearalenone (ZEA) in food and feed seriously harms human and animal health. How to reduce its toxicity is an important direction of current research on food safety. This study aim to assess the effects of procyanidins (PC) on cell apoptosis caused by ZEA and to clarify the role of Nrf2 in the process. Swine testicle (ST) cells were treated with ZEA (57.5 μmol/L) and/or PC (10 mg/L) for 24 h. Cell viability was detected by CCK-8 assay. Cell apoptosis and the level of ROS were detected by flow cytometry. The expression levels of mRNA and protein was detected by qRT-PCR and western blotting. Our results showed that ZEA reduced the antioxidant capacity of the ST cells, induced the cell apoptosis and inhibited the gene and protein expression of Nrf2 and its downstream genes (ho-1,nqo1), while PC improved the cell antioxidant capacity, reduced the degree of ZEA-induced cell apoptosis and promoted the gene and protein expression of Nrf2 and its downstream genes. However, when the Nrf2 small molecule inhibitor ML385 was added, the ability of PC to inhibit ZEA-induced cell apoptosis and promote the expression of Nrf2 and its downstream genes were decreased. Our results demonstrated that ZEA induced oxidative stress and apoptosis of ST cells, which were alleviated by PC intervention via activating Nrf2 signaling pathway. This finding of this study provided a molecular basis for the clinical application of PC to prevent ZEN-caused reproductive toxicity.
- MeSH
- antioxidancia metabolismus farmakologie MeSH
- apoptóza MeSH
- faktor 2 související s NF-E2 genetika metabolismus MeSH
- oxidační stres MeSH
- prasata MeSH
- proantokyanidiny * farmakologie MeSH
- reaktivní formy kyslíku metabolismus MeSH
- signální transdukce MeSH
- testis metabolismus MeSH
- zearalenon * metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
PIWI-interacting RNAs (piRNAs) support the germline by suppressing retrotransposons. Studies of the pathway in mice have strongly shaped the view that mammalian piRNAs are essential for male but not for female fertility. Here, we report that the role of the piRNA pathway substantially differs in golden hamsters (Mesocricetus auratus), the piRNA pathway setup of which more closely resembles that of other mammals, including humans. The loss of the Mov10l1 RNA helicase-an essential piRNA biogenesis factor-leads to striking phenotypes in both sexes. In contrast to mice, female Mov10l1-/- hamsters are sterile because their oocytes do not sustain zygotic development. Furthermore, Mov10l1-/- male hamsters have impaired establishment of spermatogonia accompanied by transcriptome dysregulation and an expression surge of a young retrotransposon subfamily. Our results show that the mammalian piRNA pathway has essential roles in both sexes and its adaptive nature allows it to manage emerging genomic threats and acquire new critical roles in the germline.
- MeSH
- křečci praví MeSH
- křeček rodu Mesocricetus metabolismus MeSH
- malá interferující RNA genetika MeSH
- oocyty metabolismus patologie MeSH
- retroelementy fyziologie MeSH
- RNA-helikasy genetika MeSH
- spermatogeneze genetika fyziologie MeSH
- spermatogonie metabolismus patologie MeSH
- testis metabolismus MeSH
- umlčování genů fyziologie MeSH
- zvířata MeSH
- Check Tag
- křečci praví MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Androgens represent the main hormones responsible for maintaining hormonal balance and function in the prostate and testis. As they are involved in prostate and testicular carcinogenesis, more detailed information of their active concentration at the site of action is required. Since the introduction of the term intracrinology as the local formation of active steroid hormones from inactive precursors of the adrenal gland, mainly dehydroepiandrosterone (DHEA) and DHEA-S, it is evident that blood circulating levels of sex steroid hormones need not reflect their actual concentrations in the tissue. Here, we review and critically evaluate available methods for the analysis of human intraprostatic and intratesticular steroid concentrations. Since analytical approaches have much in common in both tissues, we discuss them together. Preanalytical steps, including various techniques for separation of the analytes, are compared, followed by the end-point measurement. Advantages and disadvantages of chromatography-mass spectrometry (LC-MS, GC-MS), immunoanalytical methods (IA), and hybrid (LC-IA) are discussed. Finally, the clinical information value of the determined steroid hormones is evaluated concerning differentiating between patients with cancer or benign hyperplasia and between patients with different degrees of infertility. Adrenal-derived 11-oxygenated androgens are mentioned as perspective prognostic markers for these purposes.
- MeSH
- androgeny metabolismus MeSH
- lidé MeSH
- nadledviny metabolismus MeSH
- pohlavní steroidní hormony metabolismus MeSH
- prostata metabolismus MeSH
- steroidy metabolismus MeSH
- testis metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Through their receptors at each level of hypothalamo-pituitary-gonadal axis glucocorticoid excess, either endogenous or administered or stress-induced, could affect steroid production in the testis and thus male fertility. The main ways by which glucocorticoids act are as follows: 1) Affecting gonadoliberin and LH synthesis and release through glucocorticoid receptors in hypothalamic neurons and pituitary gonadotropes. 2) By so far not clearly evidenced reduction of the number of LH receptors on the membrane of Leydig cells. 3) By affecting expression and function of steroidogenic enzymes in the testis. 4) By regulation of in situ access of glucocorticoid to its target cells in the testis. 5) By promotion Leydig cell apoptosis. The review provides a survey of physiological and molecular mechanisms staying behind these effects. It does not deal with the clinical effects of glucocorticoid treatment which would substantially exceed the scope of the pater.
- MeSH
- glukokortikoidy farmakologie MeSH
- lidé MeSH
- steroidy biosyntéza MeSH
- testis účinky léků metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Chronic stress is a crucial public issue that occurs when a person is repetitively stimulated by various stressors. Previous researches have reported that chronic stress induces spermatogenesis dysfunction in the reproductive system, but its molecular mechanisms remain unclear. The nectin protein family, including nectin-1 to nectin-4, is Ca(2+)-independent immunoglobulin-like cell adhesion molecules, that are widely expressed in the hippocampus, testicular tissue, epithelial cells and other sites. Nectin-3 contributes to the sperm development at the late stage, and the abnormal expression of nectin-3 impairs spermatogenesis. Some recent studies have demonstrated that stress induces a decrease in nectin-3 expression in the hippocampus via corticotropin-releasing hormone (CRH) to corticotropin-releasing hormone receptor 1 (CRHR1) pathway. Here, we tested whether chronic stress also caused a reduction in nectin-3 expression in the testis. We established a chronic social defeat stress paradigm, which provides naturalistic and complex chronic stress inmale C57BL/6 mice. After 25 days of chronic social defeat stress, the mice showed weight loss, thymic atrophy and some other typical symptoms of chronic stress (e.g.anxiety-like behavior and social avoidance behavior). We found gonad atrophy, testicular histological structure changes and semen quality reductions in the stressed mice. The stressed male mice significantly spent more time to impregnate the female mice than the control male mice. Moreover, nectin-3 protein levels in stressed mice were significantly decreased in the testes compared with those in control mice. In addition, we found that the CRHR1 expression level was increased in the testes of stressed mice. Therefore, we demonstrated a decreased level of nectin-3 expression and an increase in CRHR1 expression in the testis after exposure to chronic stress, which may provide a potential therapeutic target for the spermatogenesis dysfunction induced by chronic stress.
- MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- nektiny biosyntéza MeSH
- psychický stres metabolismus patofyziologie MeSH
- receptory hormonu uvolňujícího kortikotropin biosyntéza metabolismus MeSH
- sociální interakce MeSH
- spermatogeneze fyziologie MeSH
- testis metabolismus patofyziologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The physiological importance of CD151 tetraspanin is known from somatic cells and its outside-in signalling through integrins was described. In male germ cells, two tetraspanins, CD9 and CD81, are involved in sperm-egg membrane fusion, and similarly to integrins, they occupy characteristic regions. We report here on a newly discovered presence of CD151 in sperm, and present its expression and distribution during spermatogenesis and sperm transition during the acrosome reaction. We traced CD151 gene and protein expression in testicular cell subpopulations, with strong enrichment in spermatogonia and spermatids. The testicular and epididymal localization pattern is designated to the sperm head primary fusion site called the equatorial segment and when compared to the acrosome vesicle status, CD151 was located into the inner acrosomal membrane overlying the nucleus. Moreover, we show CD151 interaction with α6 integrin subunit, which forms a dimer with β4 as a part of cis-protein interactions within sperm prior to gamete fusion. We used mammalian species with distinct sperm morphology and sperm maturation such as mouse and bull and compared the results with human. In conclusion, the delivered findings characterise CD151 as a novel sperm tetraspanin network member and provide knowledge on its physiology in male germ cells.
- MeSH
- antigeny CD151 chemie genetika metabolismus MeSH
- exprese genu * MeSH
- fluorescenční protilátková technika MeSH
- integrin alfa6 chemie metabolismus MeSH
- konformace proteinů MeSH
- lidé MeSH
- molekulární modely MeSH
- myši MeSH
- spermie metabolismus MeSH
- testis metabolismus MeSH
- transport proteinů MeSH
- vazba proteinů MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zárodečné buňky metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Long transgenes are often used in mammalian genetics, e.g., to rescue mutations in large genes. In the course of experiments addressing the genetic basis of hybrid sterility caused by meiotic defects in mice bearing different alleles of Prdm9, we discovered that introduction of copy-number variation (CNV) via two independent insertions of long transgenes containing incomplete Prdm9 decreased testicular weight and epididymal sperm count. Transgenic animals displayed increased occurrence of seminiferous tubules with apoptotic cells at 18 days postpartum (dpp) corresponding to late meiotic prophase I, but not at 21 dpp. We hypothesized that long transgene insertions could cause asynapsis, but the immunocytochemical data revealed that the adult transgenic testes carried a similar percentage of asynaptic pachytene spermatocytes as the controls. These transgenic spermatocytes displayed less crossovers but similar numbers of unrepaired meiotic breaks. Despite slightly increased frequency of metaphase I spermatocytes with univalent chromosome(s) and reduced numbers of metaphase II spermatocytes, cytological studies did not reveal increased apoptosis in tubules containing the metaphase spermatocytes, but found an increased percentage of tubules carrying apoptotic spermatids. Sperm counts of subfertile animals inversely correlated with the transcription levels of the Psmb1 gene encoded within these two transgenes. The effect of the transgenes was dependent on sex and genetic background. Our results imply that the fertility of transgenic hybrid animals is not compromised by the impaired meiotic synapsis of homologous chromosomes, but can be negatively influenced by the increased expression of the introduced genes.
- MeSH
- apoptóza genetika MeSH
- dvouřetězcové zlomy DNA MeSH
- fertilita genetika MeSH
- genetické pozadí MeSH
- kontrolní body buněčného cyklu genetika MeSH
- myši MeSH
- pachytenní stadium genetika MeSH
- počet spermií MeSH
- spermatocyty metabolismus MeSH
- testis anatomie a histologie metabolismus MeSH
- transgeny * MeSH
- variabilita počtu kopií segmentů DNA * MeSH
- velikost orgánu MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Estrogens are steroid hormones that affect a wide range of physiological functions. The effect of estrogens on male reproductive tissues and sperm cells through specific receptors is essential for sperm development, maturation, and function. Although estrogen receptors (ERs) have been studied in several mammalian species, including humans, they have not yet been described in bull spermatozoa and reproductive tissues. In this study, we analyzed the presence of all types of ERs (ESR1, ESR2, and GPER1) in bull testicular and epididymal tissues and epididymal and ejaculated spermatozoa, and we characterize them here for the first time. We observed different localizations of each type of ER in the sperm head by immunofluorescent microscopy. Additionally, using a selected polyclonal antibody, we found that each type of ER in bull sperm extracts had two isoforms with different molecular masses. The detailed detection of ERs is a prerequisite not only for understanding the effect of estrogen on all reproductive events but also for further studying the negative effect of environmental estrogens (endocrine disruptors) on processes that lead to fertilization.
- MeSH
- epididymis metabolismus MeSH
- receptory pro estrogeny metabolismus MeSH
- receptory spřažené s G-proteiny metabolismus MeSH
- rozmnožování * MeSH
- skot metabolismus MeSH
- spermie metabolismus MeSH
- testis metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- skot metabolismus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
