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1.
Spontaneous germ cell death by apoptosis occurs during normal spermatogenesis in mammals and is thought to play a role in the physiological mechanism limiting the clonal expansion of such cell population in the male gonad. In the prepubertal rat testis, the most conspicuous dying cells are pachytene spermatocytes, which are also the primary target of the apoptosis experimentally induced by the methoxyacetic acid (MAA). Since we have recently reported that Sertoli cells, the somatic component of the seminiferous epithelium, regulate not only germ cell viability and differentiation but also their death, we have further investigated the mechanism involved in such a control.  相似文献   

2.
In this study the cellular mechanisms of male sterility in F1 hybrids (BNF1) between BALB/c and wild-derived M.MUS-NJL (NJL) was investigated. Cell proliferation and differentiation in the sterile testis were examined by bromodeoxyuridine-labeling and use of germ cell stage-specific antibodies. In BNF1 testes, spermatogonia actively proliferated with a seminiferous epithelial cycle, and were retained in the basal layer of the tubules. However, preleptotene, leptotene and zygotene spermatocytes moved to the adluminal region. Immunohistological data with germ cell stage-specific antibodies indicated the presence of few, if any, pachytene spermatocytes in BNF1 testes. Thus, spermatogenesis seemed to be blocked at the zygotene stage. For examination of germ cell-Sertoli cell interactions, testes of aggregation chimeras between BNF1 and C3H/HeN were analyzed immunohistologically with C3H-specific antibody. Results showed that spermatogenesis of C3H-germ cells was normal, even when these cells in contact with BNF1-Sertoli cells. Differentiation of BNF1-germ cells progressed from zygotene to pachytene stage spermatocytes when these cells were surrounded by C3H-Sertoli cells, but never proceeded beyond the pachytene stage. These observations suggest that at least two different cellular factors may be involved in spermatogenesis, one acting in the germ cells and the other mediated by Sertoli cells. Furthermore, mating experiments revealed that the degree of spermatogenesis varied in different F1 hybrids, and that the major sterility factor was closely linked to the T -locus on chromosome 17.  相似文献   

3.
The direct influence of germ cells and residual bodies on Sertoli cell basal and FSH-stimulated secretion of androgen-binding protein (ABP) was studied using Sertoli cells, recovered from 20-day-old rats, cultured alone or cocultured with a crude germ cell preparation from adult rats or with pachytene spermatocytes, round spermatids or populations of residual bodies enriched by centrifugal elutriation. The effect of a rat liver epithelial cell line (LEC) on Sertoli cell function was also tested. Addition of a crude germ cell preparation increased basal and FSH-stimulated ABP secretion. Pachytene spermatocytes and residual bodies adhered to the Sertoli cell monolayer to a much greater extent than did round spermatids. Addition of pachytene spermatocytes markedly enhanced basal and FSH-stimulated ABP secretion over 12 days of culture. Round spermatids and residual bodies stimulated ABP secretion although to a lesser extent than did spermatocytes. Furthermore, the increase of FSH-stimulated ABP levels was not maintained after 4 or 8 days of culture. LEC also enhanced basal and FSH-induced ABP levels but the increase of FSH-induced ABP production was only observed until Day 8 of culture. The influence of LEC on Sertoli cell secretion could be mediated through the production of an extracellular matrix. It is concluded that germ cells, particularly pachytene spermatocytes, can directly stimulate Sertoli cell secretory activity in vitro.  相似文献   

4.
Androgen receptor-interacting protein 4 (ARIP4) belongs to the SNF2 family of proteins involved in chromatin remodeling, DNA excision repair, and homologous recombination. It is a DNA-dependent ATPase, binds to DNA and mononucleosomes, and interacts with androgen receptor (AR) and modulates AR-dependent transactivation. We have examined in this study the expression and cellular localization of ARIP4 during postnatal development of mouse testis. ARIP4 was detected by immunohistochemistry in Sertoli cell nuclei at all ages studied, starting on day 5, and exhibited the highest expression level in adult mice. At the onset of spermatogenesis, ARIP4 expression became evident in spermatogonia, pachytene, and diplotene spermatocytes. Immunoreactive ARIP4 antigen was present in Leydig cell nuclei. In Sertoli cells ARIP4 was expressed in a stage-dependent manner, with high expression levels at stages II-VI and VII-VIII. ARIP4 expression patterns did not differ significantly in testes of wild-type, follicle-stimulating hormone receptor knockout, and luteinizing hormone receptor knockout mice. In testes of hypogonadal mice, ARIP4 was found mainly in interstitial cells and exhibited lower expression in Sertoli and germ cells. In vitro stimulation of rat seminiferous tubule segments with testosterone, FSH, or forskolin did not significantly change stage-specific levels of ARIP4 mRNA. Heterozygous ARIP4(+/-) mice were haploinsufficient and had reduced levels of Sertoli-cell specific androgen-regulated Rhox5 (also called Pem) mRNA. Collectively, ARIP4 is an AR coregulator in Sertoli cells in vivo, but the expression in the germ cells implies that it has also AR-independent functions in spermatogenesis.  相似文献   

5.
The finding of large, stage-specific changes in secretion of procathepsin L by rat Sertoli cells has led to the hypothesis that this proenzyme promotes the survival, replication, or differentiation of spermatogenic cells. Experiments described herein used a mouse model to test this hypothesis. To prove that mice are appropriate for this purpose, we first demonstrate that mature mouse Sertoli cells express cathepsin L mRNA in the same stage-specific manner as rat Sertoli cells and they also secrete procathepsin L. To test whether catalytically active cathepsin L is required for normal spermatogenesis, we examined the testes of 110- to 120-day-old furless mice, which express catalytically inactive cathepsin L. Morphologic examination of testes of furless mice revealed both normal and atrophic seminiferous tubules. Enumeration of atrophic tubules in furless and control mice demonstrates that lack of functional cathepsin L results in a 12-fold increase in seminiferous tubule atrophy. To determine whether lack of functional cathepsin L affects the production of male germ cells in apparently normal, nonatrophic tubules, we compared numbers in control and furless mice of preleptotene spermatocytes, pachytene spermatocytes, and round spermatids per Sertoli cell. Results demonstrate that the lack of functional cathepsin L causes a 16% reduction in formation of preleptotene spermatocytes and a 25% reduction in differentiation of these cells into pachytene spermatocyte. These results suggest that procathepsin L either directly or indirectly has two distinct functions in the testis. This proenzyme prevents atrophy of seminiferous tubules and promotes the formation of preleptotene spermatocytes and the differentiation of these meiotic cells into pachytene spermatocytes.  相似文献   

6.
7.
The cells with nuclear DNA fragmentation related to apoptosis were detected by TUNEL technique in the seminiferous epithelium of control rats and of rats with experimental hyperprolactinemia induced by metoclopramide. The percentage of convoluted tubules with apoptotic cells and the number of apoptotic cells (predominantly spermatogonia and spermatocytes) was increased in the experimental group. The results indicated stage-specific germ cell apoptosis. In the experimental group, apoptotic cells were most evident at early (I-IV), middle (VII-VIII) and late (XII-XIV) stages of the seminiferous epithelium cycle, as revealed by light and electron microscopy. We suggest that a decreased concentration of testosterone and an increased concentration of prolactin could disturb spermatogenesis and contribute to the intensive apoptosis of germ cells in rats with hyperprolactinemia. Sertoli cells which have receptors for testosterone and prolactin and play an important role in spermatogenesis and in the initiation of apoptosis in seminiferous epithelium, could mediate such an influence of both hormones.  相似文献   

8.
Immunohistochemical localization of sulphydryloxidase was examined in the testis of the Djungarian hamster from Day 0 to Day 31 of post-natal development. The sulphydryloxidase antibody labelled prespermatogonia and the first population of spermatogonia type A within the seminiferous epithelium. Additionally, Sertoli cells exhibited immunoreactivity from Day 2 to Day 11 after birth. From Day 11 onwards, sulphydryloxidase immunoreactivity was found in germ cells after the initiation spermatogenesis from pachytene primary spermatocytes, showing the highest intensity in mid-pachytene spermatocytes. The pattern of sulphydryloxidase expression during spermatogenesis was identical to that found in adult animals. It is concluded that sulphydryloxidase immunoreactivity not only serves as a marker for early stages of spermatogenesis, especially pachytene spermatocytes, confirming earlier reports, but also for spermatogonial precursors.  相似文献   

9.
10.
Three spermatogenic cell populations isolated from prepuberal mice--type B spermatogonia, preleptotene spermatocytes, and leptotene/zygotene spermatocytes--were used to elicit distinct polyclonal antisera. Surface binding specificities were determined for purified IgGs by indirect immunofluorescence and rosette assays on live cells. Binding activities were assayed both before and after absorptions with a variety of somatic and spermatogenic cells. Each of these antisera binds to surface antigens that are present on germ cells throughout spermatogenesis and are not shared by splenocytes, thymocytes, and erythrocytes. Only the antiserum raised against leptotene and zygotene spermatocytes (ALZ) recognizes a stage-specific subset of surface determinants. After appropriate absorptions, ALZ binds to the surface of early pachytene spermatocytes and germ cells at subsequent stages of differentiation, including vas deferens spermatozoa. Antigens which react with this absorbed IgG are not detected on the surface of spermatogonia or meiotic cells prior to pachynema, including leptotene and zygotene spermatocytes. The observed binding specificities may result from the synthesis of one or more surface molecules during the early meiotic stages, followed by delayed insertion into the plasma membrane during the pachytene stage of meiotic prophase. Stage-specific antigens recognized by ALZ, including both protein and probably lipid, have been localized immunochemically on nitrocellulose blots from one-dimensional SDS gels. A dithiothreitol-sensitive constituent (Mr approximately 39,000) recognized by ALZ has been identified as the major protein determinant present in early meiotic cells but absent in 8-day-old seminiferous cell suspensions containing spermatogonia and Sertoli cells. This determinant is present in populations of preleptotene, leptotene/zygotene, and early pachytene spermatocytes isolated from 17-day-old animals, an observation consistent with the hypothesis of delayed insertion into the plasma membrane.  相似文献   

11.
Germ cells and Sertoli and Leydig cell functions were studied from 7 to 180 days after an acute exposure of 2-month-old rat testes to 9 Gy of gamma rays. Body weight, testis and epididymal weights were recorded. Sertoli cell parameters (androgen-binding protein, ABP, in caput epididymis and plasma follicle stimulating hormone, FSH) and Leydig cell parameters (plasma luteinizing hormone, LH, testosterone and prostate and seminal vesicle weights) were determined together with the number of germ cells and Sertoli cells. Irradiation did not affect body weight but significantly reduced testicular and epididymal weights from day 7 and day 15 post-irradiation respectively. The cells killed by irradiation were mainly spermatogonia and preleptotene spermatocytes engaged in replicating their DNA at the time of exposure, but all spermatocytes seemed damaged as they gave abnormal descendent cells. By day 34, only elongated spermatids remained in a few tubules and thereafter very little regeneration of the seminiferous epithelium occurred, except for one rat which showed a better regeneration. Levels of ABP decreased by day 15 when the germ cell depletion had reached the pachytene spermatocytes, whereas FSH and LH levels rose when the number of elongated spermatids decreased. Levels of testosterone and the weight of the seminal vesicles did not change; occasionally, the prostate weight was slightly reduced. These results support our hypothesis that pachytene spermatocytes and elongated spermatids are involved in influencing some aspects of Sertoli cell function in the adult rat.  相似文献   

12.
13.
Androgens are especially important for the maintenance of spermatogenesis in adulthood and the experimental withdrawal of testosterone (T) production by ethane dimenthanesulfonate (EDS) is a valuable tool for studying androgen-dependent events of spermatogenesis. The aim of the present study was to investigate the specific changes in immunoexpression of androgen receptor (AR) in the testis in relation to degeneration and regeneration of Leydig cell (LC) population and seminiferous epithelium. Immunohistochemistry for AR and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) as well as TUNEL assay for apoptosis were performed on testicular sections of control and EDS-treated rats. Serum LH and T levels were measured by RIA. Our results revealed a total loss of AR immunoexpression from the nuclei of Sertoli (SCs), LCs and peritubular cells during the first week after EDS administration and that coincided with severe drop in T levels. Two weeks after EDS administration, the AR expression was recovered in these cells but normal stage-specificity in SCs was replaced by uniform intensity of AR immunostaining at all the stages of the spermatogenic cycle. The stage-specific pattern of androgen expression in SCs with a maximum at stages VII-VIII appeared 5 weeks after treatment. LC immunoreactivity for 3beta-HSD at different time points after EDS administration correlated with values of T concentration. The maximal germ cell apoptosis on day 7 was followed by total loss of elongated spermatids 2 weeks after EDS treatment. Regeneration of seminiferous epithelium 3 weeks after EDS administration and onwards occurred in tandem with the development of new LC population indicated by the appearance of 3beta-HSD-positive cells and gradual increase in T production. The specific changes in AR after EDS including their loss and recovery in Sertoli cells paralleled with degenerative and regenerative events in Leydig and germ cell populations, confirming close functional relationship between Sertoli, Leydig and germ cells.  相似文献   

14.
A germ cell nuclear antigen with approximately 44-kDa molecular weight was identified by a novel monoclonal antibody designated as Mab 2F2 from the library we have accumulated against rat testicular cells. In immature 20-day-old and adult rat testis the recognized antigen was expressed in the nuclei of early meiotic cells from preleptotene to early pachytene spermatocytes exhibiting a stage-specific appearance in the cycle of the seminiferous epithelium. The immunoreactivity was clearly associated with the meiotic chromosomes. The antigen was not detected in the late pachytene spermatocytes and more advanced stages of spermatogenesis. No labeling was observed in spermatogonia and somatic Sertoli and Leydig cells. The pattern of expression of the recognized antigen during early meiotic stages of spermatogenesis but not in mitotically dividing spermatogonia could strengthen its possible role in meiotic division.  相似文献   

15.
Ubiquitin carboxyl-terminal hydrolase 1 (UCH-L1) can be detected in mouse testicular germ cells, mainly spermatogonia and somatic Sertoli cells, but its physiological role is unknown. We show that transgenic (Tg) mice overexpressing EF1alpha promoter-driven UCH-L1 in the testis are sterile due to a block during spermatogenesis at an early stage (pachytene) of meiosis. Interestingly, almost all spermatogonia and Sertoli cells expressing excess UCH-L1, but little PCNA (proliferating cell nuclear antigen), showed no morphological signs of apoptosis or TUNEL-positive staining. Rather, germ cell apoptosis was mainly detected in primary spermatocytes having weak or negative UCH-L1 expression but strong PCNA expression. These data suggest that overexpression of UCH-L1 affects spermatogenesis during meiosis and, in particular, induces apoptosis in primary spermatocytes. In addition to results of caspases-3 upregulation and Bcl-2 downregulation, excess UCH-L1 influenced the distribution of PCNA, suggesting a specific role for UCH-L1 in the processes of mitotic proliferation and differentiation of spermatogonial stem cells during spermatogenesis.  相似文献   

16.
17.
Bcl-w, a prosurvival member of the Bcl-2 family, is essential for spermatogenesis. However, the mechanisms by which Bcl-w participates in the regulation of apoptosis in the testis are largely unknown. To explore the potential role of Bcl-w in the regulation of apoptosis in the testis, the expression of Bcl-w mRNA and protein during testicular development and spermatogenesis, the dimerization with the proapoptosis members of the Bcl-2 family, and the responses to hormonal stimulation in vitro and apoptosis-inducing signals in vivo were investigated. Both Bcl-w mRNA and protein were detected in Sertoli cells, spermatogonia, and spermatocytes, as well as in Leydig cells. The steady-state levels of Bcl-w mRNA and protein were much higher in Sertoli cells than in spermatogonia and spermatocytes. In the adult rat testis, both Bcl-w mRNA and protein in Sertoli cells displayed a stage-specific expression pattern. Bcl-w could form complexes with Bax and Bak but not with Bad. Bax and Bak were immunohistochemically localized to the same cell types as Bcl-w, but with higher expression levels in spermatocytes and spermatogonia than in Sertoli cells. FSH could up-regulate Bcl-w mRNA levels in the seminiferous tubules cultured in vitro, whereas no effect was observed when testosterone was applied. Three animal models that display spermatogonial apoptosis induced by blockade of stem cell factor/c-kit interaction by a function-blocking anti-c-kit antibody, spermatocyte apoptosis induced by methoxyacetic acid, and apoptosis of spermatogonia, spermatocytes, and spermatids induced by testosterone withdrawal after ethylene dimethane sulfonate treatment were employed to check the changes of Bcl-w, Bax, and Bak protein levels during apoptosis of specific germ cells. In all three models, the ratios of Bax/Bcl-w and Bak/Bcl-w were significantly elevated. The present study suggests that Bcl-w is an important prosurvival factor of Sertoli cells, spermatogonia, and spermatocytes and participates in the regulation of apoptosis by binding proapoptotic factors Bax and Bak. The ratios of Bax/Bcl-w and Bak/Bcl-w may be decisive for the survival of Sertoli cells, spermatogonia, and spermatocytes.  相似文献   

18.
19.
PERF 15 is a testicular germ cell specific fatty acid-binding protein (FABP) isolated from rat. Indirect immunofluorescent analysis of juvenile rat testis showed that there were some strongly PERF 15-positive spermatocytes. These cells showed unclear nuclear structure and were predicted to undergo apoptosis. Apoptosis in germ cells is an important regulatory event to limit the number of germ cells in the seminiferous epithelium, but the physiological significance and molecular mechanisms of this testicular germ cell apoptosis are poorly understood. To determine whether PERF 15 participates in germ cell apoptosis, juvenile rat testis was examined by immunohistochemical and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) methods. Strongly PERF 15-positive cells and TUNEL-positive cells were co-localized in adjacent sections. Exposure to methoxyacetic acid (MAA), known to induce apoptosis in spermatocytes, increased the number of strongly PERF 15-positive cells in 25-day-old rats' testes. Therefore, it seems that PERF 15 is involved in both spermatogenesis and testicular germ cell apoptosis.  相似文献   

20.
Isolated rat pachytene spermatocytes were incubated in chemically defined medium supplemented with pyruvate and lactate, which are known to be essential energy substrates for these germ cells. Protein synthesis by the isolated cells was investigated by means of two-dimensional polyacrylamide gel electrophoresis. The electrophoretic patterns of (35S)-labeled proteins, synthesized by the pachytene spermatocytes during incubation in the presence of (35S)methionine either from 0-2 h or from 24-26 h after isolation, were almost completely identical. The patterns of newly synthesized proteins of freshly isolated spermatocytes and spermatids, however, showed several stage-specific proteins in addition to many proteins common to both spermatogenic cell types. Hence, it was concluded that a stage-specific pattern of protein synthesis can be maintained by pachytene spermatocytes during incubation for a period of 24 h in the absence of Sertoli cells but in the presence of a proper energy source.  相似文献   

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