Activator protein-1 in human male germ cell apoptosis.

Apoptosis limits germ cell number in the testis, and its dysregulation is associated with male infertility. Here, we evaluated the role of the transcription factor activator protein 1 (AP-1) in male germ cell apoptosis in a culture of human seminiferous tubules. AP-1 DNA-binding activity increased in cultured tubules within 2.5 h, which was earlier than the onset of apoptosis as detected by caspase 3 activation and apoptotic DNA fragmentation. The c-Jun, c-Fos and JunD proteins were detected in the Sertoli cell nuclei, whereas apoptosis occurred in the germ cells. Follicle-stimulating hormone (FSH), whose receptors are expressed in the Sertoli cells, inhibited germ cell apoptosis and concomitantly suppressed AP-1 DNA-binding activity, but had no effect on nuclear factor kappaB (NF-kappaB) activation. These results suggest that AP-1 transcription factors are involved in the Sertoli cell-mediated control of germ cell apoptosis, and that inhibition of germ cell apoptosis by FSH appears to involve suppression of AP-1 activation.

Transcription factor GATA-6, cell proliferation, apoptosis, and apoptosis-related proteins Bcl-2 and Bax in human fetal testis.

The GATA family of transcription factors have been implicated in regulating the development and function of many organs. Furthermore, they have been linked to signaling cascades regulating cell fate through apoptosis. GATA-6 has been shown to be expressed in the gonads, but its cell-specific expression in the testis has remained unclear. We have studied GATA-6 expression in human fetal testis using in situ hybridization and immunohistochemistry and compared these results with the expression of the apoptosis-related proteins Bcl-2 and Bax. Furthermore, apoptosis was studied by thymidine deoxyribose-mediated deoxy-UTP nick end labeling assay, and cell proliferation by Ki-67 immunohistochemistry. GATA-6 mRNA and protein were expressed in Sertoli and Leydig cells early in gestation. Apoptotic cells were scanty between wk 16 and 40, and proliferation significantly ceased during the third trimester, supporting the view that only a little tissue remodeling occurs in the late fetal testis. Bax was present throughout the fetal period, whereas Bcl-2 expression decreased toward term. Neither of these factors correlated to the extent of apoptosis, and thus their role in the regulation of apoptosis in the fetal testis remains open. Despite strong expression, GATA-6 did not correlate with apoptosis or cell proliferation and is therefore unlikely to be directly involved in these processes in the human fetal testis.

Developmental expression and spermatogenic stage specificity of transcription factors GATA-1 and GATA-4 and their cofactors FOG-1 and FOG-2 in the mouse testis.

OBJECTIVE: The transcription factors GATA-1 and GATA-4 have been implicated in the regulation of testicular development and function. Their cofactors FOG-1 and FOG-2 are expressed in the gonads, but their cell-specific and developmental expression in the testis remains unresolved. Therefore, we analyzed GATA-1, GATA-4, FOG-1 and FOG-2 expression in detail, from undifferentiated male urogenital ridge to adult testis. METHODS: Immunohistochemistry and in situ hybridization were applied on mouse testicular samples. RESULTS: GATA-4 and FOG-2, but not GATA-1 or FOG-1, were expressed as early as in the male urogenital ridge. FOG-2 expression was localized in the Sertoli cells at embryonal day 12.5 (E12.5), but it diminished with advancing fetal testicular development. In E17.5 testis, FOG-2 was present only in the testicular capsule and a subset of fetal Leydig cells. FOG-1 was expressed from E15.5 Sertoli cells onwards, whereas GATA-1 was not detected during the fetal period at all. In the postnatal testis, FOG-2 was abundantly expressed immediately after birth, but in adult testis its expression was predominantly restricted to stage VII-XII seminiferous tubules. Stage specificity was also found for FOG-1, which, similarly to GATA-1, was abundantly expressed in stage VII-XII tubules during adulthood. CONCLUSIONS: Our results indicate that FOG-2, in addition to GATA-4, has a role in early gonadal development and sexual differentiation, and FOG-1 at later fetal stages, while GATA-1 executes its action postnatally. The findings suggest that, in contrast to the hematopoietic system and the heart, GATA-1 and GATA-4 do not use FOG-1 and FOG-2 respectively as their only cofactors during the early stages of testicular development.

Expression of nephrin in pediatric kidney diseases.

Nephrin is a podocyte cell adhesion protein located at the slit diaphragm area of the kidney glomerulus. Mutations in the nephrin gene (NPHS1) lead to congenital nephrosis, suggesting that nephrin is essential for the glomerular filtration barrier. This prompted this study of the expression of nephrin in acquired pediatric kidney diseases using in situ hybridization and immunohistochemistry. In situ hybridization for nephrin mRNA was performed in biopsy samples from patients with proteinuria caused by minimal change nephrosis, focal segmental glomerulosclerosis, and membranous nephropathy. The expression of nephrin mRNA was evaluated by grading the signal intensity visually and by counting the number of grains in separate glomeruli. No significant difference was observed in these samples as compared with controls. Immunostaining for nephrin was performed using antibodies directed against extra- and intracellular parts of the molecule. Nephrin staining gave a linear pattern along the glomerular capillary loops. In minimal change nephrosis, focal segmental glomerulosclerosis, and membranous nephropathy, the distribution of nephrin was similar to that in controls. In proliferative forms of glomerulonephritides (Henoch-Schönlein nephritis, IgA nephropathy, postinfectious and membranoproliferative glomerulonephritis), crescents and sclerotic lesions were negative for nephrin, and mesangial proliferation led to a scattered and sparse staining pattern. The staining pattern of nephrin was compared to that of ZO-1, a component of the cytoplasmic face of the slit diaphragm. The distributions of these two proteins in capillary tufts were similar in all disease entities studied. In conclusion, immunohistochemistry and in situ hybridization did not reveal major alterations in the expression of nephrin in proteinuric kidney diseases in children. Further studies are needed for more precise evaluation of the role of nephrin in these diseases.