ABSTRACT
Detection of steroid hormone receptors within a target tissue is important for an understanding of their crucial role in regulating of steroids' action. In the light of recent knowledge on the role of estrogens in male gonads the efforts were undertaken to clarify and discuss a role of androgen receptors, aromatase and estrogen receptors (ER) in mediating testosterone and/or estradiol action in testicular cells of bank voles that were kept under short or long light cycles. Immunohistochemistry was performed on paraplast embedded sections of the bank vole testes. First, androgen receptors were immunolocalized in testicular somatic cells while germ cell did not express any immunoreaction. Moreover, the ability to convert androgens to estrogens by various testicular cells was documented; aromatase immunoexpression was found in testis sections, not only in Leydig cells and Sertoli cells but also in germ cells. Finally, the expression of estrogen receptor-alpha (ERalpha) was observed in Leydig cells whereas the presence of estrogen receptor-beta (ERbeta) was detected in Sertoli and germ cells, namely spermatocytes and spermatids. The cellular distribution of androgen receptors appeared to be light -and age-dependent in adults; immunoexpression of aromatase and ERbeta was found to be both age -and photoperiod-dependent in germ cells.
Subject(s)
Androgens/metabolism , Arvicolinae/metabolism , Estrogens/metabolism , Testis/metabolism , Animals , Aromatase/metabolism , Estrogen Receptor alpha , Estrogen Receptor beta , Immunohistochemistry , Leydig Cells/metabolism , Male , Photoperiod , Receptors, Androgen/metabolism , Receptors, Estrogen/metabolism , Sertoli Cells/metabolism , Spermatids/metabolism , Spermatocytes/metabolismABSTRACT
The Steroidogenic Acute Regulatory (StAR) protein is assumed to enhance the rate-limiting step of the steroid biosynthesis. Now, it is the most likely candidate, responsible for acutely regulating transfer of cholesterol from the outer to the inner mitochondrial membrane. In this study, the immunoreactive StAR protein was observed in the mitochondria of mouse cultured Leydig cells stimulated by hCG andtesticular macrophage-conditioned medium. Immunocytochemistry was performed using a polyclonal rabbit antibody against the StAR protein. For selective staining of mitochondria in Leydig cells, the Mito Tracker dye was used. Computerized, superimposed images from double-fluorescence staining showed a remarkable degree of similarity in the distribution of the StAR protein and mitochondria, indicating mitochondrial localization of StAR.
Subject(s)
Leydig Cells/metabolism , Mitochondria/metabolism , Phosphoproteins/metabolism , Animals , Chorionic Gonadotropin/pharmacology , Fluorescent Antibody Technique , Humans , Leydig Cells/chemistry , Male , Mice , Mitochondria/chemistryABSTRACT
Histochemistry for NADPH-diaphorase detects an enzymatic activity associated with nitric oxide synthase while immunohistochemistry detects the nitric oxide synthase molecule. NADPH-diaphorase and inducible isoform of nitric oxide synthase in Leydig cells in vitro and in testis sections of the bank vole were demonstrated histochemically and immunocytochemically. Histochemical studies revealed localization of NADPH-diaphorase reaction product in the cytoplasm of cultured Leydig cells as well as in the interstitial area, mainly in Leydig cells and in vascular endothelium. Distribution pattern of NADPH-diaphorase was different in Leydig cell cytoplasm of individual cells. Using immunocytochemistry, the immunoreactivity for nitric oxide synthase was observed both in cultured Leydig cells and testis sections. Moreover, a co-localization of positively immunostained cells with those histochemically detected was noticed. Addition of hCG to the cultured medium or injections in vivo resulted in a small decrease in reaction intensity in Leydig cells. Treatment with N omega-nitro-L-arginine methyl ester resulted in distinctly weaker reactivity of the enzymes studied which was correlated with a higher testosterone and estradiol levels in Leydig cells measured radioimmunologically. The results have indicated that nitric oxide synthase is able to act directly within the male gonad regulating androgen secretion by Leydig cells.
Subject(s)
Leydig Cells/enzymology , NADPH Dehydrogenase/analysis , Nitric Oxide Synthase/analysis , Animals , Antibodies , Arvicolinae , Cells, Cultured , Chorionic Gonadotropin/pharmacology , Enzyme Inhibitors/pharmacology , Histocytochemistry , Immunohistochemistry , In Vitro Techniques , Leydig Cells/cytology , Leydig Cells/metabolism , Male , NADPH Dehydrogenase/immunology , NG-Nitroarginine Methyl Ester/pharmacology , Nitric Oxide Synthase/immunology , Nitric Oxide Synthase Type II , Nitroarginine/pharmacology , Radioimmunoassay , Testis/cytology , Testis/drug effects , Testis/metabolism , Testosterone/metabolismABSTRACT
In the testis, androgen receptors are known to mediate autocrine and paracrine effects of androgens on Leydig cell function and spermatogenesis. The pig presents some unusual features with regard to the synthesis of testosterone and estrogens in the male gonads. In testes from prepubertal males, testosterone level was lower than in testes from adult boars, while estrogen secretion was relatively high and comparable to that of mature porcine gonad. Immunolocalization of androgen receptors and intensity of immunohistochemical staining was age-dependent. In testis sections from adult boars, androgen receptors were found in nuclei of all somatic cells such as Leydig cells, Sertoli cells, and peritubular-myoid cells, whereas in sections from immature pigs only in the Leydig cell cytoplasm showed positive immunoreaction for androgen receptors. In control tissue sections incubated with omission of the primary antibody, no positive staining was observed. Detection of the androgen receptors in testicular cells of the pig is important for understanding of their central role in mediating androgen action.
