Avaliação estrutural e funcional dos testículos de camundongos adultos controles e deficiente para iNOS, tratados ou não com propil-tiouracil durante o período pós-natal

Abstract

In rodents (mice and rats), Sertoli cells proliferate until 2 to 3 weeks after birth and it is well established in the literature that FSH and thyroid hormones (T3 and T4) are responsible, respectively, for Sertoli cells proliferation and differentiation. It is also known that transient hypothyroidism induced by PTU (propylthiouracil) treatment during the postnatal period extends the mitotic activity of Sertoli cells, leading to higher numbers of these cells per testis, as well as higher testis weight and daily sperm production. The nitric oxide (NO) is an important mediator of intra and extra cellular processes, playing therefore a crucial role in several physiological and pathological conditions, and some studies have shown that NO reduces testosterone production in vitro and in vivo. In the testis, constitutive expression of iNOS (inducible nitric oxide synthase) occurs in Leydig, Sertoli and germ cells, suggesting a regulatory function of this enzyme in spermatogenesis and steroidogenesis. In this context, recent studies from our laboratory showed that iNOS deficient mice presents higher Sertoli cell proliferation and lower germ cell apoptosis, leading also to higher daily sperm production. In these mice a higher number and lower individual volume of Leydig cell is observed. Our aims in the present study were to evaluate if the association between the neonatal hypothyroidism and the iNOS deficiency is able to promote additive effect in the total number of Sertoli cell per testis. We also aimed at characterizing, by immunohistochemistry and qPCR, the expression of important markers of testis function in adult wild type and iNOS-/- mice. As expected, our results showed higher numbers of Sertoli cell per testis in the wild type mice treated with PTU and in the iNOS-/- mice. However, surprisingly, no additive effect was observed for the number of these cells in the iNOS deficient mice treated with PTU. Suggesting that Sertoli cells could be functionally more active, in the second part of this study we observed higher expression of Sox9 and androgen receptor in these cells in the testis of iNOS-/- mice; and these findings could explain, at least in part, the lower germ cell apoptosis described in the literature for iNOS deficient mice. Regarding the steroidogenic pathway, the iNOS-/- mice showed lower levels of StAR and 3β-HSD, and this could be related with the smaller individual volume of the Leydig cells. Finally, the aromatase expression showed no difference between wild type and iNOS-/-, whereas the mRNA levels of estrogen receptor α were increased in the testis of iNOS deficient mice. In summary, our data suggest the existence of a possible compensatory mechanism to counterbalance the physiological changes caused by iNOS deficiency in mice. Furthermore, a possible interaction between estrogens and nitric oxide in the functional regulation of mice testis cannot be ruled out.