Our previous studies suggested that combined low-dose gossypol with steroid hormones possesses a reversible antifertility role on adult male rat. Spermatocyte apoptosis during the meiosis of spermatogenesis is the main reason for the antifertility. This study evaluates the effects of the regimen on the mitosis of spermatogenesis. Thirty-two adult male rats were divided into four groups randomly, Group GH: rats were fed orally with gossypol acetic acid (GA, 12.5 mg/kg d) and desogestrel (DSG, 0.125 mg/kg d)/ethinylestradiol (EE, 0.025 mg/kg d)/testosterone undecanoate (TU, 100 mg/kg d); Group G: a single dose of GA (12.5 mg/kg d); Group H: the same dosage of DSG/EE/TU as in Group GH; Group C: rats were treated with vehicle (1% methyl cellulose) as control. Testes were removed at the 8th week after treatment to assess the weight, volumes, diameter, and degenerated rate of seminiferous tubule, spermatogonia is quantitated by stereological assay. TUNEL assay and proliferating cell nuclear antigen immunohistochemistry were used to determine the germ cell apoptosis and proliferation. The changes of gene expression of glial cell line-derived neurotrophic factor (GDNF) and Ets variant gene 5 (Etv5) were analyzed. The results manifested that the spermatogenesis was suppressed, the type A and intermediate spermatogonia reduced, and spermatocyte and spermatid apoptosis increased both in Group GH and Group H. The expressions of GDNF protein in these two groups were downregulated, but no change was observed in Etv5 expression. However, the numbers of spermatogonia, germ cell apoptosis and proliferation, and the expressions of GDNF and Etv5 were unchanged in Group G compared with Group C. Above all, we concluded that the mitotic phase of spermatogenesis is suppressed by the combination regimen, it might be the steroid hormones in the regimen that caused the reduction of spermatogonia through the downregulation of GDNF.
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