Abstract
Germ cell death is very common in mammalian testis, during both early testicular development and in the adult period, however the molecular mechanisms underlying the induction of germ cell apoptosis in mouse testes are largely unknown. The aim of this study was to investigate the possible involvement of Bcl-2 and Bax in induction of germ cell apoptosis in immature and mature mouse testes. To gain insights into downstream molecular cascades, we examined the expression of cytochrome c and activated caspase-3. Testes of adult and neonatal mice at 0, 6, 12 and 18 days after birth were used. Germ cell apoptosis was evaluated by Terminal dUTP nick end-labeling (TUNEL) staining and the expression of apoptosis-related proteins was examined by enzyme-immunohistochemistry. TUNEL-positive germ cells appeared at 6 days after birth, reaching a maximum number at 12 days after birth, when such cells were identified as gonocytes or spermatogonia. In contrast, by 18 days, the main population of TUNEL-positive cells comprised spermatocytes and the average number of positive cells per seminiferous tubule remained nearly constant thereafter. The expression of Bcl-2 and Bax remained constant in germ cells from 0 to 12 days after birth. However, during that period we observed a dramatic redistribution of Bax staining from the cytoplasm to the whole cell including the nucleus. In parallel with the Bax redistribution, cytochrome c staining spread to the cytoplasm and activation of caspase-3 occurred. Moreover, in mirror sections of neonatal and adult testes, we confirmed a strong correlation among TUNEL-staining and Bax redistribution accompanying the cytochrome c redistribution and activation of caspase-3. The temporal and spatial association of Bax and spatial apoptosis-related protein expression with TUNEL-positive germ cells indicates that Bax redistribution may be a trigger for the induction of germ cell apoptosis in mouse testes.
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