Leydig cell-derived heme oxygenase-1 regulates apoptosis of premeiotic germ cells in response to stress

Stress-induced downregulation of spermatogenesis remains poorly understood. This study examined the induction of heme oxygenase-1 (HO-1), a carbon monoxide-generating inducible enzyme, in modulation of spermatogenesis. Rats were exposed to cadmium chloride (CdCl2), a stressor causing oligo-zoospermia, and HO-1-induction was monitored by following HO isozyme expression. CdCl2-treated testes increased HO-1 activity and suppressed microsomal cytochromes P450, which are required for steroidogenesis. CdCl2-elicited HO-1 occurred mostly in Leydig cells and coincided with CO generation, as judged by bilirubin-IXalpha immunoreactivity. Under these circumstances, germ cells in peripheral regions of seminiferous tubules exhibited apoptosis; laser flow cytometry revealed that these apoptotic cells involve diploid and tetraploid germ cells, suggesting involvement of spermatogonia and primary spermatocytes in CdCl2-elicited apoptosis. Pretreatmentwith zinc protoporphyrin-IX, an HO inhibitor, but not copper protoporphyrin-IX, which does not block the enzyme, attenuated the CdCl2-induced apoptosis. Such antiapoptotic effects of zinc protoporphyrin-IX were repressed by supplementation of dichloromethane, a CO donor. Upon CdCl2-treatment, both Sertoli cells and the germ cells upregulated Fas ligand; this event was also suppressed by zinc protoporphyrin-IX and restored by dichloromethane. Thus, Leydig cells appear to use HO-1-derived CO to trigger apoptosis of premeiotic germ cells and thereby modulate spermatogenesis under conditions of stress.