Leydig cell apoptosis after the administration of ethane dimethanesulfonate to the adult male rat is a Fas-mediated process

Leydig cells undergo apoptosis in response to the cytotoxin ethane dimethan esulfonate (EDS), with numbers declining at 12-18 h and maximal apoptosis a t 24 h postinjection. The Bcl-2 family members, Bcl-2, Bcl-x1, and Bar, app ear not to be involved in this process. To further investigate this phenome na, a single dose of EDS was administered to adult rats to induce the killi ng of Leydig cells. The interstitial cells were examined up to 3 days after EDS administration by Western blot analysis for the Bcl-2 family members ( Bak and Bcl-w). Western blotting showed that Bak expression in the intersti tial cell preparations was unchanged after EDS, and immunohistochemistry sh owed that it was not up-regulated in Leydig cells in response to EDS. Bcl-w expression in the Leydig cells and interstitial cell preparations was unch anged until 48 h when it became undetectable, suggesting that Leydig cell-a ssociated Bcl-w is not involved in initiating apoptosis. We also investigat ed the role of the Fas system in Leydig cell apoptosis. Both Fas receptor a nd Fas ligand protein levels increased after EDS, peaking at 12-18 h and de clining thereafter. Fas receptor and ligand were shown by immunohistochemis try to be present in Leydig cells, and after EDS all Leydig cells became st rongly positive for both proteins. The intensity of staining increased in t he early stages of apoptosis and decreased as the nuclear morphology became more fragmented. These data suggest that Bcl-2 family members are not invo lved in Leydig cell apoptosis after EDS administration. However, up-regulat ion of the Fas system does occur, implicating activation of Fas receptor in the induction of Leydig cell apoptosis.