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Mechanism by which ethane dimethanesulfonate kills adult rat Leydig cells: involvement of intracellular glutathione

, : Mechanism by which ethane dimethanesulfonate kills adult rat Leydig cells: involvement of intracellular glutathione. Toxicology and Applied Pharmacology 120(1): 80-88

We have previously demonstrated that ethane-1,2-dimethane-sulfonate (EDS) kills adult, but not immature, rat Leydig cells in vivo and in vitro. The mechanism responsible for this selective toxicity is not known. Here we report that the cytotoxic effects of EDS on adult rat Leydig cells were not dependent upon new protein synthesis or cytochrome P450 enzyme activity. To determine whether inhibition of glutathione synthesis protects Leydig cells from the cytotoxic effects of EDS, adult rat Leydig cells were cultured in the presence or absence of 4 mM buthionine sulfoximine (BSO; 2 hr), a specific inhibitor of glutathione synthesis, and subsequently with increasing doses of EDS (3 hr). Following EDS addition, the ability of the cells to produce testosterone in response to LH stimulation, and to synthesize protein ([35S]methionine incorporation) were evaluated. In both cases, Leydig cells cultured in the presence of BSO were far less sensitive than Leydig cells cultured in medium alone (control) to EDS effects on testosterone production (control: EC50 = 60 micrograms EDS/ml; BSO: EC50 > 1500 micrograms EDS/ml) and [35S]methionine incorporation (control: EC50 = 95 micrograms EDS/ml; BSO: EC50 = 1560 micrograms EDS/ml). This protective effect of BSO was abolished by restoring intracellular glutathione levels with glutathione ethyl ester (8 mM; GSHEE). Interestingly, none of these treatments altered the viability (i.e., [35S]methionine incorporation) of immature rat Leydig cells (EC50 = 420 micrograms EDS/ml for control, GSHEE, and BSO groups). Taken together, these data suggest that the mechanism by which EDS kills adult rat Leydig cells may involve Leydig cell glutathione.


PMID: 8390114

DOI: 10.1006/taap.1993.1089

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