Effect of N-acetylcysteine and deferoxamine on endogenous antioxidant defense system gene expression in a rat hepatocyte model of cocaine cytotoxicity

In the present study we investigated on cultures of hepatocytes from phenob arbital-pretreated rats, the effect of the antioxidants, 0.5 mM N-acetylcys teine (NAC) or 1.5 mM deferoxamine (DFO), previously incubated for 24 h and coincubated with cocaine (0-1000 mu M) for another 24 h. Cocaine cytotoxic ity was monitored by either the lysis of the cell membranes or apoptosis. L ysis of the cell membranes was evidenced by lactate dehydrogenase leakage, apoptosis was observed by detecting a hypodiploid peak (< 2C) in DNA histog rams obtained by flow cytometry, peroxide production was quantified with 2' ,7'-dichlorodihydrofluorescein diacetate and gene expression of the antioxi dant enzymes: Mn- and Cu,Zn-superoxide dismutases, catalase and glutathione peroxidase were measured by Northern blot analysis. NAC and DFO significan tly decreased the extent of lysis of cell membranes and apoptosis, and the antiapoptotic effect was parallel to peroxide generation. By the effect of NAC and DFO, significant increases were detected in the levels of mRNA of c atalase, manganese superoxide dismutase and glutathione peroxidase. From th ese results we conclude that NAC or DFO, when incubated in the presence of cocaine, exerted a protective effect against cocaine toxicity at the level of both lysis of the membranes and apoptosis. This protective effect, in th e case of NAG, was directed towards an increase in antioxidant enzyme expre ssion, and in the case of DFO against reactive oxygen species generation. ( C) 2000 Elsevier Science B.V. All rights reserved.