Effect of antioxidants on para-aminophenol-induced toxicity in LLC-PK1 cells

The present studies were designed to investigate the susceptibility of LLC- PK1 cells to cytotoxicity induced by para-aminophenol (PAP) and the ability of antioxidants to prevent PAP-induced cytotoxicity. LLC-PK1 cells were in cubated for 4 h with varying concentrations of PAP (00.2 mM). Incubation wa s continued for 20 h and viability was monitored at 24 h after initial expo sure to PAP. For coincubation experiments, cells were incubated for 4 h wit h various antioxidants [including ascorbate, glutathione (GSH), butylated h ydroxytoluene (BHT), beta -nicotinamide adenine dinucleotide (NADH), or bet a -nicotinamide adenine dinucleotide phosphate (NADPH)] in the absence or p resence of 0.1 mM PAP. For preincubation experiments, cells were incubated for 1 h with ascorbate, GSH or NADPH. Antioxidants were removed and cells w ere exposed to 0 or 0.1 mM PAP for 4 h. Viability was determined 24 h follo wing PAP exposure. LLC-PK1 cells displayed a steep concentration-response r elationship for PAP: 0.1 mM PAP caused similar to 50% loss of viability. Co incubation with ascorbate, GSH and NADPH was without effect on cell viabili ty in the absence of PAP and attenuated PAP-induced losses in viability. In contrast. NADH was ineffective in preventing PAP-induced cytotoxicity. BHT alone produced a significant loss of cell viability and was ineffective in preventing PAP cytotoxicity. Inability of NADH to prevent PAP-induced cyto toxicity was related to rapid degradation of NADH in aqueous solution. Prei ncubation of cells with ascorbate or GSH but not NADPH was associated with attenuation of PAP-induced cytotoxicity. These data suggest that (1) FAP is cytotoxic to LLC-PK1 cells, (2) a portion of PAP cytotoxicity is due to no nenzymatic oxidation that occurs in the incubation medium, and (3) a portio n of PAP cytotoxicity is due to enzymatic or nonenzymatic oxidation that oc curs: within cells. (C) 2000 Elsevier Science Ireland Ltd. All rights reser ved.