Modulation of genotoxic and cytotoxic effects of aromatic amines in monolayers of rat hepatocytes

Cultured rat hepatocytes exposed to 2-acetylaminofl uorene (AAF), 2-aminofl uorene (AF) or N-hydroxy-2-acetylaminofluorene (N-OH-AFF) for 3 hrs resulted in an increase in DNA repair measured as unscheduled DNA synthesis, with N-OH-AAF > AAF > AF. Cytotoxic effects were only seen with N-OH-AAF above 10^–6 M. -Naphthof avone increased the unscheduled DNA synthesis and cytotoxic effects of N-OH-AAF, whereas it decreased DNA repair and the covalent binding of AAF to cellular proteins. In contrast, very little effects of paraoxon were seen on the repair synthesis elicited by AAF, AF or N-OH-AAF. The addition of ascorbate reduced the covalent binding of AAF, the DNA repair synthesis caused by AAF and N-OH-AAF, and the cytotoxic effects of N-OH-AAF. The addition of pentachlorophenol or salicylamide all resulted in similar effects as ascorbate, through reduction of sulfation. Galactosamine, an inhibitor of glucuronidation, and the nucleophile GSH caused no or only minor effects of the activation of AAF, AF or N-OH-AAF as judged from the endpoints tested. These results are consistent with an arylnitrenium ion, a sulfate ester or a free radical as the arylamine metabolite causing cellular DNA damage, whereas the sulfate ester or a radical intermediate may be responsible for the cytotoxic effects of N-OH-AAF.Abbreviations AAF 2-acetylaminofluorene - AF 2-aminofluorene - N-OH-AAF N-hydroxy-2-acetylaminofluorene - cytochrome P-450 a collective term for all forms of the cytochrome P-450 polysubstrate monooxygenase - DMSO dimethyl sulfoxide - HU hydroxyurea - S-9 9000 g supernatants - LDH lactate dehydrogenase - UDS unscheduled DNA synthesis - ANF -naphthoflavone - GSH glutathione - PCP pentachlorophenol - MET metyrapone - PAR paraoxon - DEM dimethylmaleate