A co-cultivation system as a model for in vitro studies of modulating effects of naturally occurring indoles on the genotoxicity of model compounds

Abstract

An in vitro model system, consisting of primary chick embryo hepatocytes as the metabolizing system co-cultured with V79 Chinese hamster cells as the target cells for the SCE assay, was used to study the modulating effects of naturally occurring indoles on the genotoxicity of benzo[ a]pyrene (B[ a]P) and N-nitrosodimethylamine (NDMA). At 25 μg/ml, indole-3-carbinol (I3C) induced ethoxyresorufin- O-deethylase (ERROD) activity 3.9-fold and ethoxycoumarin- O-deethylase (ETCO) activity 2-fold. Indole-3-acetonitrile (I3A; 35 μg/ml) induced ERROD activity 1.6-fold and ETCO activity 2-fold. Pretreatment with I3C resulted in a 20–40% decrease in genotoxicity for both B[ a]P and NDMA. Pretreatment with I3A resulted in a similar decrease in genotoxicity for B[ a]P, whereas no decrease was observed for NDMA. When a microsomal fraction prepared from I3C-pretreated hepatocytes was used, no decrease in B[ a]P-induced genotoxicity was found; in contrast, use of an S-9 fraction prepared from the same cells resulted in a 45% decrease. Exposure of cells to B[ a]P after pretreatment with I3C for suboptimal times also resulted in a significant decrease in genotoxic activity. The data indicate that the protective effects of pretreatment with the indoles, observed in this study, are not directly correlated with induction of cytochrome P-450-associated enzymes. Instead they seem to be more the result of a changed balance in the enzyme systems involved in the indole biotransformation process and point to an essential role of conjugating enzyme systems.