Comparison of co-cultivation of V79 cells with rat hepatocytes and rat H4IIE hepatoma cells for studying nitrosamine-induced hprt gene mutations.

Abstract

The induction of mutations by nitrosamines in the hprt locus of V79 Chinese hamster cells was examined after metabolic activation in a co-cultivation system using either freshly isolated rat hepatocytes or H4IIE rat hepatoma cells and the results obtained were compared with systems which employ the rat liver microsomal fraction (S9-mix). This study was also designed as a first approach to investigating the induction of point mutations by tobacco-specific nitrosamines in mammalian cells in order to obtain information about the significance of these compounds in connection with the carcinogenicity of tobacco smoke. The mutagenicity of two tobacco-specific nitrosamines, 4-(methylnitroso)-1-(3-pyridol)-1-butanone (NNK) and N'-nitrosonornicotine (NNN), were investigated and compared to two extensively investigated nitrosamines, i.e. dimethylnitrosamine (DMN) and diethylnitrosamine (DEN). DMN was activated to mutagenic species by primary hepatocytes at mumolar concentrations, i.e. 1/100 of the concentrations required for mutagenesis by DEN and NNK. NNN was not activated to mutagenic species by liver S9 or primary hepatocytes. The findings shown here on the mutagenicities of NNK and NNN with liver preparations are in agreement with their relative carcinogenic potencies. When the established liver cell line H4IIE was used for metabolic activation, DMN and was found to be mutagenic, whereas the results for NNN were borderline and for DEN and NNK were without effect. The fate of these compounds via different metabolic pathways is discussed in terms of systems for detection of mutagenic metabolites and type of mutation induced.