Restriction site mutation (RSM) analysis of 2-acetylaminofluorene (2-AAF)-induced mouse liver mutations and comparison with the measurement of in vivo micronucleus induction in the bone marrows of (2-AAF)-treated mice

We report here the successful application of the restriction site mutation (RSM) assay in detecting 2-acetylaminofluorene (2-AAF)-induced mouse liver mutations. A total of seven 2-AAF-induced liver mutations were detected out of a total of 304 analyses performed on 2-AAF-treated liver tissue. No mut ations were detected in the 190 RSM analyses performed on untreated liver t issue. The 2-AAF-induced point mutations comprised 60% GC-->TA transversion s, 30% GC-->AT transitions, 10% GC-->CG transversions, and 1 insertional ev ent was also detected. All seven mutations were detected in intron 6 of the mouse p53 gene, with no mutations detectable in exons 4 or 5, supporting o ur previous data on the greater mutability of intron regions. In addition t o the RSM analysis, we also report the application of the in vivo bone marr ow micronucleus assay in detecting the clastogenicity of 2-AAF. We detected a small, but statistically significant, increase in the number of micronuc lei induced by 2-AAF, but only after 2,000 cells were scored. This also con firms previous data showing that 2-AAF is a weak clastogen. Finally, we att empted to compare the sensitivity of the two assays to 2-AAF-induced genoto xicity, as had been previously undertaken with ENU. Both assays detected ge notoxicity in their respective tissues; however, different endpoints were a nalysed. The RSM assay appears to be more adaptable than the micronucleus a ssay, due to its tissue and organism independence and has the potential to provide more molecular information on genotoxicity. (C) 2000 Wiley-Liss, In c.