We are investigating chromosomal lesions that may lead to chronic diseases and asked whether micronucleus induction is an early event leading to chromosomal mutation. We examined aneugens and clastogens and compared the induction of micronuclei with mutation induction in L5178Y mouse cells. Our results indicated that aneugens, although they induced micronuclei containing whole chromosomes, were unable to induce TFT resistance. We tentatively concluded that the induction of micronuclei containing whole chromosomes is not an early event leading to mutations in these cells. The clastogens examined thus far induce both micronuclei and mutation. To examine the nature of the genetic lesions, we used informative microsatellites that span the entire chromosome 11 and are polymorphic in these cells. We examined 122 spontaneous mutants. 36 showed no LOH at any of the loci and possibly harbor point mutations. The lesions found in remaining 86 were large. Our data showed that small and large colonies cannot predict the size of the lesion and cannot be used to predict clastogenicity or point mutagenicity respectively. We gained even more information concerning the nature of the lesions by coupling LOH analysis with chromosome paint analysis. We determined that recombination is responsible for many of the spontaneous mutants. These findings show that the L5178Y mouse cells may be an appropriate tool for in vitro measurement of recombination. The data are also inconsistent with the existence of a putative growth gene proximal to the tk+ allele though it may be distal. 5-azacytidine, one of the clastogens, induced micronuclei at concentrations at which it is mutagenic at the tk locus and that the micronuclei contained mostly chromosomal fragments. Chromatid bridges were observed in anaphase and, in some cases, were sustained into interphase. 5-Azacytidine is mutagenic to AS52 cells and over half of the 149 mutants that we examined have no mutations in the coding region of the gpt gene. We examined four azacytidine analogs for micronucleus and mutation induction. All four compounds induced micronuclei containing chromosomal fragments. There was a general trend in the biological activity of these analogs: compounds that are specifically blocked at the 5 position such as 5-azacytidine and 5-fluoro-2e-deoxycytidine effect changes in cell morphology, cytotoxicity, TFT resistance and the induction of micronuclei at very low doses. 5-Azacytidine analogs that possess more chemically accessible 5 positions such as 5,6-dihydro-5-azacytidine and 6-azacytidine either require doses that are orders of magnitude greater to induce these effects or are unable to induce changes in cell morphology and TFT resistance at doses below which the compound is lethal to the cells.
