THE CONCOMITANT DETECTION OF GENE MUTATION AND MICRONUCLEUS INDUCTION BY MITOMYCIN-C INVIVO USING LACZ TRANSGENIC MICE

A new assay system that can simultaneously provide gene mutagenicity and clastogenicity data in vivo is described. Transgenic mice (Muta(TM) Mouse) harboring the lacZ gene as a target for mutation analysis were injected intraperitoneally with mitomycin C (MMC), either once or on 5 successive days. Micronucleus assays were performed with small amounts of peripheral blood collected from a tail vessel. The spontaneous frequency of micronucleated reticulocytes was 0.42%. For the mutation analysis, DNA was extracted from bone marrow and liver cells at several harvest times. The lacZ gene was rescued by lambda packaging and infection of E. coli C (lac-), followed by plating on agarose plates containing X-gal. The spontaneous lacZ mutant frequencies were 37 and 29 X 10(-6) in bone marrow and liver, respectively. In the micronucleus assay, single treatments with 1.0 and 2.0 mg/kg of MMC induced micronuclei in 3.6 and 5.8% of reticulocytes, respectively, peaking 48 h after treatment. Muta(TM)Mouse sensitivity to micronucleus induction was similar to nontransgenic strains used routinely for the micronucleus test. On the other hand, single treatments with MMC at 1.0 and 2.0 mg/kg did not induce any significant increases in the frequency of lacZ- mutants in bone marrow or liver. N-Ethyl-N-nitrosourea, used at 100 mg/kg as a positive control, yielded a 5-fold increase in mutant frequency above untreated animals in bone marrow only. After 5-day treatments, MMC induced approximately a 2-fold increase in mutant frequency in bone marrow only for the sublethal dose of 2 mg/kg. Therefore, this study indicated that the strong clastogenic activity of MMC in bone marrow was not accompanied by significant gene mutagenic activity.