Quantitative and qualitative studies of micronucleus induction in mouse erythrocytes using flow cytometry. I. Measurement of micronucleus induction in peripheral blood polychromatic erythrocytes by chemicals with known and suspected genotoxicity.

Quantitative and qualitative aspects of the in vivo micronucleus-inducing potential of five chemicals were studied using flow cytometric enumeration of micronucleated polychromatic peripheral blood erythrocytes in mice. The chemicals were hydroquinone, vinblastine sulphate, chloral hydrate (tested in two different mouse strains), 5-bromo-2-deoxyuridine and 2-chlorobenzylidene malonitrile. Repeat samplings of peripheral blood were made at 0, 24, 40, 48 and 72 h and for low doses of 5-bromo-2-deoxyuridine 96 h after i.p. treatment. The agents hydroquinone (lowest effective dose 25 mg/kg), vinblastine sulphate (lowest effective dose 0.05 mg/kg) and 5-bromo-2-deoxyuridine (lowest effective dose 200 mg/kg) gave rise to significant increases in the frequencies of micronucleated polychromatic erythrocytes. No significant induction of micronucleated polychromatic erythrocytes by 2-chlorobenzylidene malonitrile or chloral hydrate was found. The frequencies of induced micronucleated polychromatic erythrocytes peaked at 40 h after hydroquinone treatment, at 48 h after vinblastine treatment and at 72 h after 5-bromo-2-deoxyuridine treatment with evident dose-dependent differences in the kinetics of the induction of micronucleated polychromatic erythrocytes. The mean relative Hoechst 33342 fluorescence of the populations of induced micronucleated polychromatic erythrocytes was used as an indicator of the DNA content of induced micronuclei. These values were found to be in agreement with the presumed mechanisms of micronucleus induction for hydroquinone, vinblastine sulphate and 5-bromo-2-deoxyuridine. Flow cytometric enumeration of micronucleated polychromatic erythrocytes in peripheral blood is an efficient method for the study of in vivo micronucleus induction, combining rapid analysis and high sensitivity with information on possible mechanisms of micronucleus induction. The method also allows a substantial reduction in the number of animals needed.