MEASUREMENT AND CHARACTERIZATION OF MICRONUCLEI IN CULTURED PRIMARY LUNG-CELLS OF MICE FOLLOWING INHALATION EXPOSURE TO BENZENE

The genotoxic effects of benzene in lung cells of mice exposed to sing le acute doses by inhalation have been estimated by cytogenetic analys is of micronuclei in primary cultures of lung fibroblasts, Mice were n ose-only exposed to 1000 p.p.m. for 30 or 60 min or to 3500 p.p.m. for 30 min and sacrificed 24 h after the end of exposure. Lung fibroblast s were cultured attached to coverslips for 72 h, the last 48 h in the presence of 0.75 mu g/ml cytochalasin B, Micronuclei were scored in bi nucleate cells. The mechanism(s) of micronucleus induction was charact erized by immunofluorescent staining of kinetochore proteins (CREST st aining), which allowed micronuclei due to chromosome loss (kinetochore -positive) to be distinguished from those produced by chromosome break age (kinetochore-negative). Three- and 4-fold statistically significan t increases in total micronucleus frequencies were observed in all ben zene-exposed mice with respect to unexposed controls. The effect was n either concentration nor time dependent. This is compatible,vith a pla teau dose-effect relationship for the effects on bone marrow, which is explained by saturation of metabolism. Both chromosome loss and chrom osome breakage appear to contribute to micronucleus formation, suggest ing that in addition to chromosome rearrangements, aneuploidy may be a relevant early genotoxic event associated with benzene carcinogenicit y, Under the same treatment conditions no micronucleus induction could be shown in spleen lymphocytes, suggesting that with very short benze ne exposures cells at the first contact site with local metabolizing c apacity have a higher probability of genetic alterations potentially l eading to neoplasia.