Benzene is a widely recognized human and animal carcinogen. In spite of considerable research, relatively little is known about the genotoxic events that accompany benzene exposure in vivo. To gain insights into the mechanisms underlying the genotoxic effects of benzene, we have characterized the origin of the micronuclei that are formed in bone marrow erythrocytes and splenic lymphocytes of benzene-treated mice using two molecular cytogenetic approaches: (a) fluorescence in situ hybridization with a centromeric DNA probe; and (b) staining with the calcinosis-Raynaud's phenomenon-esophageal dismobility-sclerodactyly-telangiectasia syndrome of scleroderma (CREST) antibody, an antibody recognizing a centromeric protein. Following the p.o. administration of benzene (220 or 440 mg/kg) to male CD-1 mice, a significant increase in micronuclei was observed in the bone marrow erythrocytes. In situ hybridization with a centromeric DNA probe and immunofluorescent staining with the CREST antibody indicated that the micronuclei in bone marrow erythrocytes were formed from both chromosome loss and breakage. The majority of the micronuclei originated from chromosome breakage. A dose-related increase in micronucleated cells was also observed in splenocyte cultures established from these benzene-treated animals. In contrast to the bone marrow erythrocyte results, the majority of benzene-induced micronuclei in the cytokinesis-blocked splenocytes labeled with the CREST antibody indicating that these micronuclei were the result of whole chromosome loss. These data demonstrate that both aneuploidy and chromosomal breakage are early genotoxic events induced by benzene or its metabolites in vivo and also indicate that the nature of the chromosomal alterations may vary depending on the target organ or cell type.

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This work was supported by funds provided by the Cigarette and Tobacco Surtax Fund of the State of California through the Tobacco-related Disease Research Program of the University of California, Grant 1KT36.

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