DETECTION OF HYPERDIPLOIDY AND CHROMOSOME BREAKAGE IN INTERPHASE HUMAN-LYMPHOCYTES FOLLOWING EXPOSURE TO THE BENZENE METABOLITE HYDROQUINONE USING MULTICOLOR FLUORESCENCE IN-SITU HYBRIDIZATION WITH DNA PROBES

Increased frequencies of structural and numerical chromosomal aberrati ons have been observed in the lymphocytes of benzene-exposed workers. Similar aberrations occurring in bone-marrow cells may contribute to t he increased incidence of leukemia seen in these populations. Fluoresc ence in situ hybridization with chromosome-specific DNA probes is a re latively new technique which shows promise for the identification of a neuploidy-inducing agents. In these studies, fluorescence in situ hybr idization with several chromosome-specific DNA probes was used to inve stigate the ability of the benzene metabolite hydroquinone to induce h yperdiploidy in interphase human lymphocytes. Using a classical satell ite probe specific for human chromosome 9, a significant dose-related increase in the frequency of cells containing 3 or more hybridization regions was observed following the in vitro exposure of lymphocytes to hydroquinone at concentrations from 75 to 150 mu M. At the 100-mu M c oncentration of hydroquinone, the frequency of nuclei containing 3 or more hybridization regions was determined using probes for chromosomes I, 7 and 9. Significantly higher frequencies of affected nuclei were observed using the chromosome 1 and 9 probes when compared to the chro mosome 7 probe. To establish whether this difference was due to the no nrandom involvement of these chromosomes in hydroquinone-induced hyper diploidy or to chromosomal breakage within the chromosomal region targ eted by these probes, a multicolor fluorescence in situ hybridization approach was developed using probes to two adjacent regions on chromos ome 1. Using this tandem-labeling approach, the frequency of nuclei wi th multiple hybridization regions and the origin of the regions was de termined by scoring slides labeled simultaneously with the chromosome 7 alpha satellite probe and the adjacent alpha and classical satellite probes for chromosome 1. The results of these studies confirmed that hydroquinone exposure resulted in a significant increase in hyperdiplo id nuclei, but indicated that the different frequency of nuclei contai ning 3 or more hybridization regions observed using the chromosome 1 a nd 7 probes, was due to breakage within the chromosomal region targete d by the chromosome 1 classical satellite probe. These results indicat e that hydroquinone may contribute significantly to the numerical and structural aberrations observed in benzene-exposed workers. In additio n, the multicolor fluorescence in situ hybridization approach utilized in these studies promises to be a powerful technique for the detectio n of chromosomal breakage occurring in interphase human cells.