GSTT1-DEPENDENT INDUCTION OF CENTROMERE-NEGATIVE AND CENTROMERE-POSITIVE MICRONUCLEI BY 1,2 3,4-DIEPOXYBUTANE IN CULTURED HUMAN-LYMPHOCYTES/

The role of the glutathione S-transferase T1 gene (GSTT1) in determini ng genotoxic response to 1,2:3,4-diepoxybutane (DEB), an epoxide metab olite of 1,3-butadiene, was studied by analysis of micronuclei (MN) in cultured human lymphocytes using the cytokinesis block method, Fluore scence in situ hybridization (FISH) with an alphoid satellite DNA prob e specific for the centromeres of all human chromosomes was applied to identify MN harboring whole chromosomes, Whole-blood lymphocyte cultu res of 11 GSTM1 (glutathione S-transferase M1)-positive individuals ha ving at least one GSTM1 allele), of whom six were GSTT1-positive (with at least one GSTT1 allele) and five GSTT1-null (GSTT1 homozygously de leted), were treated for 48 h (starting 24 h after culture initiation) with two different concentrations (2 and 5 mM) of DEB, The GSTT1-null individuals were excessively sensitive to DEB, showing, on average, s imilar to 2.5 times higher induced MN frequency (control frequency sub tracted) than the GSTT1-positive donors, both at 2 mM (mean/1000 binuc leate cells 29.8 versus 11.8, P < 0.05) and 5 mM (87.6 versus 34.0, P < 0.001) DEB, In accordance with the known strong clastogenicity of DE B, MN without centromeric FISH signals were particularly increased, th e difference between the two GSTT1 genotypes being statistically signi ficant at both concentrations of DEB (mean induced MN/1000 binucleate cells 23.1 versus 9.9, P < 0.05, at 2 mM; 69.7 versus 24.2, P < 0.001, at 5 mM), In addition, centromere-positive (C+) MN were induced, sugg esting that DEB also has some aneuploidogenic activity, The GSTT1-null genotype showed a significantly (P < 0.05) higher mean frequency of i nduced C+ MN than the GSTT1-positive genotype, at both 2 (6.7 versus 1 .9) and 5 mM (17.9 versus 9.8) DEB, At the higher dose mean nuclear di vision index was lower in the GSTT1-null group (1.80) than in the GSTT 1-positive group (2.05, P < 0.01), These findings support earlier resu lts from the analysis of sister chromatid exchange showing that indivi dual sensitivity to the genotoxic and cytotoxic effects of DEB is larg ely explained by the lack of the GSTT1 gene.