HPRT MUTANT FREQUENCY AND GSTM1 GENOTYPE IN NONSMOKING HEALTHY-INDIVIDUALS

The T-cell cloning assay which combines mitogen- and growth factor-dep endent expansion of lymphocyte clones with thioguanine selection of hy poxanthine-guanine phosphoribosyl transferase (hprt)-negative cells ha s been extensively used for studying human somatic gene mutation in vi vo. However, large interindividual variations in the hprt mutant frequ ency (MF), much of which is not explained by donor attributes such as age and smoking habit, and interlaboratory variations in the experimen tal methodology, including cloning efficiency (CE), call for further d evelopments of the cloning protocol and additional population studies. Using an improved T-cell cloning method, we have studied in vivo hprt MF of 76 non-smoking healthy males aged 23-77 years. The addition of 5% human serum to the growth medium was found to produce a consistentl y high CE of 61% in average. The MF, ranging from 1.4 to 22.6 x 10(-6) With a mean of 8.6 x 10(-6), increased significantly (P < 0.0001) wit h age, by 2% per year. A significant (P = 0.002) inverse relationship between MF and CE was observed. Using a PCR-based technique for GSTM1- genotyping, we also studied the relationship between MF and GSTM1 poly morphism. The 38 (50%) GSTM1-negative individuals showed a 20% higher mean MF than the 38 (50%) GSTM1-positive individuals. The difference w as however not significant, neither before (P = 0.1) nor after (P = 0. 5) correction for CE and the significantly (P = 0.04) higher mean age in the GSTM1-negative group. This study shows that age contributes mor e than GSTM1 polymorphism to the large interindividual variation in th e hprt MF of non-smokers. The relationship between GSTM1 polymorphism and hprt MF in smokers remains to be investigated. (C) 1995 Wiley-Liss , Inc.