A COMPARATIVE-STUDY ON THE EFFECT OF MNU ON HUMAN LYMPHOCYTE-CULTURESIN-VITRO EVALUATED BY O-6-MDG FORMATION, MICRONUCLEI AND SISTER-CHROMATID EXCHANGES INDUCTION

N-Nitroso-compounds are a large group of chemicals present in a number of environmental sources and many of them are mutagens as well as car cinogens in experimental animals. Among the known N-nitroso-compounds, N-nitroso-N-methylurea (MNU) is a strong mutagen. In this study an ef fort has been made to compare the ability of MNU to methylate the O-6- guanosine site in DNA and to induce micronuclei and sister chromatid e xchanges in human lymphocyte cultures in vitro. To quantitate O-6-meth yldeoxyguanosine (O-6-mdG) a highly sensitive immunoassay, immune-slot -blot (ISB), has been used. For the evaluation of micro nuclei (MN) th e cytokinesis block micronucleus method has been used. Different conce ntrations (75, 100, 125 mu g/ml) were tested. At the highest concentra tion tested for the MN induction, 125 mu g/ml, the occurrence of binuc leates and micro nuclei is higher than twice in relation to control an d a reduction in NDI is also observed. The same concentrations were us ed for the estimation of sister chromatid exchanges (SCEs) induction. The mean number of SCEs at 125 mu g/ml is almost three times that of t he control level. The concentrations tested for the quantitation of O- 6-mdG were 200, 300 and 400 mu g/ml and this was done because for the test system we used and for the given experimental conditions the firs t indication of O-6-mdG formation was at 200 mu g/ml. Our results show that methylation of O-6-guanosine increases with concentration and at 400 mu g/ml the concentration of O-6-mdG is 5.83 fmol/mu g DNA, while at the control level it is 2.40 fmol/mu g DNA. Since O-6-mdG formatio n is observed in higher concentrations than those of MN and SCE induct ion it would be interpreted that O-6-mdG levels are not correlated wit h the studied cytogenetic effects although one has to take into consid eration the total promutagenic lesions in DNA, induced by MNU, as well as AGT repair activity.