The use of mixed lymphocytes in DNA repair assays

253 Capelli, E., M. Stefanini, E. Giulotto and F . Nuzzo, Istituto di Genetica, Universit~ di Pavia, Laboratorio di Genetica Biochimica ed Evoluzionistica del C.N.R., Pavia (Italy) The use of mixed lymphocytes in DNA repair assays Human lymphocytes in Go phase are a suitable material for the determination of DNA damage since they are able to perform unscheduled DNA synthesis. The requirement for relatively large amounts of cells and the individual variability in the capacity to repair DNA damage can be serious limitations to the development of test systems based on the use of human blood. Therefore we checked the possibility of using pooled lymphocytes separated from the blood of different donors in the DNA repair assays on known mutagenic agents such as UV light, MMS, EMS, epichlorohydrin and hycanthone. The cells were mixed immediately before testing and the amount of [3H]thymidine incorporation in the presence of hydroxyurea was measured after treatment with the mutagenic agent. We also found that samples of lymphocytes stored in liquid nitrogen and pooled after thawing retained their capacity to perform repair synthesis after UV irradiation. The overall results indicate that mixed lymphocytes can be conveniently used in DNA repair synthesis assays. DiLemia, R., G. Della Valle, M.L. Tenchini and E. Tibaldi a, Istituto di Biologia Generale, Facolt~ di Medicina, UniversitY, Milan, and a Istituto di Zoologia, Faeolt~ di Scienze, UniversitY, Milan (Italy) Comparisvn between a specific rRNA inhibitor (MPB) and X-ray effects on satellite associations in human chromosomes Satellite associations of human acrocentric chromosomes depend on the presence of functional ribosomal genes localized on secondary constrictions. Therefore, satellite association frequency of these chromosomes and the average number of associated chromosomes per cell may be considered as characteristic parameters of the different individuals. We have used these parameters to test in human peripheral blood cultures both effects of rRNA transcription specific inhibitor and ionizing radiations. In particular, we have analyzed the satellite associations of lymphocyte metaphase plates to compare the effects of MPB (at d o s e s o f 5, 10, 20, 30, 50, 100 and 200 #g/ml), reported to be a reversible inhibitor of rRNA synthesis, and X-rays (at doses of 50, 100, 150 and 200 Rad). Both chemical and physical agents induce in the same way significant fluctuations in the values of the measured parameters. A decrease of relative satellite association frequency of "D" group chromosomes and the average number of associated chromosomes per cell were found. These data may be explained by the possibility that the damage mechanism affecting the nucleolar organizers is at random. In fact if theoretically all functional NORs carried on acrocentric chromosomes are involved at random in satellite associations, " D " group chromosomes are obviously more frequent (50%) and they are consequently proportionally more damaged. Our results may be in support of this assumption.