Influence of DMSO on Carbon K ultrasoft X-rays induced chromosome aberrations in V79 Chinese hamster cells.

Ultrasoft X-rays have been shown to be very efficient in inducing chromosomal aberrations in mammalian cells. The present study was aimed to evaluate the modifying effects of DMSO (a potent scavenger of free radicals) on the frequencies of chromosome aberrations induced by soft X-rays. Confluent held G1 Chinese hamster cells (V79) were irradiated with Carbon K ultrasoft X-rays in the presence and absence of 1M DMSO and frequencies of chromosome aberrations in the first division cells were determined. DMSO reduced the frequencies of exchange types of aberrations (dicentrics and centric rings) by a factor of 2.1-3.5. The results indicate that free radicals induced by ultrasoft X-rays contribute to a great extent to the induction of chromosome aberrations. The possible implications of these results in interpreting the mechanisms involved in the high efficiency of ultrasoft X-rays in the induction of chromosome aberrations are discussed.

Relationship between chromatin structure, DNA damage and repair following X-irradiation of human lymphocytes.

Earlier studies using the technique of premature chromosome condensation (PCC) have shown that in human lymphocytes, exchange type of aberrations are formed immediately following low doses (<2 Gy) of X-rays, whereas at higher doses these aberrations increase with the duration of recovery. This reflects the relative roles of slow and fast repair in the formation of exchange aberrations. The underlying basis for slow and fast repairing components of the DNA repair may be related to differential localization of the initial damage in the genome, i.e., between relaxed and condensed chromatin. We have tried to gain some insight into this problem by (a) X-irradiating lymphocytes in the presence of dimethyl sulfoxide (DMSO) a potent scavenger of radiation-induced .OH radicals followed by PCC and (b) probing the damage and repair in two specific chromosomes, 18 and 19, which are relatively poor and rich in transcribing genes by COMET-FISH, a combination of Comet assay and fluorescence in situ hybridization (FISH) techniques. Results obtained show (a) that both fast appearing and slowly formed exchange aberrations seem to take place in relaxed chromatin, since they are affected to a similar extent by DMSO, (b) significant differential DNA breakage of chromosome 18 compared to chromosome 19 in both G0 and G1 phases of the cell cycle as detected by Comet assay, indicating that relaxed chromatin containing high densities of transcriptionally active genes shows less fragmentation due to fast repair (chromosome 19) compared to chromosome 18, and (c) that relaxed chromatin is repaired or mis-repaired faster than more compact chromatin.

Matrix association of early- and late-replicating chromatin studied by single-cell electrophoresis.

CHO-K1 cells were synchronized at the G(1)/S border by mitotic shake-off and aphidicolin incubation. Pulse-labeling with tritium was done at 30 min, 2 or 5 h into the S-phase, with chase incubations for different times in non-radioactive medium. The cells were subjected to neutral microelectrophoresis to extend the DNA into "comets," after which the label was visualized through autoradiography. At zero chase time, all label was positioned in the head. The displacement of label into the tails increased with time, reaching a maximum at about 5 h after the pulse. A lag phase of 2-3 h was observed for the early-labeled cells before the displacement started. Also, more label was released after overnight serum starvation, but this was reversed through a 3-h incubation at normal growth conditions. It was found that late-replicating chromatin is organized in larger domains than early-replicating chromatin, and DNA polymerase seems to be an important organizer. Early-replicating chromatin has other important attachments to the nuclear matrix, dependent on metabolic activity.