SISTER CHROMATID EXCHANGE IN CHROMOSOMES OF UNTREATED AND DRUG TREATED HUMAN FIBROBLASTS

A number of variables contribute to the formation of sister chromatid exchange (SCE) in human fibroblasts in vitro. Cell culture conditions, drug effects and chronic drug exposures were investigated in the present study. In addition, the effects on SCE distribution in the genome of untreated and drug treated cells were studied.; SCE rates in untreated cells varied with serum concentration (5%-20%) and serum characteristics (whole, dialyzed, heat-inactivated or virus tested). The number of cells present at the time of analysis also contributed to variations in SCE rates. An inoculum of 1 x 10('5) fibroblasts in 5 ml of cell culture medium gave the highest mitotic indices and the least variability in SCE rates.; Four chemotherapeutic agents, each reported to cause chromosome aberrations and each with differing chemical reaction mechanisms and sites of involvement in cellular metabolism, were evaluated for their effects on SCE rates and mitotic indices. The two alkylating agents, Busulfan (BUS) and Mitomycin C (MMC), were found to increase SCE rates, while Methotrexate (MTX) and Vincristine (VNC) did not. Each exposure reduced the mitotic index. This suggests that those lesions leading to an SCE can accumulate in the cell or that the cells are more sensitive to SCE inducing events if they have been previously exposed to the agent.; Following acute exposure to MMC or BUS, fibroblasts rapidly recovered from the damage, as reflected in SCE rates and mitotic indices. As the number of exposures increased, recovery from the damage was slower. This may suggest a residual damage effect, because as the SCE rate returned to normal the mitotic index also recovered.; Lastly, sister chromatid exchanges were localized by simultaneous G-banding and differential sister chromatid staining of chromosomes obtained from untreated fibroblasts and fibroblasts treated with 10('-6)M BUS, 10('-6)M MTX, 10('-6)M VNC or 10('-8) M MMC. In untreated human fibroblasts, the SCE sites were found to be located in the euchromatic light G-bands and band-interband junctions more frequently than in the dark G-bands. There was no change in the number of location of SCEs in cells treated with Methotrexate or Vincristine. Mitomycin C treated fibroblasts had more SCEs in the dark G-bands than in the light G-bands. The Busulfan treated cells showed elevated SCE rates but no change in the distribution of SCE sites from that obtained in untreated cells.; The SCE sites were also evaluated for their distribution between the mid-arm regions of the chromosomes and the centromeric and telomeric regions. It was found that the telomeric and centromeric regions of the chromosomes of untreated cells were less frequently involved in SCE than were the midarm regions. Mitomycin C distorted this pattern with the distribution of SCE sites along the length of the metaphase chromosome becoming random.; These localization results may reflect the constraints placed on SCE formation by the nature of the euchromatin and heterochromatin. Euchromatin, being less condensed and more accessible than heterochromatin to its environment, is more frequently involved in SCE in untreated cells. Since this area contains the active unique sequence DNA, it cannot survive heavy damage. The heterochromatin, which contains the moderate and unique sequence DNA, can better tolerate residual damage.