Effect of antioxidant vitamin supplementation on DNA damage and repair in human lymphoblastoid cells

In this study the possible protective effects of ascorbic acid and alpha-tocopherol (singly and in combination) on Raji lymphoblastoid cells exposed to various doses of X-rays or hydrogen peroxide (H2O2) are investigated. DNA strand breaks and alkali-labile sites were measured using the alkaline comet assay. Survival and hypoxanthine guanine phosphoribosyl transferase mutant frequency were measured using the colony-forming assay. Ascorbic acid (60 mu M) and alpha-tocapherol (30 mu M) were added singly or together to cell culture medium 24 hours before treatment and were present during treatment. After the 24-hour supplementation period with ascorbic acid atone, alpha-tocopherol alone, and ascorbic acid + alpha-tocopherol, the level of endogenous DNA damage was significantly lower (p < 0.05) than in the nonsupplemented culture, as assessed by the comet assay. By use of the comet assay, it was observed that ascorbic acid exhibited an overall protective effect against DNA damage induced after X-ray treatment, whereas alpha-tocopherol exhibited an overall protective effect against DNA damage induced after H2O2 treatment. Significant increases were observed in the percent survival after 1-Gy X-rays and 5 and 20 mu M H2O2 in those cultures supplemented with ascorbic acid alone and alpha-tocopherol alone relative to the nonsupplemented cultures. The endogenous level of mutant frequency was also significantly decreased in the presence of ascorbic acid relative to the nonsupplemented culture. These findings are consistent with the concept that ascorbic acid and alpha-tocopherol can, under certain conditions, protect against oxidatively induced DNA damage.