Comparative effect of the thiols dithiothreitol, cysteamine and WR-151326 on survival and on the induction of DNA damage in cultured Chinese hamster ovary cells exposed to gamma-radiation

Abstract

We compared the ability of three thiols--dithiothreitol (DTT), cysteamine and WR-151326--to protect aerated Chinese hamster ovary cells from the lethal and DNA-damaging effects of gamma-radiation. These results were compared with earlier measurements for WR-1065 and WR-255591. The time-course and the concentration dependence of protection against cell killing was determined after 10 Gy of gamma-rays. The aminothiols cysteamine and WR-151326 protected at much lower extracellular concentrations than the simple thiol DTT; however, there was no clear difference between the behaviour of cysteamine, WR-151326, WR-1065 and WR-255591 in this respect. Protection by DTT and cysteamine was complete within 1 min, whereas for WR-151326, WR-1065 and WR-255591 about 30 min was required before protection began to reach a plateau. Based on these data, complete radiation survival curves were generated for each thiol and protection factors calculated. Effects on the induction of DNA single-strand breaks (ssb) and double-strand breaks (dsb) by gamma-rays were measured using alkaline (pH 12.1) and neutral (pH 7.0 and 9.6) elution, respectively. All three thiols protected against ssb induction, although to a significantly lower extent than against cell killing measured under identical conditions. Each thiol also protected against dsb induction. After high radiation doses the protection factors for dsb induction were also less than the protection factors for cell survival; however, when dsb were assayed using the low-dose replicate plating neutral elution method, the relative effect of each thiol on cell survival and on dsb induction appeared to be equivalent. The hierarchy of protection against both ssb and dsb induction (based on the extracellular thiol concentration required to produce a given degree of protection) was similar to that for cell survival, i.e., WR-151326 congruent to cysteamine less than DTT.