Radiation-induced double-strand breaks in mammalian DNA: influence of temperature and DMSO

Abstract

Purpose : To investigate the effects of subphysiological irradiation temperature (2-28°C) and the influence of the radical scavenger DMSO on the induction of double-strand breaks (DSB) in chromosomal DNA from a human breast cancer cell line (MCF7) as well as in intact cells. The rejoining of DSB in cells irradiated at 2°C or 37°C was also investigated. Materials and methods : Agarose plugs with [ 14 C]thymidine labelled MCF-7 cells were lysed in EDTA-NLS-proteinase-K buffer. The plugs containing chromosomal DNA were irradiated with Xrays under different temperatures and scavenging conditions. Intact MCF-7 cells were irradiated in Petri dishes and plugs were made. The cells were then lysed in EDTA-NLS-proteinaseK buffer. The induction of DSB was studied by constant field gel electrophoresis and expressed as DSB/100Mbp, calculated from the fraction of activity released into the gel. Results : The induction of DSB in chromosomal DNA was reduced by a decrease in temperature. This protective effect of low temperature was inhibited when the DNA was irradiated in the presence of DMSO. No difference was found when intact cells were irradiated at different temperatures. However, the rapid phase of rejoining was slower in cells irradiated at 37°C than at 2°C. Conclusions : The induction of DSB in naked DNA was reduced by hypothermic irradiation. The temperature had no influence on the induction of DSB in the presence of a high concentration of DMSO, indicating that the temperature effect is mediated via the indirect effects of ionizing radiation. Results are difficult to interpret in intact cells. Rejoining during irradiation at the higher temperature may counteract an increased induction. The difference in rejoining may be interpreted in terms of qualitative differences between breaks induced at the two temperatures.