Radiation-induced DNA single-strand break induction and repair in murine tissues measured by the comet assay

• Main Author: Zheng, Haihua
• Language: English
• Published: 2009
• Online Access: http://hdl.handle.net/2429/4534

DNA damage produced in C3H/HeN murine tissues by ionizing radiation was characterized at the level of the individual cell with the goal of defining tissue dependent differences in DNA single-strand break (SSB) induction and repair. Subsequently, nicotinamide, was examined as a modifier of initial DNA damage and SSB rejoining following irradiation. The alkaline comet assay, a single-cell gel electrophoresis method, was used to examine cells from SCCVM tumours, spleen, bone marrow, liver, jejunum, testis, thymus and cerebellum. Cells from all tissues irradiated in vitro showed similar radiosensitivity. However, for in vivo irradiation, rapid SSB rejoining which occurs in cells during irradiation led to differences between tissues. Also, tumour and testis showed less damage in vivo than other normal tissues. Consistent with previous studies, these two tissues were found to contain radiobiologically hypoxic cells. Efficiency of SSB rejoining was cell type-dependent; cells from SCCVII tumors rejoined breaks about 5 times more rapidly than cells from cerebellum. Heterogeneity in speed of rejoining was minimal among cells of a tissue, and no significant damage remained 4 hours following 15 Gy. However, extensive DNA degradation was observed in all tissues except brain 48 hours after 15 Gy. DNA ladder patterns in agarose gels, typical of apoptosis, were observed 4 hours after 2-10 Gy in spleen and thymus. The vitamin B analogue, nicotinamide, was shown to improve testis and tumour oxygenation, in agreement with other studies. A new observation was that nicotinamide (500 mg/kg or more) given before irradiation inhibited SSB rejoining in cells of all tissues except brain. Furthermore, radiation-induced DNA degradation was found to be greatly accelerated by nicotinamide. Both effects are likely to involve poly(ADP-ribose) polymerase inhibition. However, while nicotinamide significantly retarded radiation-induced SSB rejoining in tumors, the biological significance of this effect is questionable since nicotinamide did not enhance oxygen-independent killing of irradiated tumor cells.