Immunolocalization of 8-oxoguanine in nutrient-deprived mammalian tissue culture cells

Nutrient deprivation, as experienced by tissues during ischemia and other f orms of injury may induce oxidative DNA damage. To explore this possibility , nutrient-deprived human HeLa and simian Cos7 cells were evaluated for 8-o xoguanine, a well established marker for oxidative DNA damage. Monoclonal a ntibodies directed against 8-oxoguanine were used to detect and localize th is lesion in vitro and in situ. Under controlled culture conditions, short term nutrient deprivation was sufficient to induce the accumulation of 8-ox oguanine in actively growing HeLa and Cos7 cells. Immunofluorescence analyses revealed that 8-oxoguanine was primarily locali zed to the nucleus in nutrient-deprived cells, and this nuclear accumulatio n could be reversed to base levels once cells were returned to normal growi ng conditions. Furthermore, confocal immunofluorescence with anti-8-oxoguan ine antibodies revealed several distinct areas of intense staining at the p eriphery and interior of interphase nuclei in nutrient deprived cells. Usin g this novel approach, we have found that relatively mild endogenous and ex ogenous factors contribute to oxidative DNA damage. Because 8-oxoguanine is mutagenic, we believe further studies may provide evidence to support the hypothesis that endogenous DNA damage is involved in the etiology of both d egenerative diseases and cancer.