Nitroxides Block DNA Scission and Protect Cells from Oxidative Damage

The protective effect of cyclic stable nitroxide free radicals, having SOD-like activity, against oxidative damage was studied by using Escherichia coli xthA DNA repair-deficient mutant hypersensitive to H₂O₂. Oxidative damage induced by H₂O₂ was assayed by monitoring cell survival. The metal chelator 1,10-phenanthroline (OP), which readily intercalates into DNA, potentiated the H₂O₂-induced damage. The extent of in vivo DNA scission and degradation was studied and compared with the loss of cell viability. The extent of DNA breakage correlated with cell killing, supporting previous suggestions that DNA is the crucial cellular target of H₂O₂ cytotoxicity. The xthA cells were protected by catalase but not by superoxide dismutase (SOD). Both five- and six-membered ring nitroxides, having SOD-like activity, protected growing and resting cells from H₂O₂ toxicity, without lowering H₂O₂ concentration. To check whether nitroxides protect against O₂·⁻-independent injury also, experiments were repeated under hypoxia. These nitroxides also protected hypoxic cells against H₂O₂, suggesting alternative modes of protection. Since nitroxides were found to reoxidize DNA-bound iron(II), the present results suggest that nitroxides protect by oxidizing reduced transition metals, thus interfering with the Fenton reaction.