Induction of DNA strand breakage and base oxidation by nitroxyl anion through hydroxyl radical production

Nitroxyl anion (NO−), the one-electron reduction product of nitric oxide (NO√), has been reported to be formed under various physiological conditions and to be cytotoxic, although the mechanism responsible for the toxic effects has not been identified. We have studied the effects of NO− generated from Angeli’s salt (sodium trioxodinitrate) or Piloty’s acid (N-hydoxybenzenesulfonamide) on DNA strand breakage and DNA base oxidation in vitro. Induction of strand breakage was dose- and time-dependent upon incubation of plasmid pBR322 with Angeli’s salt or Piloty’s acid. Similarly, 8-oxo-2′-deoxyguanosine and malondialdehyde were formed when calf-thymus DNA or 2′-deoxyribose, respectively, were incubated with Angeli’s salt. Electron acceptors (ferricyanide, 4-hydroxy-TEMPO), that convert NO− to NO√, inhibited the reactions, indicating that NO−, but not NO√, is responsible for the reactions. Furthermore, the reactions were also inhibited by the presence of hydroxyl radical (HO√) scavengers, antioxidants, metal chelators and superoxide dismutase and catalase, implying involvement of free HO√. These results suggest that NO− is a possible endogenous source of HO√, that may be formed either directly from the reaction product of NO− with NO√ (N2O2•−) or indirectly through H2O2 formation. Thus NO− may play an important role as a cause of diverse pathophysiological conditions such as inflammation and neurodegenerative diseases.