Protective Effect And Mechanism Of Nitric Oxide Against Hydroxyl Radical-induced DNA Damage

ObjectiveNitric oxide (NO) is one of the most important biological molecules in human body, at the same time is also a kind of highly unstable biological free radicals. NO is used as the research object of this study. It’s the first time to achieve NO saturated aqueous solution by simple method which is used to do the DNA experiment in vitro, then explore the activity of NO protecting hydroxyl-induced DNA damage and analyze its possible mechanism of protecting DNA oxidative damage.MethodsPrepration of NO saturated water solution:NO(g) was collected by drainage method, then joined the degassing of distilled water to the bottle filled with NO(g).Protective effect against hydroxyl radical-induced DNA damage detection: the protective effect in vivo against hydroxyl radical-induced DNA damage was measured by ultraviolet spectrophotometry and HPLC. Then, we analyzed the protective effect of NO against mouse hippocampal HT-22cells oxidative damage induced by hydroxyl radical.The mechanism of NO against hydroxyl radical-induced DNA damage:We used diazotization coupling spectrometry to demonstrate the existence of nitrous acid. Investigate the radical-scavenging abilities of DPPH· and Fe3+reducing power ability to detect the antioxidant ability of NO to discuss possible mechanisms.ResultsNO could effectively protect hydroxyl-induced DNA damage in a dose dependent manner, The IC50value was calculated as0.19±0.01mmol/L. We found NO can protect hydroxyl-induced bases damage by comparing the spectra. The IC50values were calculated as0.89±0.013,1.12±0.043,1.21±0.32,1.22±0.036,1.13±0.021mmol/L, respectively, for Adenine, Guanine, Cytosine, Thymine, Uracil.5μg/mL NO could reduce the oxidative damage of cell induced by hydroxyl radical and improve cell vitality.We detected the existence of HNO2in the production mixed NO and·OH by the methods of ultraviolet spectrometry. The regression equation of standard curve was Y=0.4843X+0.0246(R=0.99914) measured by ultraviolet spectrometry and the concentration was0.19mmol/L. In this study, the antioxidant was investigated by various assays in vitro. The IC50values were calculated as27.74±0.50and313.5±0.05μmol/mL, respectively for DPPH· and Fe3+reducing power.ConclusionNO can acavenge free radicals and also has reduction ability. It can be used as an antioxidant protection to reduce Hydroxyl-induced DNA damage.Nitric oxide exerts its protective role via the reaction with·OH radical to produce nitrous acid (HNO2), in which NO partly donates electron (e) to·OH radical.