Purines inhibit poly(ADP-ribose) polymerase activation and modulate oxidant-induced cell death

Purines such as adenosine, inosine, and hypoxanthine are known to have pote nt antiinflammatory effects. These effects generally are believed to be med iated by cell surface adenosine receptors. Here we provide evidence that pu rines protect against oxidant-induced cell injury by inhibiting the activat ion of the nuclear enzyme poly(ADP-ribose) polymerase (PARP). Upon binding to broken DNA, PARP cleaves NAD(+) into nicotinamide and ADP-ribose and pol ymerizes the latter on nuclear acceptor tor proteins such as histones and P ARP itself. Overactivation of PARP depletes cellular NAD(+) and ATP stores and causes necrotic cell death. We have identified some purines (hypoxanthi ne, inosine, and adenosine) as potential endogenous PARP inhibitors, We hav e found that purines (hypoxanthine > inosine > adenosine) dose-dependently inhibited PARP activation in peroxynitrite-treated macrophages and also inh ibited the activity of the purified PARP enzyme. Consistently with their PA RP inhibitory effects, the purines also protected interferon gamma + endoto xin (IFN/LPS) -stimulated RAW macrophages from the inhibition of mitochondr ial respiration and inhibited nitrite production from IFN/LPS-stimulated ma crophages. We have selected hypoxanthine as the most potent cytoprotective agent and PARP inhibitor among the three purine compounds, and investigated the mechanism of its cytoprotective effect. We have found that hypoxanthin e protects thymocytes from death induced by the cytotoxic oxidant peroxynit rite, In line with the PARP inhibitory effect of purines, hypoxanthine has prevented necrotic cell death while increasing caspase activity and DNA fra gmentation, As previously shown with other PARP inhibitors, hypoxanthine ac ted proximal to mitochondrial alterations as hypoxanthine inhibited the per oxynitrite-induced mitochondrial depolarization and secondary superoxide pr oduction. Our data imply that purines may serve as endogenous PARP inhibito rs. We propose that, by affecting PARP activation, purines may modulate the pattern of cell death during shock, inflammation, and reperfusion injury.