Purines inhibit poly(ADP-ribose) polymerase activation and modulate oxidant-induced cell death.
Purines such as adenosine, inosine, and hypoxanthine are known to have potent antiinflammatory effects. These effects generally are believed to be mediated by cell surface adenosine receptors. Here we provide evidence that purines protect against oxidant-induced cell injury by inhibiting the activation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP). Upon binding to broken DNA, PARP cleaves NAD+ into nicotinamide and ADP-ribose and polymerizes the latter on nuclear acceptor proteins such as histones and PARP itself. Overactivation of PARP depletes cellular NAD+ and ATP stores and causes necrotic cell death. We have identified some purines (hypoxanthine, inosine, and adenosine) as potential endogenous PARP inhibitors. We have found that purines (hypoxanthine > inosine > adenosine) dose-dependently inhibited PARP activation in peroxynitrite-treated macrophages and also inhibited the activity of the purified PARP enzyme. Consistently with their PARP inhibitory effects, the purines also protected interferon gamma + endotoxin (IFN/LPS) -stimulated RAW macrophages from the inhibition of mitochondrial respiration and inhibited nitrite production from IFN/LPS-stimulated macrophages. 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 hypoxanthine protects thymocytes from death induced by the cytotoxic oxidant peroxynitrite. In line with the PARP inhibitory effect of purines, hypoxanthine has prevented necrotic cell death while increasing caspase activity and DNA fragmentation. As previously shown with other PARP inhibitors, hypoxanthine acted proximal to mitochondrial alterations as hypoxanthine inhibited the peroxynitrite-induced mitochondrial depolarization and secondary superoxide production. Our data imply that purines may serve as endogenous PARP inhibitors. We propose that, by affecting PARP activation, purines may modulate the pattern of cell death during shock, inflammation, and reperfusion injury.
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Inotek Corporation, Beverly, Massachusetts 01915, USA.
Szabó
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D000818:Animals; D002308:Cardiolipins / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects*; D002460:Cell Line; D019069:Cell Respiration / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003574:Cytochrome c Group / Q000378:metabolism; D019610:Cytoprotection / Q000187:drug effects*; D053938:DNA Fragmentation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry; D019271:Hypoxanthine / Q000494:pharmacology; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D009336:Necrosis; D009566:Nitrates / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D016877:Oxidants / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D011687:Purines / Q000378:metabolism / Q000494:pharmacology*; D013481:Superoxides / Q000378:metabolism; D013950:Thymus Gland / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism
D002308:Cardiolipins; D003574:Cytochrome c Group; D009566:Nitrates; D016877:Oxidants; D011687:Purines; D013481:Superoxides; C039642:peroxynitric acid; D019271:Hypoxanthine; D009569:Nitric Oxide; D011065:Poly(ADP-ribose) Polymerases; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases