Virág L, Szabó C
Inotek Corporation, Beverly, Massachusetts 01915, USA.
FASEB J. 2001 Jan;15(1):99-107. doi: 10.1096/fj.00-0299com.
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.
已知嘌呤如腺苷、肌苷和次黄嘌呤具有强大的抗炎作用。一般认为这些作用是由细胞表面腺苷受体介导的。在此我们提供证据表明，嘌呤通过抑制核酶聚（ADP - 核糖）聚合酶（PARP）的激活来保护细胞免受氧化剂诱导的损伤。PARP与断裂的DNA结合后，将NAD + 裂解为烟酰胺和ADP - 核糖，并将后者聚合到核受体蛋白如组蛋白和PARP自身上。PARP的过度激活会耗尽细胞内的NAD + 和ATP储备，并导致坏死性细胞死亡。我们已确定一些嘌呤（次黄嘌呤、肌苷和腺苷）为潜在的内源性PARP抑制剂。我们发现嘌呤（次黄嘌呤＞肌苷＞腺苷）在剂量依赖性地抑制过氧亚硝酸盐处理的巨噬细胞中PARP的激活，并且还抑制纯化的PARP酶的活性。与它们的PARP抑制作用一致，嘌呤还保护干扰素γ + 内毒素（IFN/LPS）刺激的RAW巨噬细胞免受线粒体呼吸抑制，并抑制IFN/LPS刺激的巨噬细胞产生亚硝酸盐。我们选择次黄嘌呤作为三种嘌呤化合物中最有效的细胞保护剂和PARP抑制剂，并研究了其细胞保护作用的机制。我们发现次黄嘌呤可保护胸腺细胞免受细胞毒性氧化剂过氧亚硝酸盐诱导的死亡。与嘌呤的PARP抑制作用一致，次黄嘌呤可防止坏死性细胞死亡，同时增加半胱天冬酶活性和DNA片段化。如先前用其他PARP抑制剂所显示的那样，次黄嘌呤在线粒体改变的近端起作用，因为次黄嘌呤抑制过氧亚硝酸盐诱导的线粒体去极化和继发性超氧化物产生。我们的数据表明嘌呤可能作为内源性PARP抑制剂。我们提出，通过影响PARP激活，嘌呤可能在休克、炎症和再灌注损伤期间调节细胞死亡模式。
