DNA strand breakage, activation of poly(ADP-ribose) synthetase, and cellular energy depletion are involved in the cytotoxicity in macrophages and smooth muscle cells exposed to peroxynitrite
DNA strand breakage, activation of poly(ADP-ribose) synthetase, and cellular energy depletion are involved in the cytotoxicity in macrophages and smooth muscle cells exposed to peroxynitrite
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DOI：
10.1073/pnas.93.5.1753
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发表时间：
1996-03-05
影响因子：
11.1
通讯作者：
Salzman, AL
中科院分区：
文献类型：
--
作者：
Szabo, C;Zingarelli, B;Salzman, AL
自由基一氧化氮和超氧阴离子反应形成过氧亚硝酸盐（ONOO-），一种剧毒的氧化剂。体内ONOO-的形成已在休克和炎症中得到证实。在此，我们提供的证据表明，暴露于ONOO-的细胞毒性是由DNA链断裂和随后的DNA修复酶聚（ADP核糖）合成酶（PARS）的激活介导的。暴露于ONOO- (100 μ M ~ 1 mM)可抑制培养的J774巨噬细胞和大鼠主动脉平滑肌细胞的线粒体呼吸。细胞呼吸丧失迅速，在ONOO-暴露后1-3小时达到峰值，并且可逆，在6-24小时后恢复。线粒体呼吸的抑制与DNA链断裂的剂量依赖性增加平行，在暴露于ONOO-后20-30分钟达到最大。我们观察到暴露于ONOO-的细胞中PARS活性呈剂量依赖性增加。PARS抑制剂如3-氨基苯甲酰胺（1 mM）阻止了暴露于ONOO-的细胞呼吸的抑制。ONOO介导的DNA链断裂激活PARS导致细胞内能量储存显著减少，反映在细胞内NAD（+）和ATP含量下降。3-氨基苯甲酰胺可阻止ONOO-暴露细胞中NAD（+）和ATP的丢失。相反，一氧化氮供体s -亚硝基-n -乙酰基- dl -青霉胺或二乙基三胺一氧化氮复合物的加入对细胞呼吸的损害与DNA链断裂的发展无关，浓度高达1mm，并且在很大程度上对PARS抑制不起作用。我们的研究结果表明，DNA损伤和PARS（一种消耗能量的无效修复周期）的激活在ONOO介导的细胞损伤中起着核心作用。
The free radicals nitric oxide and superoxide anion react to form peroxynitrite (ONOO-), a highly toxic oxidant species. In vivo formation of ONOO- has been demonstrated in shock and inflammation. Herein we provide evidence that cytotoxicity in cells exposed to ONOO- is mediated by DNA strand breakage and the subsequent activation of the DNA repair enzyme poly(ADP ribose) synthetase (PARS). Exposure to ONOO- (100 mu M to 1 mM) inhibited mitochondrial respiration in cultured J774 macrophages and in rat aortic smooth muscle cells. The loss of cellular respiration was rapid, peaking 1-3 h after ONOO- exposure, and reversible, with recovery after a period of 6-24 h. The inhibition of mitochondrial respiration was paralleled by a dose-dependent increase in DNA strand breakage, reaching its maximum at 20-30 min after exposure to ONOO-. We observed a dose-dependent increase in the activity of PARS in cells exposed to ONOO-. Inhibitors of PARS such as 3-aminobenzamide (1 mM) prevented the inhibition of cellular respiration in cells exposed to ONOO-. Activation of PARS by ONOO--mediated DNA strand breakage resulted in a significant decrease in intracellular energy stores, as reflected by a decline of intracellular NAD(+) and ATP content. 3-Aminobenzamide prevented the loss of NAD(+) and ATP in cells exposed to ONOO-. In contrast, impairment of cellular respiration by the addition of the nitric oxide donors S-nitroso-N-acetyl-DL-penicillamine or diethyltriamine nitric oxide complex, was not associated with the development of DNA strand breaks, in concentrations up to 1 mM, and was largely refractory to PARS inhibition. Our results suggest that DNA damage and activation of PARS, an energy-consuming futile repair cycle, play a central role in ONOO--mediated cellular injury.