THE ROLE OF NITRIC-OXIDE IN LIPOXIN A(4)-INDUCED POLYMORPHONUCLEAR NEUTROPHIL-DEPENDENT CYTOTOXICITY TO HUMAN VASCULAR ENDOTHELIUM IN-VITRO
THE ROLE OF NITRIC-OXIDE IN LIPOXIN A(4)-INDUCED POLYMORPHONUCLEAR NEUTROPHIL-DEPENDENT CYTOTOXICITY TO HUMAN VASCULAR ENDOTHELIUM IN-VITRO
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DOI：
10.1002/art.1780380609
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发表时间：
1995-06-01
影响因子：
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通讯作者：
GYLLENHAMMAR, H
中科院分区：
文献类型：
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作者：
BRATT, J;GYLLENHAMMAR, H
Objective.探讨花生四烯酸脂氧合酶5-和15-脂氧合酶产物脂氧素A（4）（LXA（4））和佛波醇酯佛波醇肉豆蔻酸酯（PMA）诱导人中性粒细胞（PMN）对人脐静脉内皮细胞（HUVEC）的细胞毒作用机制。HUVEC生长至汇合并用Cr-51标记。加入中性粒细胞和刺激物，4 h后测定上清液中Cr-51的释放。LXA（4）和PMA均能产生高度显著的PMN依赖性细胞溶解作用。LXA（4）激活的细胞毒性可被N-G-单甲基-L-精氨酸（L-NMA）和硝基-L-精氨酸甲酯（NO合成酶的特异性抑制剂）抑制。此外，细胞外NO的清除剂氧合血红蛋白（HbO（2））以剂量依赖性方式阻止LXA（4）诱导的细胞溶解。与此形成鲜明对比的是，L-NMA对PMA诱导的细胞溶解作用无明显影响，而HbO（2）则表现出一定的抑制作用。在无PMN的实验中，加入NO供体S-亚硝基-N-乙酰青霉胺诱导HUVEC明显的细胞溶解，HbO可抑制此作用，但L-NMA不抑制此作用。此外，L-精氨酸或精氨酸类似物不影响超氧阴离子的生产在无细胞次黄嘌呤/黄嘌呤氧化酶系统。LXA（4）和PMA均能诱导PMN产生超氧阴离子和HUVEC产生NO。在血管炎的体外模型中，HUVEC产生的NO与产生超氧阴离子的PMN相互作用，对内皮细胞损伤具有显著意义。这可能是由于随后通过NO与O-2（-）之间的自由基-自由基相互作用形成细胞毒性产物，如过氧亚硝酸盐及其代谢产物。尽管LXA（4）和PMA在最佳浓度下诱导的细胞溶解相当，但与介导细胞毒性的其他机制相比，NO的相对重要性是刺激依赖性的，并且NO在LXA（4）诱导的PMN依赖性内皮损伤中相对更重要。
Objective. To assess the mechanism for the cytotoxicity of human polymorphonuclear neutrophils (PMN) to human umbilical vein endothelial cells (HUVEC) induced by the 5- and 15-lipoxygenase product of arachidonate, lipoxin A(4) (LXA(4)), and the phorbol ester, phorbol myristate acetate (PMA).Methods. HUVEC were grown to confluence and labeled with Cr-51. PMN and stimuli were added, and the release of Cr-51 info supernatants was assessed after 4 hours.Results. Both LXA(4) and PMA conferred highly significant PMN-dependent cytolysis. The cytotoxicity activated by LXA(4) was inhibited by N-G-monomethyl-L-arginine (L-NMA) and by nitro-L-arginine methyl ester, specific inhibitors of the nitric oxide (NO)-producing enzyme NO synthase. Also, the scavenger of extracellular NO, oxyhemoglobin (HbO(2)), prevented LXA(4)-induced cytolysis in a dose-dependent manner. In sharp contrast, L-NMA did not significantly affect the cytolysis induced by PMA, whereas HbO(2) showed a modest inhibitory action. In experiments without PMN, addition of the NO donor S-nitroso-N-acetyl-penicillamine to HUVEC induced marked cytolysis, which was inhibited by HbO,, but not by L-NMA. Addition of L-arginine or arginine analogs did not affect superoxide anion production in a cell-free hypoxanthine/xanthine oxidase system. Both LXA(4) and PMA induced the production of superoxide anion from PMN and of NO from HUVEC.Conclusion. NO produced by HUVEC, interacting with PMN which produce superoxide anions, is of marked significance for the endothelial cell damage in this in vitro model of vasculitis, This is probably due to the subsequent formation, via a radical-radical interaction between NO and O-2(-), of cytotoxic products, such as peroxynitrite and its metabolites, Furthermore, although LXA(4) and PMA induced comparable cytolysis at optimal concentrations, the relative importance of NO compared with other mechanisms mediating cytotoxicity was stimulus dependent, and NO was relatively more important for LXA(4)-induced PMN-dependent endothelial injury.