THE REGULATION OF ENDOTHELIAL CONSTITUTIVE NITRIC OXIDE SYNTHASE IN REGENERATED ENDOTHLIAL CELLS

我们检验了球囊剥脱后再生内皮中超氧阴离子加速内皮源性一氧化氮降解的假设。为了测量在正常猪冠状动脉（对照血管）和球囊暴露后两周的冠状动脉（损伤血管）内超氧阴离子的产生，我们使用基于光泽精的化学发光的超氧阴离子测定，使用光度计（Shimazu）。在病理学研究中，在受损血管中观察到中度内膜增厚。在单独的等长张力研究中，5-羟色胺诱导的血管收缩在损伤的血管段中显著增强。损伤血管产生的超氧阴离子比对照血管多2 - 3倍（n = 8）。有趣的是，裸露的对照血管和裸露的损伤血管之间没有差异，表明再生的内皮细胞是超氧阴离子产生的来源。为了进一步研究再生内皮细胞超氧阴离子过量产生的分子机制，我们切除了受损的冠状动脉，用免疫组织化学方法定量酶的表达，用抗小鼠内皮细胞组成型一氧化氮合酶（ecNOS），小鼠铜锌超氧化物歧化酶（Cu-Zn SOD），小鼠黄嘌呤脱氢酶（XDH）的单克隆抗体。我们观察到再生内皮细胞内ecNOS表达显著减弱，而Cu-Zn SOD或XDH表达不变。球囊剥脱后再生内皮细胞内超氧阴离子产生增加和ecNOS表达减弱可能在早期动脉粥样硬化过程中起重要作用。

We tested the hypothesis that accelerated degradation of endothelium-derived nitric oxide by superoxide anion in regenerated endothelium after baloon denudation. To measure superoxide anion production within normal pig coronary arteries (control vessels) and coronary arteries two weeks after baloon dunudation (injured vessels), we used an assay for superoxide anion based on the chemiluminescence of lucigenin using luminometer (Shimazu). In pathological studies, moderate intimal thickening was observed in injured vessels. In separate isometric tension studies, serotonin- induced vasoconstriction was markedly augmented in injured vessel segments. Injured vessels produced two to threefold more superoxide anion than control vessels (n = 8). Interestingly, there was no difference between denuded control vessels and denuded injured vessels, suggesting that the regenerated endothlium is a source of superoxide anion production. To further investigate the molecular mechanisms of excess production of superoxide anion within regenerated endothlium, we excised injured coronary vessels and quantitated the expression of enzyme by immunohistochemical method using monoclonal antibody against mouse endothelial constitutive nitric oxide synthase (ecNOS), mouse Cu-Zn containing superoxide dismutase (Cu-Zn SOD), and mouse xanthine dehydrogenase (XDH). We observed ecNOS expression was significantly attenuated within regenerated endothlium, whereas either Cu-Zn SOD or XDH expression unchanged. Increased endothelial superoxide anion production and attenuated expression of ecNOS within regenerated endothlium after baloon denudation may play an important role in the early atherosclerotic process.