Balance-shift in the production of NO and superoxide via endothelial nitric oxide synthase in atherogenesis

来自内皮NO合成酶（eNOS）的NO被认为是抗动脉粥样硬化分子。事实上，最近对eNOS基因缺陷小鼠的研究表明，缺乏eNOS基因的小鼠动脉粥样硬化病变形成加速。另一方面，研究表明，在四氢生物蝶呤（BH4）缺乏或缺乏eNOS酶活性的情况下，eNOS功能失调，产生超氧化物而不是NO。BH4是合成NO的重要辅助因子。这种现象被称为eNOS解耦。在本研究中，我们研究了eNOS过表达如何影响高胆固醇血症下动脉粥样硬化的发生。我们将载脂蛋白e敲除小鼠（载脂蛋白E-KO）与内皮细胞中过表达eNOS的eNOS转基因小鼠（eNOS- tg）杂交。我们发现，在高胆固醇血症下，与载脂蛋白E-KO/eNOS-Tg相比，载脂蛋白E-KO/eNOS-Tg的主动脉超氧化物生成增加，一氧化氮生成相对较低。因此，在载脂蛋白E-KO/eNOS- tg中存在eNOS解耦。我们还发现，eNOS过表达加速了载脂蛋白E-KO中动脉粥样硬化病变的形成。与载脂蛋白E-KO相比，载脂蛋白E-KO/eNOS-Tg组血管BH4水平降低。外源性BH4慢性治疗载脂蛋白E-KO/eNOS-Tg可减少主动脉超氧化物的产生，增加NO的产生，减少动脉粥样硬化病变面积。通过与GTPCH 1转基因小鼠杂交进一步过表达BH4合成限速酶GTPCH 1，也减少了载脂蛋白E-KO/eNOS-Tg的动脉粥样硬化病变面积。因此，血管组织中BH4水平和eNOS蛋白水平之间似乎存在一种微小的平衡。当高胆固醇血症患者BH4水平降低时，eNOS解偶联并产生超氧化物而非一氧化氮，从而加速动脉粥样硬化的发生。

NO from the endothelial NO synthase (eNOS) is believed to act as anti-atherogenic molecule. Indeed recent studies in eNOS gene-deficient mice showed the accelerated atherosclerotic lesion formation in mice that lacks eNOS gene. On the other hand, it is revealed that eNOS becomes dysfunctional and produces superoxide rather than NO under conditions in which tetrahydrobiopterin (BH4), an essential cofactor for NO synthesis, is lacking or deficient for eNOS enzymatic activity. This phenomenon is called eNOS uncoupling. In the present study we examined how eNOS overexpression affects atherogenesis under hypercholesterolemia. We crossed apo E-knockout mice (apo E-KO) with eNOS transgenic mice that overexpress eNOS in the endothelium (eNOS-Tg). We found that under hypercholesterolemia superoxide production was increased and NO production was relatively low in the aortas from apo E-KO/eNOS-Tg compared with those from apo E-KO. Therefore the presence of eNOS uncoupling was suggested in apo E-KO/eNOS-Tg. We also revealed that eNOS overexpression accelerated atherosclerotic lesion formation in apo E-KO. Vascular levels of BH4 were decreased in apo E-KO/eNOS-Tg compared with apo E-KO. Chronic treatment of apo E-KO/eNOS-Tg with exogenous BH4 decreased superoxide production and increased NO production from aortas, and reduced the atherosclerotic lesion area. Further overexpression of GTPCH 1, the rate limiting enzyme of BH4 synthesis, by crossing with GTPCH 1 transgenic mice also decreased atherosclerotic lesion area in apo E-KO/eNOS-Tg. Therefore, it seems that there is a minute balance between BH4 levels and eNOS protein levels in vascular tissue. When BH4 levels decrease under hypercholesterlemia, eNOS becomes uncoupling and produces superoxide rather than NO, which serves to accelerate atherogenesis.