Regulation of gene expression of inducible nitric oxide synthase (iNOS) in cardiovascular system and its molecular mechanism of action

诱导型一氧化氮合酶过度产生一氧化氮参与内毒素休克和动脉粥样硬化病变的发生。为了阐明iNOS源性NO的作用，我们研究了iNOS基因在培养的大鼠血管平滑肌细胞（VSMC）中的表达及其作用的分子机制。脂质体转染诱导型一氧化氮合酶cDNA构建体进入VSMC引起大量的NO产生伴随着细胞凋亡和DNA合成的抑制，提示NO作为促凋亡因子的作用，以防止动脉粥样硬化内膜增厚。加入NO供体可引起培养的大鼠内皮细胞（EC）大量凋亡，其作用可被内皮素-1（ET-1）阻断，提示NO是促凋亡因子，而ET-1是抗凋亡因子，二者相互作用维持内皮细胞的完整性。通过IkBα的磷酸化和降解激活核因子（NF）-kB，从而导致iNOS基因启动子的反式激活。然而，NO供体通过阻断IkBα的磷酸化和随后的降解来抑制细胞因子诱导的iNOS基因表达，表明由iNOS产生的内源性NO以自反馈方式抑制iNOS的过表达。糖皮质激素还通过阻断IkBα的磷酸化和降解来抑制细胞因子诱导的iNOS基因表达。与此相反，非甾体抗炎药（NSAID），如阿司匹林和水杨酸钠，通过阻断（后）翻译水平抑制细胞因子刺激的NO产生，而不影响NF-κ B激活或iNOS基因转录。这些数据表明，糖皮质激素和NSAID发挥其抗炎作用，通过阻断NO的生产，通过不同的网站的行动。

Excessive production of NO by iNOS is implicated in the development of endotoxic shock and atherosclerotic lesion. To elucidate the role of iNOS-derived NO, we have studied the molecular mechanisms of iNOS gene expression and its actions in cultured rat vascular smooth muscle cells (VSMC). Transfection of iNOS cDNA construct into VSMC by lipofection caused a massive generation of NO accompanied by apoptosis and inhibition of DNA synthesis, suggesting the role of NO as proapoptotic factor to prevent intimal thickening in atherosclerosis. Addition of NO donors caused massive apoptosis of cultured rat endothelial cells (EC), whose effect was blocked by endothelin-1 (ET-1), suggesting that NO functions as a proapoptotic factor, whereas ET-1 function as an anti-apoptotic factor, thereby conteracting with each other to maintain endothelial integrity.Inflammatory cytokines, such as interleukin (IL)-1 and tumor necrosis factor (TNF)-α, activate nuclear factor (NF)-kB by phosphorylation and degradation of IkBα, thereby leading to transactivation of iNOS gene promoter. However, NO donors inhibited cytokines-induced iNOS gene expression by blocking phosphorylation and subsequent degradation of IkBα, suggesting that endogenous NO generated from iNOS inhibits overexpression of iNOS in an autofeedback fashion. Glucocorticoid also inhibited cytokines-induced iNOS gene expression by blockade of IkBα phosphorylation and degradation. In contrast, nonsteroidal anti-inlflammatory drugs (NSAID), such as aspirin and sodium salicylate, inhibited cytokines-stimulated NO production by blocking (post) translational level without affecting NF-kB activation or iNOS gene transcription. These data suggest that glucocorticoid and NSAID exert their anti-inflammatory effects by blocking NO production via different sites of action.