Johnson David K, Schillinger Kurt J, Kwait David M, Hughes Chambers V, McNamara Erin J, Ishmael Fauod, O'Donnell Robert W, Chang Ming-Mei, Hogg Michael G, Dordick Jonathan S, Santhanam Lakshmi, Ziegler Linda M, Holland James A
Department of Chemistry, State University of New York College at Geneseo, USA.
Endothelium. 2002;9(3):191-203. doi: 10.1080/10623320213638.
NADPH oxidase is a major enzymatic source of oxygen free radicals in stimulated endothelial cells (ECs). The ortho-methoxy-substituted catechol, apocynin (4-hydroxy-3-methoxyacetophenone), isolated from the traditional medicinal plant Picrorhiza kurroa, inhibits the release of superoxide anion (O2*-) by this enzyme. The compound acts by blocking the assembly of a functional NADPH oxidase complex. The underlying chemistry of this inhibitory activity, and its physiological significance to EC proliferation, have been investigated. A critical event is the reaction of ortho-methoxy-substituted catechols with reactive oxygen species (ROS) and peroxidase. Analysis of this reaction reveals that apocynin is converted to a symmetrical dimer through the formation of a 5,5' carbon-carbon bond. Both reduced glutathione and L-cysteine inhibit this dimerization process. Catechols without the ortho-methoxy-substituted group do not undergo this chemical reaction. Superoxide production by an endothelial cell-free system incubated with apocynin was nearly completely inhibited after a lagtime for inhibition of ca. 2 min. Conversely, O2*- production was nearly completely inhibited, without a lagtime, by incubation with the dimeric form of apocynin. The apocynin dimer undergoes a two-electron transfer reaction with standard redox potentials of -0.75 and -1.34 V as determined by cyclic voltammetry. Inhibition of endothelial NADPH oxidase by apocynin caused a dose-dependent inhibition of cell proliferation. These findings identify a metabolite of an ortho-methoxy-substituted catechol, which may be the active compound formed within stimulated ECs that prevents NADPH oxidase complex assembly and activation.
NADPH氧化酶是受刺激的内皮细胞（ECs）中氧自由基的主要酶源。从传统药用植物胡黄连中分离出的邻甲氧基取代儿茶酚——载脂蛋白（4-羟基-3-甲氧基苯乙酮），可抑制该酶释放超氧阴离子（O2*-）。该化合物通过阻断功能性NADPH氧化酶复合物的组装起作用。已对这种抑制活性的潜在化学机制及其对EC增殖的生理意义进行了研究。一个关键事件是邻甲氧基取代儿茶酚与活性氧（ROS）和过氧化物酶的反应。对该反应的分析表明，载脂蛋白通过形成5,5'碳-碳键转化为对称二聚体。还原型谷胱甘肽和L-半胱氨酸均抑制这种二聚化过程。没有邻甲氧基取代基的儿茶酚不会发生这种化学反应。与载脂蛋白一起孵育的无内皮细胞系统产生超氧化物的过程在约2分钟的抑制延迟时间后几乎完全被抑制。相反，与载脂蛋白的二聚体形式孵育可几乎完全抑制O2*-的产生，且无延迟时间。通过循环伏安法测定，载脂蛋白二聚体经历双电子转移反应，标准氧化还原电位分别为-0.75和-1.34 V。载脂蛋白对内皮NADPH氧化酶的抑制导致细胞增殖呈剂量依赖性抑制。这些发现确定了一种邻甲氧基取代儿茶酚的代谢产物，它可能是在受刺激的ECs内形成的活性化合物，可阻止NADPH氧化酶复合物的组装和激活。
