Wang Jingxiong, Wang Huizhen, Zhang Yiqiang, Gao Huanhuan, Nattel Stanley, Wang Zhiguo
Research Center, Montreal Heart Institute, Montreal, Quebec H1T 1C8.
J Biol Chem. 2004 Apr 2;279(14):13289-92. doi: 10.1074/jbc.C400025200. Epub 2004 Feb 17.
Congestive heart failure (CHF) is associated with susceptibility to lethal arrhythmias and typically increases levels of tumor necrosis factor-alpha (TNF-alpha) and its receptor, TNFR1. CHF down-regulates rapid delayed-rectifier K(+) current (I(Kr)) and delays cardiac repolarization. We studied the effects of TNF-alpha on cloned HERG K(+) channel (human ether-a-go-go-related gene) in HEK293 cells and native I(Kr) in canine cardiomyocytes with whole-cell patch clamp techniques. TNF-alpha consistently and reversibly decreased HERG current (I(HERG)). Effects of TNF-alpha were concentration-dependent, increased with longer incubation period, and occurred at clinically relevant concentrations. TNF-alpha had similar inhibitory effects on I(Kr) and markedly prolonged action potential duration (APD) in canine cardiomyocytes. Immunoblotting analysis demonstrated that HERG protein level was slightly higher in canine hearts with tachypacing-induced CHF than in healthy hearts, and TNF-alpha slightly increased HERG protein level in CHF but not in healthy hearts. In cells pretreated with the inhibitory anti-TNFR1 antibody, TNF-alpha lost its ability to suppress I(HERG), indicating a requirement of TNFR1 activation for HERG suppression. Vitamin E or MnTBAP (Mn(III) tetrakis(4-benzoic acid) porphyrin chloride), a superoxide dismutase mimic) prevented, whereas the superoxide anion generating system xanthine/xanthine oxidase mimicked, TNF-alpha-induced I(HERG) depression. TNF-alpha caused robust increases in intracellular reactive oxygen species, and vitamin E and MnTBAP abolished the increases, in both HEK293 cells and canine ventricular myocytes. We conclude that the TNF-alpha/TNFR1 system impairs HERG/I(Kr) function mainly by stimulating reactive oxygen species, particularly superoxide anion, but not by altering HERG expression; the effect may contribute to APD prolongation by TNF-alpha and may be a novel mechanism for electrophysiological abnormalities and sudden death in CHF.
充血性心力衰竭（CHF）与致死性心律失常易感性相关，通常会使肿瘤坏死因子-α（TNF-α）及其受体TNFR1的水平升高。CHF会下调快速延迟整流钾电流（I(Kr)）并延迟心脏复极。我们采用全细胞膜片钳技术研究了TNF-α对HEK293细胞中克隆的HERG钾通道（人ether-a-go-go相关基因）以及犬心肌细胞中天然I(Kr)的影响。TNF-α持续且可逆地降低HERG电流（I(HERG)）。TNF-α的作用呈浓度依赖性，随着孵育时间延长而增强，且在临床相关浓度下即可出现。TNF-α对犬心肌细胞中的I(Kr)有类似的抑制作用，并显著延长动作电位时程（APD）。免疫印迹分析表明，快速起搏诱导的CHF犬心脏中HERG蛋白水平略高于健康心脏，TNF-α使CHF心脏中的HERG蛋白水平略有升高，但对健康心脏无此作用。在用抑制性抗TNFR1抗体预处理的细胞中，TNF-α失去了抑制I(HERG)的能力，表明HERG抑制需要TNFR1激活。维生素E或MnTBAP（锰（III）四（4-苯甲酸）卟啉氯化物，一种超氧化物歧化酶模拟物）可预防TNF-α诱导的I(HERG)抑制，而超氧阴离子生成系统黄嘌呤/黄嘌呤氧化酶则模拟了TNF-α诱导的I(HERG)抑制。TNF-α导致细胞内活性氧大量增加，维生素E和MnTBAP在HEK293细胞和犬心室肌细胞中均消除了这种增加。我们得出结论，TNF-α/TNFR1系统主要通过刺激活性氧，尤其是超氧阴离子来损害HERG/I(Kr)功能，而非通过改变HERG表达；这种作用可能导致TNF-α延长APD，并且可能是CHF中电生理异常和猝死的新机制。
