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H_(2)O_(2)对人心房肌细胞I_(Kur)和I_(Ca,L)的浓度依赖性双向影响

张光伟, 谷天祥, 王春, 喻磊, 温倜

中国医科大学附属第一医院心脏外科.辽宁,沈阳 110001；中国医科大学附属第四医院中心实验室.辽宁,沈阳 110001

摘要

该实验旨在观察活性氧(reactive oxygen species,ROS)对人心房肌细胞电生理活动特性的影响。取有心房颤动(atrial fibrillation,AF)和非AF心脏手术患者(各12例)右心耳组织,用酶消化法得到单个心房肌细胞。两组细胞(每组{sl n}=75)分别随机分为三个亚组:对照组({sl n}=12)、H_(2)O_(2)组(0.1、0.2、0.5、0.75、1、2、5、10#mu#mol/LH_(2)O_(2),每个浓度{sl n}=7)和维生素C(ROS清除剂)组(1#mu#mol/L维生素C,{sl n}=7)。实验采用全细胞膜片钳方法记录电生理活动。与非AF对照组相比,AF对照组超快速延迟整流钾电流(ultrarapid delayed rectifier K~(+) current,I_(Kur))和L--型钙电流(L--type calcium current,I_(Ca,L))电流密度(pA/pF)均明显降低(6.27±0.67 vs 3.77±0.56,{sl P}<0.05;6.31±0.60 vs 3.34±0.32,{sl P}<0.05),动作电位时程(action potential duration,APD)(ms)也明显缩短(405±13 vs 354±12,{sl P}<0.05)。在非AF和AF组中,H_(2)O_(2)对心房肌细胞I_(Kur)和I_(Ca,L)的电流密度均有浓度依赖性双向影响-----高抑低促。非AF组中,H_(2)O_(2)浓度为0.2#mu#mol/L时有最大增强作用,而0.75#mu#mol/L为分界浓度,大于0.75#mu#mol/L时,随H_(2)O_(2)浓度增加I_(Kur)和I_(Ca,L)的电流密度逐渐降低;在另一方面,0.2、1、2、5和10#mu#mol/LH_(2)O_(2)孵育的心房肌细胞APD_(90)与同组对照组相比均明显缩短({sl P}<0.05),而与AF对照组相比无明显差异。在AF组中,H_(2)O_(2)的最大效应浓度为0.5#mu#mol/L,而1#mu#mol/L为分界浓度。维生素C可以逆转H_(2)O_(2)的上述作用,但单独给予维生素C并不改变通道特性。H_(2)O_(2)诱导正常人心房肌细胞发生电生理活动特性改变与AF时心肌电重构(atrial electrical remodeling,AER)相似,显示ROS可能诱发AF;同时,H_(2)O_(2)又能加重AF时AER,对AF有维持作用。以上结果提示ROS清除剂可能对预防和治疗AF有重要意义。

关键词： 氧化应激; 心肌; 心房颤动; 离子通道; 抗氧化剂

Two--way concentration--dependent effect of H_(2)O_(2) on I_(Kur) and I_(Ca,L) in human atrial myocytes

Zhang Guangwei, Gu Tianxiang, Wang Chun, Yu Lei, Wen Ti

Department of Cardiac Surgery,the First Hospital of China Medical University.Shenyang 110001,Liaoning；China

Abstract

It has been shown that oxidative stress correlates with atrial fibrillation(AF).The purpose of the present study was to investigate the effects of reactive oxygen species(ROS) on the electrophysiological activity of human atrial myocytes.Right atrial appendages were obtained from patients with AF(AF group,n=12) and without AF(non-AF group,n=12).Single human atrial myocytes were isolated through enzymatic dissociation with type XXIV protease and type V collagenase,then divided into three subgroups:control group(n=12),H_(2)O_(2) group(0.1,0.2,0.5,0.75,1,2,5,10 #mu#mol/L,n=7 at each concentration) and vitamin C(antioxidant) group(1 #mu#mol/L,n=7).Ultrarapid delayed rectifier K~(+) current(I_(Kur)),L-type calcium current(I_(Ca,L)) and action potential duration(APD) were recorded by whole-cell patch clamp.In AF control group,the maximum current densities of I_(Kur) and I_(Ca,L) were significantly lower than that in non-AF control group(both P<0.05) and APD90 was significantly shorter as well(P<0.05).In both non-AF and AF groups,H_(2)O_(2) showed two-way concentration-dependent effect on I_(Kur) and I_(Ca,L).The maximum current densities of I_(Kur) and I_(Ca,L) was significantly increased at lower H_(2)O_(2) concentration,but was decreased at higher H_(2)O_(2) concentration.In non-AF group,0.2 #mu#mol/L H_(2)O_(2) caused a peak increase in the maximum currunt dentities of I_(Kur) [(8.92±0.51) pA/pF,P<0.05] and I_(Ca,L) [(9.32±0.67) pA/ pF,P<0.05].H_(2)O_(2) at a concentration higher than 0.75 #mu#mol/L decreased I_(Kur) and I_(Ca,L).When the H_(2)O_(2) concentrations were 0.2,1,2,5 and 10 #mu#mol/L,APD_(90) was significantly shorter compared with that in non-AF control group(P<0.05),meanwhile it had no significant difference from that in AF control group.In AF group,the peak effective concentration of H_(2)O_(2) was 0.5 #mu#mol/L,and the turning concentration was 1 #mu#mol/L.The H_(2)O_(2) concentration-current density curve in AF group was similar to that in non-AF group,but the turning point shifted to the right,indicating that the way that H_(2)O_(2) acted on ion channels in AF was the same as that in non-AF,however,the sensitivity of ion channels to H_(2)O_(2) was decreased in AF.Vitamin C reversed these changes induced by H_(2)O_(2),and did not affect the characteristics of ion channels.H_(2)O_(2)-induced electrophysiological changes in human atrial myocytes were similar to atrial electrical remodeling(AER) in AF,suggesting that ROS might induce AF.Meanwhile,H_(2)O_(2) also could aggravate AER in AF,contrib-uting to the maintenance of AF.The results suggest that antioxidants might play a significant role in the prevention and treatment of AF.

Key words: oxidative stress;cardiac myocyte;atrial fibrillation;ion channels;antioxidants

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引用本文：

张光伟, 谷天祥, 王春, 喻磊, 温倜. H_(2)O_(2)对人心房肌细胞I_(Kur)和I_(Ca,L)的浓度依赖性双向影响[J]. 生理学报 2008; 60 (6): 695-703.

Zhang Guangwei, Gu Tianxiang, Wang Chun, Yu Lei, Wen Ti. Two--way concentration--dependent effect of H_(2)O_(2) on I_(Kur) and I_(Ca,L) in human atrial myocytes. Acta Physiol Sin 2008; 60 (6): 695-703 (in Chinese with English abstract).