双孔钾离子通道在低钾血症诱导的心律失常中的病理作用

TWIK-1 two-pore domain potassium channels are highly expressed in the human heart, but their cardiac functional roles are unknown. Hypokalemia causes cardiac arrhythmias, but its pathological mechanism is not well understood. Our recent study indicated that in hypokalemia TWIK-1 potassium channels change ion selectivity, become permeable to external sodium, and conduct inward leak sodium currents, contributing to cardiac paradoxical depolarization, a well-known abnormal behavior of human cardiomyocytes. In order to further investigate the mechanism of hypokalemia-induced cardiac arrhythmias, we hypothesize that in hypokalemia TWIK-1 potassium channels change function and result in abnormal excitability of human cardiomyocytes, which is crucial to the mechanism of hypokalemia-induced cardiac arrhythmias. To test the hypothesis, we will examine the influences of TWIK-1 potassium channels on electrophysiological properties of human cardiomyocytes. We will employ the cardiomyocytes derived from human induced pluripotent stem cells (iPSC) and human embryonic stem cells (ESC) as models, and study how TWIK-1 potassium channels affect the action potential and resting membrane potentials of human cardiomycytes with strategies in real time PCR, Western Blotting, Lenti-viral vectors, RNAi, electrophysiology, and computer modeling. The proposed research will indicate how TWIK-1 potassium channels precisely regulate electric excitability of cardiac cells in hypokalemia, imply functional roles of TWIK-1 potassium channels in human atrial and ventricular cardiomyocytes, and provide novel insights on etiology of hypokalemia-induced cardiac arrhythmias.

TWIK-1双孔钾离子通道在人心脏高度表达，但对其在人心脏的功能仍知之甚少。低钾血症可诱导心律失常甚至心脏停跳，其病理机制不明。低血钾时心肌细胞静息膜电位表示出矛盾去极化而不是复极化。我们前期研究揭示：低血钾时TWIK-1钾通道变成可通透钠离子，并传导内向钠漏电流，进而引起人心肌细胞的矛盾去极化。但低血钾症诱导心律失常的病理分子机制还有待进一步探讨。为此，我们提出假说：低血钾时TWIK-1钾通道的功能改变导致人心肌细胞兴奋性异常。为了验证这一假说，我们将通过人类干细胞诱导心肌细胞模型，采用病毒载体转染、RNAi干扰, 电生理等手段，从分子和细胞水平剖析低钾血症状况下TWIK-1钾通道对人心肌细动作电位和静息膜电位的调控，阐述TWIK-1钾通道在人心肌细胞的功能。本课题将从TWIK-1钾通道这个新视点为揭示低血钾诱导心律失常的机制奠定基础，并为低血钾症及其并发症的防治提供新的思路。

低血钾症可诱导心律失常甚至心脏停跳，但其病理机制不明。低血钾时人心肌细胞展示出异常的电生理特征。这一异常现象是这类心律失常的关键所在。TWIK-1双孔钾离子通道在人心脏里高度表达，但迄今对其在心脏的功能仍知之甚少。本项研究重点在探讨TWIK-1双孔钾离子通道是如何造成人心肌细胞兴奋性异，并导致心律失常的分子机制。在基金资助下，主要的研究内容、重要结果、关键数据和科学意义总结如下：（1）系统研究了内流整合钾通道Kir2.1 与双孔钾离子通道K2P1协同作用，共同维持心肌细胞静息状态下双重膜电位，揭示了钾通道在低血钾症中的分子机制。（2）探索在低血钾症中功能缺失的Kir2.1钾通道是如何影响双重静息膜电位，从而导致心律失常的致病机制。重点研究了E224G突变位点对Kir2.1钾通道功能的影响。（3）构建模型小鼠，研究过表达KCNK1基因编码的K2P1钾通道在低血钾症诱导的心律失常中的致病机制。（4）在基金资助下，筛查出了一例新发的与心源性猝死和儿茶酚胺敏感性多形性室性心动过速相关的RYR2钙离子通道基因突变；构建了基因工程小鼠，在模式动物层面上验证了该突变为致病基因突变，揭示了突变引发室速和造成心源性猝死的致病机制。在项目执行的四年中，申请立项的研究内容均按计划实施，研究目标已全部完成。发表SCI论文2篇，培养博士后以及博士研究生3人。