hERG钾通道功能降低介导心肌梗死后恶性心律失常的发生机制及ω-3多不饱和脂肪酸的防治作用

Life-threatening ventricular arrhythmias and sudden cardiac death (SCD) are the leading causes of death in myocardial infarction (AMI) patients. Presently, the mechanisms underlying AMI-related sudden death are not fully understood, and effective preventive measures are lacking. QT interval prolongation, observed on the surface electrocardiogram (ECG), is common in AMI patients. Dysfunction of cardiac potassium channel, hERG, delays cardiac repolarization and cause QT interval prolongation, representing most fundamental mechanisms of sudden cardiac death. The stability of membrane hERG channel was closely related with cytosolic Ca2+ or Ca2+-activated cardiac enzymes. ω-3 polyunsaturated fatty acids (PUFAs) have been proved to prevent SCD by reducing Ca2+ overload. Thus, we hypothesize that AMI-related Ca2+ overload and increased Ca2+-activated cardiac enzymes accelerate the degradation of hERG channel, which prolongs QT interval and increases SCD risks. ω-3 PUFAs could reduce SCD risk through decreasing Ca2+ overload, stabilizing hERG channel and shortening QT interval. In the present study, we will use hERG-HEK cells and rabbit AMI model, together with molecular and biological methods (ELISA, flow cytometry, real-time PCR, Western blot, patch clamp, and surface ECG recordings), to investigate causes of dysfunction of hERG channel after AMI, to evaluate the key steps of AMI-related QT prolongation via dysfunction of hERG, and to elucidate the prevention effects and molecular mechanisms of ω-3 PUFAs on AMI-related arrhythmias. These results will extend our understanding of the mechanisms of AMI related ventricular arrhythmias, and will lay a foundation for prevention of AMI related SCD by ω-3 PUFAs.

恶性心律失常、心源性猝死（SCD）是心梗的主要死因，机制尚不明确，也缺乏有效防治措施。心梗后常见心电图QT间期延长。心脏hERG通道降低引起QT间期延长及SCD，高Ca2+和蛋白酶降低hERG通道，ω-3 PUFAs可降低细胞内Ca2+而减少SCD。我们假设心梗后Ca2+超载或心肌酶通过加速hERG通道降解机制，引起QT间期延长，增加心律失常风险；ω-3 PUFAs通过减轻Ca2+超载，稳定细胞膜hERG通道，缩短QT间期而防治心律失常。本研究拟以培养细胞和心梗兔为实验模型，采用心电图、分子生物学、流式细胞仪及膜片钳等方法，阐明心梗后引起hERG通道功能降低、QT间期延长、心律失常发生的关键环节，明确ω-3 PUFAs稳定细胞膜hERG通道而预防心梗后心律失常、减少SCD的分子机制。预期结果将为心梗后心律失常发生机制提供新的证据，也为利用ω-3 PUFAs防治心梗后心律失常奠定理论基础。

恶性心律失常、心源性猝死（SCD）是心梗的主要死因，机制尚不明确，也缺乏有效防治措施。心脏hERG通道的表达及功能降低引起可以引起QT间期延长及SCD。本研究分别采用急性心梗动物模型及和培养细胞为实验对象，采用分子生物学、ELISA、流式细胞仪、膜片钳及心电图等方法，探讨了hERG通道功能降低是否介导心梗后恶性心律失常发生的分子机制。我们发现急性心梗患者心电图QTc间期逐渐延长，于24-72h达峰值。进一步发现家兔急性心梗后心电图QT间期显著延长，且心梗兔心肌细胞膜ERG通道蛋白显著降低，提示心梗后QT间期延长可能与心肌细胞上ERG/hERG通道蛋白降低有关。考虑到急性心梗后心肌组织存在缺血低氧，我们发现低氧显著降低hERG通道在细胞膜上的表达以及功能。研究显示去乙酰化酶6（HDAC6）的去乙酰化修饰作用能降低离子通道的稳定性，我们发现低氧显著增加HDAC6蛋白表达及HDAC6酶活性。免疫共沉淀实验显示HDAC6与hERG存在相互作用，过表达HDAC6降低hERG蛋白表达，而抑制或敲减HDAC6则明显增加hERG蛋白的表达及功能。进一步探讨HDAC6降低hERG通道的机制发现，HDAC6通过增加hERG蛋白的泛素化而降低hERG通道的表达，构建突变破坏hERG通道位于胞浆侧的乙酰化修饰位点可以彻底消除低氧及HDAC6诱导的hERG通道泛素化降解。进一步发现心梗后小鼠心脏组织ERG蛋白泛素化水平增加，ERG蛋白表达显著降低，而HDAC6表达则显著增加，抑制HDAC6表达能降低ERG蛋白的泛素化降解，增加ERG的表达，提示HDAC6诱导的ERG泛素化降解介导心肌梗死后ERG的下调。这些结果一方面有助于深入理解心梗患者恶性心律失常或SCD发作的机制，另一方面也为心梗患者心律失常或猝死防治提供了新的针对性靶点。