长链非编码RNA KCNA2-AS靶向调控kv1.2钾通道在缺血性慢性心力衰竭室性心律失常发生中的作用研究

Ventricular arrhythmias are the leading cause of sudden cardiac death in patients with ischemic chronic heart failure (CHF). Reduced density of delayed rectifier K+ currents (IK) has been considered as an important mechanism underlying ventricular arrhythmias in ischemic CHF. Down-regulation of Kv1.2 can cause reduced IK, and many antiarrhythmic drugs have been shown to act via Kv1.2. In our previous study, we observed that the expression level of Kv1.2 was decreased in ischemic CHF. Recently, accumulating evidence suggests that long noncoding RNA (lncRNA) plays crucial roles in gene regulation. We had successfully identified a novel antisense lncRNA termed KCNA2-AS, which could specially down-regulate the expression of Kv1.2. We also observed increased level of KCNA2-AS in ischemic CHF. In the present study, we will first identify the impacts of KCNA2-AS on ventricular arrhythmias in a guinea pig model of ischemic CHF, and then investigate the molecular and electrophysiological mechanism by which the KCNA2-AS/Kv1.2 pathway influences ventricular arrhythmogenesis on both the cellular and whole animal level. Finally, we will explore the potential role of MZF-1 in regulating the expression of KCNA2-AS in ischemic CHF. Our studies are likely to add to our understanding of the mechanisms of ventricular arrhythmia in ischemic CHF, and may contribute to the design of new drugs that are clinically useful to the treatment of this disorder.

缺血性慢性心衰（CHF）患者易发生室性心律失常导致猝死，延迟整流钾电流（IK）下降是其重要因素。Kv1.2表达下调可导致IK降低，是多种抗心律失常药的作用靶点。我们研究发现缺血性CHF心肌Kv1.2表达明显降低。长链非编码RNA（lncRNA）在基因调控中起重要作用。我们已克隆出一个新lncRNA KCNA2-AS，其可靶向下调Kv1.2表达，且在缺血性CHF心肌表达明显升高。我们将在前期工作基础上制备豚鼠缺血性CHF模型并使心肌过表达/沉默KCNA2-AS，研究KCNA2-AS对缺血性CHF室性心律失常发生的影响；在整体和细胞两个水平研究KCNA2-AS/Kv1.2通路影响缺血性CHF室性心律失常发生的分子及电生理机制；最后从转录因子MZF-1角度探讨缺血性CHF时KCNA2-AS表达上调的可能机制。本研究不仅可以加深理解缺血性CHF室性心律失常发生机制，还可为临床治疗提供新的药物靶点。

背景：慢性心衰（CHF）患者易发生室性心律失常导致猝死，缓慢型延迟整流钾电流（Iks）下降使动作电位延长是其重要因素,但其具体分子作用机制尚未明确。.目的：近年来在神经系统发现一条新的内源性长链非编码RNA名为Kcna2-AS，它与Kcna2（编码Kv1.2钾通道的基因）的大部分序列互补，并能反向调节Kv1.2表达。我们在大鼠慢性心衰模型中通过Kcna2-AS靶向调节Kv1.2的表达来观察Iks、动作电位（AP）、心电图变化，探讨心衰后室性心律失常发生的确切机制，为今后临床应用提供客观依据。.方法与结果：利用qRT-PCR和Western blot技术发现慢性心衰模型心肌组织和心肌细胞肥大模型中Kcna2-AS表达较正常组升高，全细胞膜片钳技术记录到Kv1.2表达下降伴随Iks密度降低、AP延长（P<0.05）。运用腺相关病毒AAV9载体将心衰大鼠Kcna2过表达后，下降的Iks密度增大、延长的AP缩短（P<0.05）。把Kcna2沉默后，这些现象消失。将心衰大鼠Kcna2-AS沉默后，Kv1.2表达升高、下降的Iks密度增大、延长的AP缩短（P<0.05）。而将Kcna2-AS过表达后，这些现象消失且Kv1.2表达更低（P<0.05）。心电图显示：把心衰大鼠Kcna2-AS沉默或Kcna2过表达后室性心律失常事件易发性减少，而把Kcna2-AS过表达或Kcna2沉默后室性心律失常事件易发性增大（P<0.05）。.结论：发生慢性心衰时，长链非编码RNA Kcna2-AS能够靶向下调Kv1.2钾通道表达，从而使Iks电流减少，导致3期复极化延长后动作电位时程增大，增加室性心律失常事件的易发性。所以在慢性心衰时下调Kcna2-AS可增加Kv1.2的表达，上调Iks电流密度，缩短动作电位延长，从而降低室性心律失常事件的发生风险。本研究将为今后临床治疗和预防心衰后室性心律失常提供新的靶点。

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