Arrhythmia Mechanisms in Sarcomeric Cardiomyopathies

描述(申请人提供)：肌丝钙敏感性增加是导致家族性肥厚型心肌病(HCM)的肌瘤突变的共同特征，心肌梗死(MI)后也描述了同样的钙敏感性增加。这两种疾病都与室性心律失常和猝死的高风险相关，但肌丝钙敏感性和心律失常易感性的机制仍然知之甚少。这一更新应用将研究肌丝钙敏感性和触发的心律失常之间的机制联系。在上一个资助周期中，我们发现两个钙增敏肌钙蛋白T(TnT)突变(I79N、F110I)和钙增敏化合物EMD57033都显著增加了胞浆的钙结合亲和力(即表观KD)，但对胞浆最大缓冲容量(Bmax)没有影响。这些数据首次建立了TNT突变对胞浆钙缓冲特性的直接影响。在生理心率下，肌丝钙结合亲和力增加的主要结果是胞浆中舒张末钙的增加。令人惊讶的是，与先前的预测相反，肌浆网(SR)中的钙含量并没有减少。相反，肌丝钙结合增加的净效应是在心率加快期间胞浆中钙的积累，在短暂停顿后导致肌浆网钙超载、动作电位(AP)延长和早期后除极。再加上我们在HCM和MI后小鼠模型中发现的停顿依赖性触发严重室性心律失常的新结果，这些具有挑衅性的新结果使我们提出了以下假设：肌丝钙结合亲和力增加是导致停顿依赖性SR钙超载的根本缺陷，从而增加触发心律失常的易感性。目标1中的实验将测试我们的假设是否可以推广到导致遗传性心肌病(即家族性肥厚性心肌病或扩张型心肌病[DCM])的其他肌节突变。目标2将确定增加的肌丝钙结合如何改变不同物种的钙处理和AP调节。目的3将测试肌丝钙结合增加是否会导致获得性心脏病(即缺血性心肌病)的心律失常风险。目的1.验证肌丝钙敏感性改变导致肌丝钙结合亲和力和胞浆钙缓冲目标发生一致性变化的假说2.确定肌丝钙结合亲和力增加对不同物种心肌细胞钙稳态和动作电位调节的影响目的3.验证肌丝钙结合增加导致慢性心肌梗死后触发心律失常的假说

DESCRIPTION (provided by applicant): Increased myofilament Ca sensitivity is a common feature of sarcomeric mutations that cause familial hypertrophic cardiomyopathy (HCM) and the same Ca sensitivity increase has also been described after myocardial infarction (MI). Both diseases are associated with a high risk for ventricular arrhythmia and sudden death, but the mechanisms linking myofilament Ca sensitivity and arrhythmia susceptibility remain poorly understood. This renewal application will investigate the mechanistic link between myofilament Ca sensitivity and triggered arrhythmia. During the last funding cycle, we discovered that two Ca-sensitizing Troponin T (TnT) mutations (I79N, F110I) and the Ca sensitizing compound EMD57033 both significantly increase the Ca binding affinity of the cytosol (i.e., apparent Kd), but have no effect on the maximal cytosolic buffering capacity (i.e., Bmax). These data establish for the first time a direct effect of the TnT mutations on cytosolic Ca buffering properties. The main consequence of the increased myofilament Ca binding affinity at physiological heart rates was an increase in end-diastolic Ca in the cytosol. Surprisingly, and in contrast to previous predictions, Ca content in the sarcoplasmic reticulum (SR) was NOT reduced. Rather, the net effect of increased myofilament Ca binding was the accumulation of Ca in the cytosol during periods of rapid heart rates, which after brief pauses then led to SR Ca overload, action potential (AP) prolongation and early afterdepolarizations. Together with our finding of pause-dependent triggering of serious ventricular arrhythmia both in HCM and post-MI mouse models, these provocative new results have led us to formulate the following hypothesis: Increased myofilament Ca binding affinity is a fundamental defect that causes pause-dependent SR Ca overload and thereby increases triggered arrhythmia susceptibility. Experiments in Aim 1 will test whether our hypothesis can be generalized to other sarcomeric mutations that cause inherited cardiomyopathies (i.e., familial HCM or dilated cardiomyopathy [DCM]). Aim 2 will determine how increased myofilament Ca binding alters Ca handling and AP regulation in different species. Aim 3 will test whether increased myofilament Ca binding causes arrhythmia risk in acquired heart disease (i.e., ischemic cardiomyopathy). Aim 1. To test the hypothesis that sarcomeric mutations that change myofilament Ca sensitivity cause concordant changes in myofilament Ca binding affinity and cytosolic Ca buffering Aim 2. To determine the effect of increased myofilament Ca binding affinity on myocyte Ca homeostasis and action potential regulation in different species Aim 3. To test the hypothesis that increased myofilament Ca binding contributes to triggered arrhythmia after chronic MI