Molecular Determinants of Regional Differences in Human Ventricular Repolarization and Remodeling

摘要 仅在美国就有超过500万成年人患有心力衰竭， 增加室性心律失常和猝死的风险。分子、细胞和系统机制 然而，人们对心力衰竭与猝死风险增加之间的联系知之甚少， 尽管我们付出了大量的努力和精力，但对患者进行风险分层仍然是一个巨大的挑战。虽然许多实验 （动物/细胞）心力衰竭模型已经被开发和广泛研究，仅有限地了解到 已经提供了人类心律失常机制。为了改变这一点并推动这一领域的发展，我们 进行了全面的研究工作，旨在确定参与生理机制， 人心脏膜兴奋性的调节与病理生理性电重构 与人类心力衰竭有关。利用转化心血管 华盛顿大学的生物库和储存库，用于获取非失败和失败的人类心脏，我们 已经建立了用于分离和体外维持人心室肌细胞的稳健、可靠的方法， 肌细胞在这里，我们利用这些独特的资源来直接测试假设， 三种电压依赖性传导通路的调节和重塑的差异对 通过心室的活动的协调传播和正常心律的维持： Kv4.3编码的快速瞬态外向K+电流，Ito，f;最近发现的新型非失活Kv电流 Nav1.5编码的电压门控Na+电流，INa。在目标#1中，我们将定义函数 异质性Ito，f重构对LV动作电位波形的影响，并确定其分子机制。 非衰竭人LV中天然Ito，f通道和衰竭人LV中Ito，f重塑的决定因素。在aim中 #2，我们将检验这一假设，即在表达和重塑中也存在跨壁差异， 并定义了细胞类型特异性差异在Iss表达和重塑中的功能后果 左心室动作电位波形目标#3中的实验将检验跨壁 Nav1.5编码的INa通道的表达、性质和重塑的差异，特别是晚期 非衰竭和衰竭人LV肌细胞中INa、INa、L的组分，并定义对LV的功能影响 动作电位波形的异质性INa，L表达和重构。 这些研究将提供新的，临床相关的，对细胞/分子机制的见解， 对人心室肌Ito、f、Iss和INa的生理调节和病理生理重构的影响 渠道这些见解将改变人类心肌细胞和整个心脏模型的完善， 转化为新的，基于机制的策略，以靶向特定的细胞类型，以减少危及生命的风险， 患有人类心力衰竭的患者的室性心律失常。

ABSTRACT Heart failure, which afflicts more than five million adults in the United States alone, is associated with markedly increased risk of ventricular arrhythmias and sudden death. The molecular, cellular and systemic mechanisms linking heart failure to increased sudden death risk, however, are poorly understood and, despite considerable attention and effort, risk stratifying patients remains an enormous challenge. Although numerous experimental (animal/cellular) heart failure models have been developed and extensively studied, only limited insights into human arrhythmia mechanisms have been provided. Motivated to change this and advance the field, we have undertaken a comprehensive research effort aimed at defining the mechanisms involved in the physiological regulation of membrane excitability in the human heart and the pathophysiological electrical remodeling associated with human heart failure. Utilizing the infrastructure developed in the Translational Cardiovascular Biobank and Repository at Washington University for the acquisition of non-failing and failing human hearts, we have established robust, reliable methods for the isolation and in vitro maintenance of human ventricular myocytes. Here, we utilize these unique resources to test directly the hypothesis that there are regional differences in the regulation and remodeling of three voltage-dependent conductance pathways critical for the coordinated propagation of activity through the ventricles and the maintenance of normal cardiac rhythms: the Kv4.3-encoded, fast transient, outward K+ current, Ito,f; the recently identified, novel, non-inactivating Kv current component, Iss; and, the Nav1.5-encoded voltage-gated Na+ current, INa. In aim #1, we will define the functional impact of heterogeneous Ito,f remodeling on LV action potential waveforms, and identify the molecular determinants of native Ito,f channels in non-failing human LV and of Ito,f remodeling in failing human LV. In aim #2, we will test the hypothesis that there are also transmural differences in the expression and the remodeling of Iss, and define the functional consequences of cell type-specific differences in Iss expression and remodeling on LV action potential waveforms. Experiments in aim #3 will test the hypothesis that there are transmural differences in the expression, properties and remodeling of Nav1.5-encoded INa channels, particularly the late component of INa, INa,L, in non-failing and failing human LV myocytes and define the functional impact on LV action potential waveforms of heterogeneous INa,L expression and remodeling. These studies will provide new, clinically relevant, insights into the cellular/molecular mechanisms contributing to the physiological regulation and pathophysiological remodeling of native human ventricular Ito,f, Iss and INa channels. These insights will transform the refinement of human cardiac myocyte and whole heart models and translate to novel, mechanism-based strategies to target specific cell types to reduce the risk of life-threatening ventricular arrhythmias in patients suffering human heart failure.