Molecular mechanisms of stretch-induced electrical remodeling in the heart

项目摘要 室性快速性心律失常是心脏性猝死（SCD）的主要原因。虽然导致肥大和失代偿性心力衰竭的机械负荷改变是致命性室性心律失常的最强大的风险因素之一，但对心脏机械功能障碍与电功能障碍之间的联系机制仍知之甚少。因此，SCD仍然是一个尚未解决的重大公共卫生问题。申请人心室电重构（ventricular electrical remodeling，VER）是心室肌电生理特性的一种持续性变化，是由局部机械牵张通过机械-电反馈机制引起的。他实验室的初步数据显示，起搏犬心脏的高应变心肌中Gq/11-蛋白偶联机械感受器通路的表达发生了显著变化。考虑到该通路在心脏机械传导和压力超负荷性肥大中的重要作用，它是解释VER的理想候选者。申请人假设心肌牵张通过增强的Gq/11激活引起VER和心律失常。在凯斯西储大学的现任职位上，他设计了一种斑马鱼全心脏拉伸系统，该系统将高精度拉伸和高分辨率电生理学与完整脊椎动物心脏中的高通量基因操作相结合。来自野生型和转基因斑马鱼胚胎心脏的微囊在场起搏期间通过附加的10微米直径的薄碳纤维拉伸数小时以诱导持续性VER。拉伸后，使用高分辨率荧光成像测定VER。申请人的初步数据表明，他的斑马鱼模型再现了他的导师的VER犬模型的关键电生理表型。申请人将使用他的斑马鱼VER模型来确定 (1)诱发VER的关键生物力学和生物电条件， (2)牵张诱导的VER如何增强心脏对心律失常的易感性， (3)导致牵张诱发性VER的分子信号机制。 这些研究将有助于确定预防和治疗心脏病患者VER和相关心律失常的新治疗靶点。通过K99/R 00奖对该项目的支持将在候选人的职业发展中发挥关键和必要的作用。他的近期目标是： (1)通过本提案直接产生的额外强化指导、技术培训和广泛的智力发展，巩固他的研究经验。申请人在生物物理学和实验模型开发方面有相当丰富的经验，但需要在分子和综合生物学方面进行额外的培训，以发展心血管生物学的独立学术生涯。 (2)在5年奖励期结束前成功获得R 01资助。 高度结构化的职业发展计划是该提案的内在组成部分，旨在大大提高候选人长期职业目标的实现：建立一个充满活力，独立资助的跨学科研究计划，结合生物医学工程，高通量遗传学和高分辨率电生理学研究心律失常的分子机制。

Project Summary Ventricular tachyarrhythmias are a principal cause of sudden cardiac death (SCD). While altered mechanical loading leading to hypertrophy and decompensated heart failure is one of the most powerful risk factors for fatal ventricular arrhythmias, the mechanisms that link mechanical to electrical dysfunction in the heart remain poorly understood. Consequently, SCD remains a major unresolved public health problem. The applicant¿s mentor recently reported that ventricular electrical remodeling (VER), a persistent change in the electrophysiological properties of ventricular myocardium in response a change in activation sequence, is triggered by local mechanical stretch via a mechano-electrical feedback mechanism. Preliminary data from his laboratory revealed significant expression changes in the Gq/11-protein coupled mechanoreceptor pathway in high-strain myocardium from paced canine hearts. Considering the prominent roles of this pathway in cardiac mechanotransduction and pressure-overload hypertrophy, it is an ideal candidate to explain VER. The applicant hypothesizes that myocardial stretch causes VER and arrhythmias via enhanced Gq/11 activation. In his current position at Case Western Reserve University, he engineered a zebrafish whole-heart stretch system that combines high-precision stretch and high-resolution electrophysiology with high-throughput gene manipulation in an intact vertebrate heart. Ventricles from wild-type and genetically modified zebrafish embryo hearts are stretched during field-pacing for several hours via attached 10-micrometer-diameter thin carbon fibers to induce persistent VER. Following stretch, VER is determined using high-resolution fluorescence imaging. The applicant¿s preliminary data show that his zebrafish model reproduces the key electrophysiological phenotypes of his mentor¿s canine model of VER. The applicant will use his zebrafish VER model to determine (1) the key biomechanical and bioelectrical conditions that induce VER, (2) how stretch-induced VER enhances the susceptibility of the heart to arrhythmias and (3) the molecular signaling mechanisms that cause stretch-induced VER. These studies will help identify new therapeutic targets for preventing and treating VER and associated arrhythmias in patients with heart disease. Support for this project via a K99/R00 award would play a pivotal and requisite role in the candidate¿s career development. His immediate goals are: (1) to solidify his research experience through additional intensive mentorship, technical training and broad intellectual development that will result directly from this proposal. The applicant has considerable experience in biophysics and experimental model development, but requires additional training in molecular and integrative biology to develop an independent academic career in cardiovascular biology. (2) to successfully obtain R01 funding by the end of the 5-year award period. A highly structured career development plan is an intrinsic component of this proposal and is designed to greatly enhance the accomplishment of the candidate¿s long-term career goal: to establish a vibrant, independently funded, interdisciplinary research program that combines biomedical engineering, high-throughput genetics and high-resolution electrophysiology to study the molecular mechanisms of cardiac arrhythmias.