Sex Hormones and Cardiac Arrhythmia in Transgenic LQT2 Rabbits

描述（由申请人提供）：心源性猝死（SCD）每年夺去大约35万美国人的生命。新出现的证据表明，遗传易感性SCD的重要作用，然而，分子决定因素仍然难以捉摸。该提案的总体目标是通过应用创新技术和新使用长QT综合征2（LQT 2）的新遗传模型来研究SCD的新机制。这种多管齐下的方法包括研究经激素治疗的青春期前、卵巢切除的LQT 2家兔，以探索性激素对心律失常影响的新机制范例，使用体外和计算机实验方法，将整合新的分子、细胞、组织和理论模型。由于性别和交感神经系统在LQT 2中触发SCD中起关键作用，因此将详细研究性激素和自主因素对SCD风险的影响。本研究的主要目的有四个：目的1：研究性激素对心脏复极的影响（QT间期和心脏不应期）、自发性TdP和心源性猝死的发生率，采用遥测ECG监测和体内侵入性电生理研究;目的2：分析动作电位时程、复极离散度、传导、触发活动和电/机械恢复的异质性，以及使用动作电位和钙瞬变的光学映射的传导阻滞。目标3：以限定（基于光学研究）通过使用蛋白质表达研究和定量实时PCR分析编码复极化钾电流、钙电流和控制Ca++循环的蛋白质的或亚基的基因表达中的性别或性激素相关差异以及肾上腺素能受体，来确定心脏复极化和兴奋中性别差异的分子决定因素，和细胞电生理学研究;目的4：使用膜片钳和荧光指示剂技术，在对照条件下和自主神经受体刺激期间，对从经血管紧张素Ⅱ治疗的LQT 2家兔中分离的肌细胞中的动作电位和钙处理特性进行比较分析。B）使用计算机建模来检查这些LQT 2模型中实验定义的细胞/分子改变如何影响模拟心脏组织中的后去极化和致心律失常底物形成。这些研究将有助于了解触发和维持心律失常的机制，在心脏治疗的兔子，因此导致更好地了解性别相关的心律失常在长QT综合征。公共卫生相关性：转基因兔心律失常模型的表征，性激素的影响，以及突变LQTS基因编码功能障碍的离子通道和危及生命的室性快速性心律失常之间的关系的鉴定，应提供更完整的知识到复极障碍的遗传和电生理因素。这些心室复极改变的研究，应有助于重要的新的见解与获得性心脏疾病，伴随缺血性和非缺血性心肌病和QT延长药物的心脏猝死机制。这一知识的提高应导致更有效的战略，以预防猝死在广泛的遗传性和获得性心脏疾病与有意义的公共卫生利益。

DESCRIPTION (provided by applicant): Sudden cardiac death (SCD) claims the lives of approximately 350,000 Americans each year. Emerging evidence indicates an important role for genetic predisposition to SCD; however, the molecular determinants have remained elusive. The overall goal of this proposal is to investigate new mechanisms that underlie SCD through the application of innovative technology and the novel use of new genetic models of long QT syndrome 2 (LQT2). This multi-pronged approach includes the investigation of hormone-treated prepubertal, ovariectomized LQT2 rabbits to explore new mechanistic paradigms that underlie the effects of sex hormones on cardiac arrhythmias using in vitro and in silico experimental approaches that will integrate novel molecular, cellular, tissue, and theoretical models. Because gender and the sympathetic nervous system plays a key role in triggering SCD in LQT2, the influence of sex hormones and the autonomic factors on SCD risk will be studied in detail. The proposal is composed of Four Specific Aims: Aim 1: To characterize of the sex hormone induced changes in cardiac repolarization (QT duration and cardiac refractory periods), incidence of spontaneous TdP and sudden cardiac death with the use of telemetric ECG monitoring and in vivo invasive electrophysiological studies; Aim 2: To analyze the action potential duration, dispersion of repolarization, conduction, triggered activity and the heterogeneities in electrical/mechanical restitution, and conduction block using optical mapping of action potential and calcium transients. Aim 3: To define (based on the optical studies) the molecular determinants of the gender differences in cardiac repolarization and excitation by analyzing the gender or sex hormone related differences in the expression of genes coding for the or subunits of the repolarizing potassium currents, calcium currents, and proteins that control Ca++ cycling, and the adrenergic receptors with the use of protein expression studies and quantitative real-time PCR, and cellular electrophysiological studies; and Aim 4: To perform a comparative analysis of action potential and calcium-handling properties in myocytes isolated from hormone-treated LQT2 rabbits under control conditions and during autonomic receptor stimulation, using patch-clamp and fluorescent-indicator techniques. b) To use computer modeling to examine how experimentally defined cellular/molecular alterations in these LQT2 models influence afterdepolarizations and arrhythmogenic substrate formation in simulated cardiac tissue. These studies will contribute to the understanding of the mechanisms that trigger and maintain arrhythmias in hormone-treated rabbits, and therefore lead to a better understanding of gender-related arrhythmias in long QT syndrome. PUBLIC HEALTH RELEVANCE: The characterization of transgenic rabbit models for cardiac arrhythmias, the effects of sex hormones, and the identification of the relationship between mutant LQTS genes that encode for dysfunctional ion channels and life-threatening ventricular tachyarrhythmias should provide more complete knowledge into the genetic and electrophysiologic factors involved in repolarization disorders. These studies into altered ventricular repolarization should contribute important new insights into sudden cardiac death mechanisms associated with acquired cardiac disorders that accompany ischemic and nonischemic cardiomyopathy and QT-prolonging drugs. This enhanced knowledge should lead to more effective strategies for prevention of sudden death in a broad spectrum of genetic and acquired cardiac disorders with meaningful public health benefits.