Calcium-independent enhancement of contractility in normal and failing cardiomyoc

正常和衰竭心肌中非钙依赖性的收缩力增强

• 批准号: 7531746
• 负责人: Charles E Murry
• 金额: $ 19.5万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-15 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7531746
• 关键词: 2' -deoxyadenosine triphosphate; Actins; Adrenergic Agents; Adult; Adverse effects; Affect; Animal Model; Area; Binding; Calcium; Cardiac; Cardiac Myocytes; Cause of Death; Cell Survival; Condition; Contractile Proteins; Data; Development; Drug effect disorder; Enzymes; Goals; Heart; Heart Diseases; Heart failure; Homeostasis; In Situ; In Vitro; Infarction; Kinetics; Longitudinal Studies; Microfilaments; Myocardial Infarction; Myocardium; Myosin ATPase; Performance; Pharmacologic Substance; Phosphorylation; Preparation; Production; Property; Protein Isoforms; Public Health; Rattus; Relaxation; Rest; Ribonucleotide Reductase; Sarcomeres; Skin; Study Section; Systole; Therapeutic; Thin Filament; Tissues; Transfection; Troponin C; United States; Viral; Viral Vector; adenoviral-mediated; adrenergic; base; gene therapy; heart function; improved; mutant; nucleotide analog

DESCRIPTION (provided by applicant): Heart disease is the leading cause of death in the United States and has been rising dramatically around the world. This is an exploratory application to develop strategies for improving cardiac performance by targeting the contractile apparatus of cardiomyocyte sarcomeres. While the current application focuses on in vitro studies, the long term goal is to enhance systolic function without compromising diastolic function of the heart, and still allow responsiveness to adrenergic stimulation. We will target the cardiac thin filament activation and the actin-myosin `crossbridge' cycle directly, such that cardiomyocyte contraction (but not relaxation) is enhanced without the need for increased intracellular [Ca2+]. To increase thin filament activation at a given [Ca2+] we will replace native troponin C in myofilaments with a mutant TnC (L48Q) that has enhanced Ca2+ binding properties (aim 1). To enhance crossbridge cycling we will increase the cellular production of 2 deoxy-ATP (dATP) via increased expression of the enzyme that converts ATP to dATP in cardiomyocytes (ribonucleotide reductase; RR) (aim 2). We have provided significant data demonstrating these approaches improve contractility in demembranated (skinned) cardiac tissue, without affecting relaxation kinetics or resting stiffness. For the proposed studies we will use a viral transfection strategy to determine if similar increases in cardiac contractility occur in intact adult cardiomyocytes in culture. We will also determine whether these approaches affect sarcomere contractile protein isoform and phosphorylation profiles or the level of intracellular [Ca2+] during stimulated activation and in relaxation. In the second year of the proposal we will begin development of animal models for an expanded proposal to study how these manipulations influence whole heart function in situ and in vitro in normal hearts and under pathological conditions. PUBLIC HEALTH RELEVANCE: Heart failure at its base is a reduction in cardiac myofilament contractility. Most current therapies focus on mechanisms that enhance intracellular Ca2+ during systole which, among other things, can affect diastolic function. To avoid this, our proposal targets myofilaments directly to enhance contraction without the need for increased intracellular Ca2+.

描述（由申请人提供）：心脏病是美国的首要死因，并且在世界范围内呈急剧上升趋势。这是一项探索性应用，旨在通过针对心肌细胞肌节的收缩装置来制定改善心脏性能的策略。虽然目前的应用侧重于体外研究，但长期目标是在不损害心脏舒张功能的情况下增强收缩功能，并且仍然允许对肾上腺素能刺激做出反应。我们将直接针对心脏细丝激活和肌动蛋白-肌球蛋白“横桥”循环，从而增强心肌细胞收缩（但不是松弛），而不需要增加细胞内 [Ca2+]。为了增加给定 [Ca2+] 下的细丝激活，我们将用具有增强 Ca2+ 结合特性的突变体 TnC (L48Q) 取代肌丝中的天然肌钙蛋白 C（目标 1）。为了增强跨桥循环，我们将通过增加心肌细胞中将 ATP 转化为 dATP 的酶（核糖核苷酸还原酶；RR）的表达来增加细胞内 2 脱氧 ATP (dATP) 的产生（目标 2）。我们提供的重要数据表明这些方法可以改善脱膜（带皮）心脏组织的收缩性，而不影响松弛动力学或静息僵硬度。对于拟议的研究，我们将使用病毒转染策略来确定培养的完整成年心肌细胞中是否会出现类似的心肌收缩力增加。我们还将确定这些方法是否影响肌节收缩蛋白亚型和磷酸化谱或刺激激活和放松期间细胞内 [Ca2+] 的水平。在该提案的第二年，我们将开始开发动物模型，以扩大提案的范围，以研究这些操作如何在正常心脏和病理条件下原位和体外影响整个心脏功能。公众健康相关性：心力衰竭的根本原因是心肌丝收缩力的降低。目前大多数疗法侧重于增强收缩期细胞内 Ca2+ 的机制，这尤其会影响舒张功能。为了避免这种情况，我们的建议直接针对肌丝来增强收缩，而不需要增加细胞内 Ca2+。