Integrins and Caveolin Proteins in Cardiac Hypertrophy and Failure

描述(由申请人提供)：心肌细胞(CM)感知机械变化并将其转化为生化和功能效应的机制尚不清楚。CM中的两个区域似乎对适当的机械转导至关重要，这两个区域是带有常驻整合素(ITGs)的胞间和小凹，即烧瓶状的膜内陷和相关的小窝蛋白(Cav)。我们和其他人已经开始表明CM ITGs和Cav蛋白在传递机械信号中的重要性，并且ITGs和Cav3的表达和/或突变减少可导致心肌病。ITGs和Cav-3如何在功能上相互作用，以及它们的协调反应通过什么机制在CM中传递机械信号尚不清楚。我们已经建立了1 ITG CM特异性基因敲除(KO)(称为1cKO)、Cav-3KO和CM特异性ITG和Cav-3转基因(TG)小鼠模型。我们的初步数据显示，ITG和Cav-3在成人CM中共定位和共沉淀，在CMS中增加ITGs和Cav-3蛋白的表达可以在心脏受到增加的机械负荷挑战时保护心肌功能。基于这一数据，我们提出了我们的总体假设：1)ITGs和Cav-3在CM机械转导所需蛋白质的组织方面具有合作特性，以及B)CM中这两种蛋白质的表达增加将保护因HF而面临血流动力学负荷增加的心脏。我们将通过功能丧失和功能获得的方法来实现这一目标：1)通过单独使用Cav3基因缺失和1ITG心肌细胞特异性基因敲除小鼠，或联合使用，确定1ITGs和小窝蛋白-3在心肌力传导反应中的作用。我们将评估当ITGs、Cav-3或两者都从CM中丢失时，急性和慢性机械介导的信号是如何改变的，以及当心脏肥厚并向HF表型进化时，这些蛋白的丢失如何改变重塑过程。2)明确指导心肌细胞ITG和Cav-3协同机械信号传导的相互作用。将评估ITGs和Cav3与胞核和细胞骨架蛋白之间的直接相互作用。然后，蛋白质组学将确定与ITGs或Cav3相互作用的新蛋白质，以及CM的凹陷和非凹陷生化部分的蛋白质如何随着血流动力学的挑战而变化。3)确定在肥厚诱导下，Cav-3和/或1ITGs的TG过表达如何改变机械反应，延缓或保护心脏免受心力衰竭的影响。这将使我们能够直接测试Cav3和？1ITGs在机械挑战心肌中的潜在治疗作用。临床意义：在大量患者中发现各种原因的心衰，需要频繁的住院和门诊治疗。确定心脏功能不全的根本原因，以及重要的是，可能导致这种常见疾病的新疗法的研究，是必不可少的，也将是本提案的重点。

DESCRIPTION (provided by applicant): The mechanisms by which the cardiac myocyte (CM) senses mechanical changes and converts it to biochemical and functional effects, are poorly understood. Two areas within the CM that appear critical for proper mechanotransduction are the costamere with its resident integrins (ITGs), and caveolae, flask-shaped membrane invaginations with associated Caveolin (Cav) proteins. We and others have begun to show the importance of CM ITGs and Cav proteins in transducing mechanical signals, and that reduced expression and/ or mutation of ITGs and Cav3 can cause cardiomyopathies. How ¿1 ITGs and Cav-3 functionally interact and by what mechanisms their coordinated responses transduce mechanical signals in the CM are unknown. We have generated ¿1 ITG CM-specific knockout (KO) (termed ¿1cKO), Cav-3KO and CM-specific ITG and Cav-3 transgenic (Tg) mouse models. Our preliminary data shows that ¿1 ITG and Cav-3 co-localize and co-immunoprecipitate in the adult CM and that increased expression of ITGs and Cav-3 proteins in CMs can both preserve myocardial function when the heart is challenged with an increased mechanical load. Based on this data, we formulated our overall hypotheses that: A) ¿1 ITGs and Cav-3 have cooperative properties in organization of proteins required for CM mechanotransduction and B) that increased expression of both of these proteins in the CM will protect the heart challenged with an increased hemodynamic load from HF. We will pursue this with 3 aims using loss and gain of function approaches: 1) Determine roles of ¿1 ITGs and caveolin-3 in mechanotransductive responses of the myocardium by use of Cav3 null and ¿1 ITG cardiac myocyte specific knockout mice alone, or in combination. We will evaluate how acute and chronic mechanically mediated signals are altered when ITGs, Cav-3 or both proteins are lost from the CM and how loss of these proteins modifies the remodeling process as the heart hypertrophies and evolves towards a HF phenotype. 2) Define the interactions which direct cooperative mechanical signaling of cardiomyocyte ¿1 ITG and Cav-3. Direct interactions between ITGs and Cav3, with costameric and cytoskeletal proteins will be assessed. Proteomics will then identify novel proteins which interact with ITGs or Cav3, and how proteins in the caveolar and non-caveolar biochemical fraction of the CM change with hemodynamic challenge. 3) Determine how Tg overexpression of Cav-3 and/ or ¿¿1 ITGs alters mechanical responses, and delays or protects the heart from cardiac failure in the face of hypertrophic induction. This will allow us to directl test the potential therapeutic roles of Cav3 and ¿1 ITGs in the mechanically challenged myocardium. Clinical Significance: HF of varied causes is found in a large number of patients, necessitating frequent inpatient and outpatient care. Identification of root causes of cardiac dysfunction and importantly, studies which could lead to novel therapeutics of this common disease, are essential, and will be the focus of this proposal.