Cardiomyocyte TRPV4 and cardiac dysfunction following ischemia-reperfusion in the aged heart.

衰老心脏缺血再灌注后心肌细胞 TRPV4 和心功能障碍。

• 批准号: 9280032
• 负责人: Timothy Lee Domeier
• 金额: $ 37.57万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-03-03 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9280032
• 关键词: Acidosis; Acute myocardial infarction; Admission activity; Age; American; Arrhythmia; Bioavailable; Blood flow; Calcium; Calcium Signaling; Cardiac; Cardiac Myocytes; Cations; Cell Death; Cell membrane; Cessation of life; Chlorides; Complement; Coronary Arteriosclerosis; Data; Development; Echocardiography; Elderly; Exhibits; Experimental Designs; Fluorescence Microscopy; Functional disorder; Genetic; Goals; Heart; Homeostasis; Hospitals; Individual; Ion Channel; Ischemia; Knockout Mice; Knowledge; Laboratories; Left; Mediating; Membrane Potentials; Metabolic; Metabolic stress; Microscopy; Mitochondria; Modality; Modeling; Monitor; Mus; Muscle Cells; Myocardial Infarction; Myocardial Ischemia; Myocardial dysfunction; Necrosis; Oral; Organ; Pathway interactions; Pharmacology; Phase; Physiological; Pilot Projects; Population; Process; Propidium Diiodide; Reperfusion Injury; Reperfusion Therapy; Research; Resolution; Risk Factors; Role; Signal Transduction; Staining method; Stains; Stimulus; Stress; Testing; Time; Tissues; Transgenic Mice; Transgenic Organisms; Translating; Vanilloid; age group; aged; fluorescence imaging; genetic approach; high risk; imaging approach; in vivo; indexing; inhibitor/antagonist; member; mitochondrial dysfunction; mitochondrial membrane; multi-photon; novel; novel strategies; older patient; overexpression; pressure; prevent; receptor; restoration; sensor; tetramethylrhodamine; therapeutic development; translational study

Project Summary/Abstract Advancing age is the primary risk factor for coronary artery disease and myocardial infarction (MI). A major cause of arrhythmia and tissue damage following MI is reperfusion injury (ischemia-reperfusion or I-R) which associates with excessive calcium concentration within cardiomyocytes, a process termed calcium overload. Our laboratory recently showed that the transient receptor potential vanilloid, member 4 (TRPV4) cation channel is highly expressed within cardiomyocytes of the aged (but not young) heart and contributes to intracellular calcium overload following hypoosmotic stress. Osmotic changes in cardiac tissue are pronounced during I-R, and reperfusion associates with substantial hypoosmotic stress on cardiomyocytes. Therefore, this proposal tests the central hypothesis that TRPV4 contributes to calcium overload and cardiac dysfunction following I-R. To test this hypothesis we utilize young (3-6 months, with low TRPV4 expression) and aged (24-26 months, with high TRPV4 expression) C576BL/6 mice, young cardiomyocyte-specific TRPV4 transgenic mice (“overexpressors” with high TRPV4 expression), and aged TRPV4 knock-out mice (with no TRPV4 expression). To complement this genetic approach we also utilize a pharmacological approach with specific antagonists of TRPV4. Specific Aim 1 tests the hypothesis that TRPV4 produces cardiomyocyte calcium overload, excessive contractility, and arrhythmia following I-R. Isolated cardiomyocytes subjected to simulated I-R (combined metabolic and osmotic stress) will be used to examine the effect of TRPV4 on calcium signaling modalities using high-resolution confocal fluorescence microscopy. Similarly, transgenic mouse hearts which encode the GCaMP6f calcium sensor will be subjected to ex vivo I-R with pro-arrhythmic calcium signaling monitored within the intact organ. Langendorff-perfused hearts will be utilized to test the role of TRPV4 on contractility and arrhythmia following I- R. Specific Aim 2 tests the hypothesis that TRPV4 contributes to cardiomyocyte death, tissue damage, and adverse cardiac remodeling following I-R. Fluorescence imaging approaches of mitochondrial membrane potential and plasma membrane integrity will elucidate the role of TRPV4 on cardiomyocyte dysfunction and death in real time following simulated I-R or in ex vivo perfused hearts following I-R. Translational studies using orally bioavailable TRPV4 inhibitors will be used to examine the role of TRPV4 in adverse cardiac remodeling and tissue damage following I-R in vivo. The goal of this project is to rigorously examine the role of the TRPV4 ion channel in cardiac dysfunction and tissue damage following I-R, with the long-term goal of translating our research findings into new treatments for aged individuals following MI.

项目总结/摘要 年龄增长是冠状动脉疾病和心肌梗死（MI）的主要危险因素。一个主要原因 心肌梗死后心律失常和组织损伤的主要原因是再灌注损伤（缺血-再灌注或I-R）， 心肌细胞内钙浓度过高，这一过程称为钙超载。本实验室 最近表明，瞬时受体电位香草酸，成员4（TRPV 4）阳离子通道是高度 在老年（而非年轻）心脏的心肌细胞内表达，并有助于细胞内钙 低渗应激后超负荷。在I-R期间心脏组织中的渗透变化是明显的， 再灌注与对心肌细胞的实质性低渗应激有关。因此，这一提议考验了 核心假设TRPV 4导致I-R后钙超载和心功能障碍。测试 该假设我们利用年轻（3-6个月，具有低TRPV 4表达）和老年（24-26个月，具有高TRPV 4表达）。 TRPV 4表达）C576 BL/6小鼠，年轻心肌细胞特异性TRPV 4转基因小鼠（“过表达者”） 具有高TRPV 4表达）和老年TRPV 4敲除小鼠（没有TRPV 4表达）。以补充 这种遗传方法我们还利用了TRPV 4特异性拮抗剂的药理学方法。具体 目的1检验TRPV 4引起心肌细胞钙超载、过度收缩和心肌细胞凋亡的假设。 I-R后心律失常分离的心肌细胞进行模拟I-R（结合代谢和渗透压 应激）将被用于使用高分辨率成像技术检查TRPV 4对钙信号传导模式的影响。 共聚焦荧光显微镜。类似地，编码GCaMP 6 f钙的转基因小鼠心脏， 传感器将经受离体I-R，其中在完整器官内监测促钙离子信号传导。 Langendorff灌注的心脏将用于测试TRPV 4对I-2000后收缩性和心律失常的作用。 R.特异性目的2检验了TRPV 4导致心肌细胞死亡、组织损伤和 I-R后的不良心脏重塑。线粒体膜的荧光成像方法 潜在的和质膜完整性将阐明TRPV 4对心肌细胞功能障碍的作用， 模拟I-R后真实的死亡或I-R后离体灌注心脏中的死亡。翻译研究使用 口服生物可利用的TRPV 4抑制剂将用于检查TRPV 4在不利的心脏重塑中的作用 和体内I-R后的组织损伤。该项目的目标是严格检查TRPV 4的作用 离子通道在I-R后心功能障碍和组织损伤中的作用，长期目标是将我们的研究成果转化为 研究发现为老年人MI后的新疗法。