Connexin Distribution in Physiological Versus Pathological Cardiac Hypertrophy

生理性与病理性心脏肥大中的连接蛋白分布

• 批准号: 8195558
• 负责人: GEORGE COOPER
• 金额: --
• 依托单位: RALPH H JOHNSON VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-10-01 至 2013-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8195558
• 关键词: Accounting; Address; Adenoviruses; Adrenergic Agents; Adrenergic Agonists; Adrenergic Receptor; Adult; Affinity; Agonist; Aliquot; Animal Model; Animals; Applications Grants; Arrestins; Arrhythmia; Atrial Heart Septal Defects; Attenuated; Basic Science; Binding; Biopsy; Cadherins; Canis familiaris; Cardiac; Cardiac Myocytes; Caring; Catheterization; Cell Adhesion; Cell Nucleolus; Cell Nucleus; Cell surface; Cells; Characteristics; Chronic; Clinical; Complex; Confocal Microscopy; Congenital Heart Defects; Connexin 43; Connexins; Connexon; Control Animal; Cyclic AMP; Cytoplasm; Cytoskeleton; Data; Dependence; Diffuse; Diffusion; Distal; Docking; Down-Regulation; Dynein ATPase; Echocardiography; Electrocardiogram; Electrons; Elements; Environment; Event; Exhibits; Failure; Family; Family Felidae; Felis catus; Functional disorder; Funding; Gap Junctions; Genes; Genetic Transcription; Genetic screening method; Goals; Golgi Apparatus; Grant; Grant Review; Growth; Heart; Heart Diseases; Heart Hypertrophy; Heart failure; Homeostasis; Human; Hypertrophy; Immunoblotting; Infection; Interphase Cell; Intervention; Intracellular Transport; Kinesin; Lead; Left; Left ventricular structure; Life; Life Cycle Stages; Location; Lung; Lysosomes; MAP4; Maps; Measurement; Measures; Mechanics; Mediating; Medical center; Membrane; Membrane Transport Proteins; Messenger RNA; Metalcaptase; Methods; Microfilaments; Microtubule Depolymerization; Microtubule Proteins; Microtubule-Associated Proteins; Microtubules; Minor; Modeling; Molecular; Morphology; Motion; Motor; Mus; Myocardial; Myocardium; Myofibrils; Nocodazole; Normal Cell; Normal Range; Optics; Pathway interactions; Patients; Pharmaceutical Preparations; Phase; Phenotype; Phosphoric Monoester Hydrolases; Phosphotransferases; Physiological; Plus End of the Microtubule; Positioning Attribute; Prevention; Principal Investigator; Process; Program Reviews; Progress Reports; Property; Propranolol; Protein Biosynthesis; Protein Dephosphorylation; Protein Isoforms; Protein phosphatase; Proteins; Protocols documentation; Pulmonary artery structure; Recruitment Activity; Research; Research Support; Ribonucleoproteins; Ribosomal RNA; Ribosomes; Right ventricular structure; Right-On; Role; Sarcolemma; Science; Series; Site; Specificity; Staining method; Stains; Stimulus; Stress; Structure; System; Systolic Pressure; Telemetry; Testing; Therapeutic Intervention; Thinking; Time; Tissues; Transfection; Transgenic Mice; Translating; Translational Research; Translations; Transport Process; Transport Vesicles; Treatment Efficacy; Tubulin; Up-Regulation; Ventricular; Vesicle; Work; adrenergic; base; beta-adrenergic receptor; constriction; density; extracellular; gap junction channel; gene therapy; hemodynamics; improved; interest; macromolecule; member; messenger ribonucleoprotein; overexpression; p21-activated kinase 1; palliative; particle; pressure; prevent; programs; public health relevance; receptor; receptor internalization; receptor recycling; research study; response; small molecule; success; trans-Golgi Network

Research supported by this grant during the previous twenty-four years has been built around extensive data showing that cardiac structure, composition, and function each respond rapidly and reversibly to changes in hemodynamic load. The first set of studies supported by this grant used isolated cells, or cardiocytes, and intact animals to demonstrate the role of load as a central regulator of cardiocyte growth. The second set of studies supported by this grant, which also used load change as the primary experimental variable, led to our discovery of a dense cardiocyte microtubule network during severe pressure-overload cardiac hypertrophy that contrib- utes to the contractile dysfunction which occurs in this setting. The initial goals for the subsequent studies of this abnormal microtubule network were to determine how it contributes to the contractile dysfunction of hypertrophied myocardium. Major findings have been that 1) it is based both on increased tubulin, and thus microtubules, and on greater microtubule stability, 2) the major car- diac microtubule-stabilizing microtubule-associated protein, MAP4, is greatly upregulated in pressure overload hypertrophy and binds extensively to microtubules, and 3) contractile dysfunction is caused by viscous loading imposed on shortening myofilaments by the dense microtubule network. However, the most important normal role of the microtubules in an interphase cell such as the cardiocyte is not to determine cellular rheological properties but rather to subserve intracellular transport of macromolecules and vesicles via the microtubule-associated kinesin and dynein families of motor proteins. Indeed, this is an absolutely essential role in the extremely diffusion-restricted cytoplasm of the adult cardiocyte. For this reason, and because of the known inhibition of microtubule-dependent intracellular transport by excessive decoration of microtubules with MAPs, we next asked if microtubule-based transport of the activated ¿-adrenergic receptor and/or mRNA - ribonucleoprotein complexes was inhibited by MAP4 binding to microtubules in pressure- overload hypertrophy. Such, in fact. was the case. Building on this most recent work, we propose to examine here the potential role of alterations in microtubule network organization and MAP4 binding in causing abnormal transport and localization of connexin43 [Cx43], a gap junction protein known to undergo functionally important alterations in quantity and localization during pathological cardiac hypertrophy. The basic research in the first objective will use isolated cells as well as oper- ated and transgenic mice to determine whether MAP4 decoration of microtubules, and the attendant densifica- tion of the microtubule network, inhibit the normal transport of Cx43 to gap junctions as well as Cx43-depen- dent electrophysiological function. The translational research in the second and third objectives will compare an equal degree & duration of pathological pressure vs. physiological volume overload hypertrophy. We will first extend the findings of the first objective to ask if MAP4 decoration of the dense microtubule network in pathological hypertrophy has a role in the altered Cx43 transport and localization that are important clinically in forming an arrhythmogenic substrate. We will then ask if ¿-receptor blockade in pathological hypertrophy, which early data indicates will prevent the abnormal microtubule phenotype, will also prevent the abnormal Cx43 phenotype in this setting. In the first objective we will use murine models, and in the second and third objectives we will use our long- standing feline models of physiological versus pathological hypertrophy. While we recognize that it is prefer- able to use a single species, in this research the initial mechanistic portion can only be done in the mouse, but the later quantitative translational portions require very reproducible animal models that can be reliably and verifiably ¿-blocked and have an equivalent degree and duration of physiological vs. pathological hypertrophy, with ex- tensively characterized cytoskeletal properties in each setting.

在过去的24年里，这项资助所支持的研究是建立在广泛的数据基础上的。 表明心脏结构、组成和功能对心脏的变化都有快速和可逆的反应。 血流动力学负荷这项资助支持的第一组研究使用了分离的细胞或心肌细胞， 动物来证明负荷作为心脏细胞生长的中心调节器的作用。第二组研究 在该基金的支持下，也使用负载变化作为主要的实验变量，导致了我们的发现 在严重的压力超负荷性心脏肥大过程中，致密的心肌细胞微管网络， 导致这种情况下发生的收缩功能障碍。 对这种异常微管网络的后续研究的最初目标是确定它是如何被激活的。 导致肥厚心肌的收缩功能障碍。主要的发现是：（1） 基于增加的微管蛋白和微管，以及更大的微管稳定性，2）主要的汽车， 压力超负荷时，双腔管稳定微管相关蛋白MAP 4大幅上调 肥大并广泛结合微管，和3）收缩功能障碍是由粘性负荷引起的 通过致密的微管网络施加于缩短的肌丝。 然而，微管在间期细胞如心肌细胞中最重要的正常作用是 不是为了确定细胞的流变学性质，而是为了促进大分子的细胞内转运 和囊泡通过微管相关的驱动蛋白和动力蛋白家族的马达蛋白。的确，这是一个 在成人心肌细胞的极度扩散受限的细胞质中发挥绝对重要的作用。基于这个理由， 并且由于已知过度修饰会抑制微管依赖性细胞内转运 微管与地图，我们接下来问，如果微管为基础的运输激活的肾上腺素能受体 和/或mRNA -核糖核蛋白复合物被MAP 4与微管的结合所抑制， 超负荷肥大事实上，是这样的。就是这样。 基于这一最新的工作，我们建议在这里研究微管改变的潜在作用， 网络组织和MAP 4结合导致连接蛋白43 [Cx43]的异常运输和定位， 已知间隙连接蛋白质在生长过程中在数量和定位上发生功能上重要的改变， 病理性心脏肥大第一个目标的基础研究将使用分离的细胞以及操作， 用小鼠和转基因小鼠来确定微管的MAP 4修饰，以及随之而来的致密化， 微管网络的作用，抑制Cx43向缝隙连接的正常转运以及Cx43依赖性。 削弱电生理功能第二个和第三个目标中的翻译研究将进行比较 病理性压力与生理性容量超负荷肥大的程度和持续时间相等。我们将 首先扩展第一个目标的研究结果，询问是否MAP 4修饰致密的微管网络， 病理性肥大在Cx43转运和定位的改变中起作用，这在临床上是重要的。 形成致炎底物。然后，我们会问，如果在病理性肥大， 早期资料表明会防止异常微管表型，也会防止异常Cx43 表型在此设置。 在第一个目标中，我们将使用鼠模型，在第二个和第三个目标中，我们将使用我们的长期- 生理与病理性肥大的猫模型。尽管我们承认这是更好的- 由于能够使用单一物种，在这项研究中，最初的机制部分只能在小鼠中完成，但 随后的定量翻译部分需要非常可再现的动物模型 阻断，具有相同程度和持续时间的生理性与病理性肥大， 紧张的特点细胞骨架的性质，在每一个设置。