Protection with Adenosine A1 Receptor Overexpression

腺苷 A1 受体过度表达的保护

• 批准号: 6457331
• 负责人: David G Van Wylen
• 金额: $ 13.02万
• 依托单位: ST. OLAF COLLEGE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2006-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6457331
• 关键词: adenosine; cell line; cytoprotection; fluorescence microscopy; membrane potentials; mitochondrial membrane; mitogen activated protein kinase; myocardial ischemia /hypoxia; potassium channel; protein kinase C; protein localization; protein tyrosine kinase; purine /pyrimidine metabolism; purinergic receptor; receptor expression; transfection

DESCRIPTION (provided by applicant): Minimizing myocardial damage resulting from ischemic heart disease remains a major challenge for health professionals. Adenosine-mediated cardioprotection has recently been harnessed through a transgenic mouse model of cardiac specific overexpression of the adenosine A1 receptor that demonstrates remarkable protection against ischernia-reperfusion injury. However, the mechanisms of this cardioprotection remain largely unknown. The broad, long-term scientific objective of the proposed research is to use an isolated cardiac myocyte model of adenosine A1 receptor overexpression to investigate the mechanisms of cardioprotection associated with adenosine A1 receptor overexpression. The specific aims of the proposed research are: Specific Aim 1: To characterize the cardioprotection against simulated ischemia associated with adenosine A1 receptor overexpression in H9c2 cells. Using standard transfection techniques to induce adenosine A1 receptor overexpression, H9c2 cells (a rat embryonic heart cell line) will be used to test the hypothesis that adenosine A1 receptor overexpression protects against simulated ischemia. Specific Aim 2: To determine the role of protein kinase C (PKC), tyrosine kinase, p38 mitogen-activated protein kinase (MAPK), and mitochondrial KATP channels in the protection associated with adenosine A1 receptor overexpression in H9c2 cells. PKC, tyrosine kinase, p38 MAPK, and mitochondrial KATP channels contribute to adenosme-mediated cardioprotection; however, their involvement in the protection afforded by adenosine A1 receptor overexpression is largely unknown. These pathways will be inhibited pharmacologically to determine their role in the protection afforded by adenosine A1 receptor overexpression. Specific Aim 3: To determine the effect of adenosine A1 receptor overexpression on the timing of PKC translocation, p38 MAPK phosphorylation, and depolarization of the mitochondrial membrane potential during simulated ischemia. Fluorescence microscopy will be used to observe the effect of adenosine A1 receptor overexpression on the subcellular location of PKC, the dynamics of p38 MAPK phosphorylation, and the time course of mitochondrial depolarization during simulated ischemia. The proposed research, which will introduce undergraduates to the important interface between molecular biology, cardiac physiology, and clinical application, will allow more informed decisions regarding the potential use of adenosine receptor manipulation in humans via gene therapy.

描述（由申请人提供）：最大限度地减少造成的心肌损伤 缺血性心脏病仍然是卫生专业人员面临的重大挑战。 腺苷介导的心脏保护作用最近已被利用 腺苷 A1 心脏特异性过表达的转基因小鼠模型 对缺血再灌注表现出显着保护作用的受体 受伤。然而，这种心脏保护机制在很大程度上仍然存在 未知。拟议研究的广泛、长期的科学目标是 使用腺苷A1受体的离体心肌细胞模型 过表达以研究心脏保护相关的机制 与腺苷 A1 受体过度表达。拟议的具体目标 研究有： 具体目标 1：表征针对模拟缺血的心脏保护作用 与 H9c2 细胞中腺苷 A1 受体过度表达有关。使用 诱导腺苷 A1 受体的标准转染技术 过度表达，H9c2 细胞（大鼠胚胎心脏细胞系）将用于 检验腺苷 A1 受体过度表达可预防的假设 模拟缺血。 具体目标 2：确定蛋白激酶 C (PKC)、酪氨酸的作用 激酶、p38 丝裂原激活蛋白激酶 (MAPK) 和线粒体 KATP 与腺苷 A1 受体过度表达相关的保护通道 在H9c2细胞中。 PKC、酪氨酸激酶、p38 MAPK 和线粒体 KATP 通道 有助于腺苷介导的心脏保护作用；然而，他们的参与 腺苷 A1 受体过度表达所提供的保护在很大程度上是 未知。这些途径将被药理学抑制以确定其 腺苷 A1 受体过度表达所提供的保护作用。 具体目标 3：确定腺苷 A1 受体过度表达的影响 PKC 易位、p38 MAPK 磷酸化的时间以及 模拟过程中线粒体膜电位的去极化 缺血。荧光显微镜将用于观察效果 腺苷 A1 受体在 PKC 亚细胞位置过度表达， p38 MAPK 磷酸化的动态以及线粒体的时间过程 模拟缺血期间的去极化。 拟议的研究将向本科生介绍重要的 分子生物学、心脏生理学和临床之间的接口 应用程序，将允许就潜在用途做出更明智的决定 通过基因疗法操纵人类的腺苷受体。