Protection with Adenosine A1 Receptor Overexpression

描述(由申请人提供)：将心肌损伤降至最低 预防缺血性心脏病仍然是卫生专业人员面临的一大挑战。 腺苷介导的心脏保护最近通过一种 心脏特异性腺苷A1过表达转基因小鼠模型的建立 对缺血再灌流有显著保护作用的受体 受伤。然而，这种心脏保护的机制在很大程度上仍然存在。 未知。拟议研究的广泛的、长期的科学目标是 应用腺苷A1受体分离的心肌细胞模型 过表达研究心脏保护相关机制 腺苷A1受体过度表达。建议的具体目标 研究内容包括： 具体目标1：表征对模拟缺血的心脏保护作用 与腺苷A1受体在H9c2细胞中的过度表达有关。vbl.使用 诱导腺苷A1受体的标准转染法 H9c2细胞(一种大鼠胚胎心脏细胞系)将被用于 测试腺苷A1受体过度表达对预防疾病的假设 模拟缺血。 特定目的2：确定蛋白激酶C(PKC)、酪氨酸的作用 P38丝裂原活化蛋白激酶(MAPK)和线粒体KATP 与腺苷A1受体过度表达相关的保护作用的通道 在H9c2细胞中。PKC、酪氨酸激酶、p38MAPK和线粒体KATP通道 有助于腺苷酶介导的心脏保护；然而，它们参与了 腺苷A1受体过度表达所提供的保护在很大程度上是 未知。这些通路将被药物抑制以确定它们的 在腺苷A1受体过度表达所提供的保护中的作用。 特异性目标3：确定腺苷A1受体过表达的影响 关于PKC转位、p38MAPK磷酸化的时间，以及 模拟过程中线粒体膜电位的去极化 缺血症。将用荧光显微镜观察其效果。 腺苷A1受体在PKC亚细胞定位上的过度表达 P38MAPK的磷酸化动力学和线粒体的时间进程 模拟缺血过程中的去极化。 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，将允许关于潜在使用的更明智的决定 通过基因治疗在人类中操纵腺苷受体。

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.