Balance of Angiotensin II/Angiotensin (1-1): A Target in Ischemic Stroke

描述（由申请人提供）：缺血性中风是美国第三大死亡原因。减少缺血性脑卒中后脑损伤和促进脑修复的途径有限。肾素-血管紧张素系统（RAS）的血管紧张素（Ang）转换酶（ACE）、Ang II和Ang AT1受体参与高血压、脑卒中等多种心血管疾病的病理生理过程。随着Ang转换酶2 （ACE2）、Ang（1-7）和Mas受体的发现，越来越多的证据表明，ACE2/Ang (1-7)/Mas轴抵消ACE/Ang II/AT1轴对血压和血流的控制。然而，这些抵消轴在缺血性中风中的作用在很大程度上是未知的。活性氧（Reactive oxygen species， ROS）是引起缺血性损伤的一个重要因素。内皮一氧化氮合酶（eNOS）产生的一氧化氮（NO）在维持正常脑血流中起重要作用。ACE/Ang II/AT1轴的激活已被证明可诱导NADPH氧化酶（NOX）衍生ROS的过量产生和eNOS衍生NO的减少。最近的一项研究表明，Ang (1-7)/Mas负向调节人内皮细胞中Ang II/AT1活化的NADPH氧化酶。有证据表明，ACE/Ang II/AT1轴的激活可通过血流依赖和独立机制恶化缺血性卒中。骨髓源性内皮祖细胞（EPCs）和缺血诱导的缺氧诱导因子-1 （HIF-1）、基质源性因子-1a （SDF-1a）及其受体CXCR4的上调参与缺血性卒中后的修复过程（血管生成和神经发生）。综上所述，我们假设ACE2/Ang (1-7)/Mas轴通过调节NOX/ROS、eNOS/NO和HIF-1/SDF-1a/CXCR4信号通路，抵消ACE/Ang II/AT1调控神经、脑血管和EPCs功能，从而在脑缺血损伤和修复中发挥重要作用。为了验证这一假设，将使用转基因小鼠模型，R+A+（肾素和血管紧张素原过表达），ACE2+ （ACE2在脑内过表达）和R+A+ACE2+ （ACE2在R+A+脑内过表达）小鼠，以及过表达ACE2的慢病毒载体（lentii -ACE2），通过多种综合生理，药理学和分子方法来追求四个特定目标：特异性Aim 1将测试ACE2/Ang (1-7)/Mas通过调节血压和脑血流来对抗ACE/Ang II/AT1在缺血性损伤中的作用。特异性目的2将通过独立于血压和脑血流的机制研究ACE2/Ang (1-7)/Mas在缺血性损伤中抵消ACE/Ang II/AT1的作用。特异性目的3将研究ACE2/Ang (1-7)/Mas在缺血性脑卒中后脑修复中对抗ACE/Ang II/AT1的作用。特异性目的4将探讨ACE2/Ang (1-7)/Mas在ACE/Ang II/AT1的拮抗中对EPCs移植促进缺血性脑卒中后修复的作用。

DESCRIPTION (provided by applicant): Ischemic stroke is the third leading cause of death in the United States. There are limited avenues for reducing cerebral damage and promoting repair after ischemic stroke. The angiotensin (Ang) converting enzyme (ACE), Ang II and Ang AT1 receptor of the renin-angiotensin system (RAS) participate in the pathophysiology of various cardiovascular diseases including hypertension and stroke. With the discovery of the Ang converting enzyme 2 (ACE2), Ang (1-7) and Mas receptor, accumulating evidence suggest that the ACE2/Ang (1-7)/Mas axis counteracts the ACE/Ang II/AT1 axis in control of blood pressure and flow. However, the role of these counteracting axes in ischemic stroke is largely unknown. Reactive oxygen species (ROS) have been recognized as a major factor contributing to ischemic damage. Nitric oxide (NO) from endothelium nitric oxide synthase (eNOS) is important in maintaining normal cerebral blood flow. Activation of ACE/Ang II/AT1 axis has been shown to induce overproduction of NADPH oxidase (NOX) derived ROS and reduction of eNOS derived NO. A recent report shows that Ang (1-7)/Mas negatively modulates Ang II/AT1 activated NADPH oxidase in human endothelial cells. Evidence suggests that activation of ACE/Ang II/AT1 axis worsens ischemic stroke via both blood flow dependent and independent mechanisms. The bone marrow (BM) derived endothelial progenitor cells (EPCs) and ischemia-induced up-regulation of hypoxia induced factor-1 (HIF-1), stromal derived factor-1a (SDF-1a) and its receptor CXCR4 participate in the repair processes (angiogenesis and neurogenesis) after ischemic stroke. Taken together, we hypothesize that the ACE2/Ang (1-7)/Mas axis counteracts ACE/Ang II/AT1 in control of neural, cerebrovascular and EPCs function through modulating NOX/ROS, eNOS/NO and HIF-1/SDF-1a/CXCR4 signaling pathways, thereby playing an important role in cerebral ischemic damage and repair. To test this hypothesis, there transgenic mice models, R+A+ (renin and angiotensinogen over-expression), ACE2+ (ACE2 over-expressed in the brain) and R+A+ACE2+ (ACE2 over-expressed in the brain of R+A+) mice, and a lentiviral vector over-expressing ACE2 (lenti-ACE2) will be used for pursuing four specific aims using a variety of integrated physiological, pharmacological and molecular approaches: Specific Aim 1 will test the role of ACE2/Ang (1-7)/Mas in counteracting ACE/Ang II/AT1 in ischemic damage through regulating blood pressure and cerebral blood flow. Specific Aim 2 will examine the role of ACE2/Ang (1-7)/Mas in counteracting ACE/Ang II/AT1 in ischemic damage through mechanisms independent of blood pressure and cerebral blood flow. Specific Aim 3 will study the role of ACE2/Ang (1-7)/Mas in counteracting ACE/Ang II/AT1 in cerebral repair after ischemic stroke. Specific Aim 4 will investigate the role of ACE2/Ang (1-7)/Mas in counteracting ACE/Ang II/AT1 in the efficacy of EPCs transplantation for promoting repair after ischemic stroke. PUBLIC HEALTH RELEVANCE: Ischemic stroke is the third leading cause of death in the United States. Currently, there are no effective drugs for reducing cerebral damage and promoting repair after ischemic stroke. The renin-angiotensin system (RAS) is a possible therapeutic target for ischemic stroke. There are two arms of RAS, the angiotensin (Ang) converting enzyme (ACE), Ang II and Ang AT1 receptor (ACE/Ang II/AT1 axis) and the Ang converting enzyme 2 (ACE2), Ang (1-7) and Mas receptor (ACE2/Ang (1-7)/Mas receptor axis). This proposal will test the hypothesis that the ACE2/Ang (1- 7)/Mas axis via counteracting the ACE/Ang II/AT1 plays important roles in acute cerebral ischemic damage and repair. The results of this project will provide new insights on the role of Ang II/Ang (1-7) balance in the pathophysiology of ischemic stroke, and thus facilitate the development of novel therapeutic targets for ischemic stroke.