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主要死亡原因。减少缺血性中风后的脑损伤和促进修复的途径有限。肾素-血管紧张素系统(RAS)中的血管紧张素转换酶(ACE)、血管紧张素转换酶II(Ang II)和血管紧张素转换酶AT1受体(Ang AT1受体)参与高血压和卒中等多种心血管疾病的病理生理过程。随着血管紧张素转换酶2(ACE2)、血管紧张素转换酶(1-7)和Mas受体的发现，越来越多的证据表明ACE2/Ang(1-7)/Mas轴在控制血压和血流方面与ACE/Ang II/AT1轴相互抵消。然而，这些反作用轴在缺血性中风中的作用在很大程度上是未知的。活性氧簇(ROS)已被认为是导致脑缺血损伤的主要因素。内皮型一氧化氮合酶(ENOS)在维持正常脑血流中起重要作用。血管紧张素转换酶(ACE)/血管紧张素Ⅱ(Ang II)/血管紧张素Ⅱ(AT1)轴的激活可诱导NADPH氧化酶(NOX)来源的ROS产生过量和eNOS来源的NO减少。最近的研究表明，Ang(1-7)/mAs对Ang II/AT1激活的人内皮细胞NADPH氧化酶具有负性调节作用。有证据表明，ACE/Ang II/AT1轴的激活通过血流依赖和独立的机制加重了缺血性卒中。骨髓来源的内皮祖细胞和缺血诱导的缺氧诱导因子-1(HIF-1)、基质衍生因子-1a(SDF-1a)及其受体CXCR4的上调参与了缺血性卒中后的修复过程(血管生成和神经生成)。综上所述，我们推测ACE2/Ang(1-7)/Mas轴通过调节NOx/ROS、eNOS/NO和HIF-1/SDF-1a/CXCR4信号通路，从而在脑缺血损伤和修复中发挥重要作用，从而抵消ACE/Ang II/AT1对神经、脑血管和内皮祖细胞的调控作用。为了验证这一假设，三个转基因小鼠模型，R+A+(肾素和血管紧张素原过度表达)，ACE2+(脑内过表达ACE2)和R+A+ACE2+(R+A+ACE2+在R+A+小鼠脑内过表达)，以及一个过表达ACE2的慢病毒载体(Lenti-ACE2)将用于通过多种综合的生理、药理和分子方法来实现四个特定目的：特异性目标1将通过调节血压和脑血流来检测ACE2/Ang(1-7)/mas在对抗ACE/Ang II/AT1中的作用。具体目标2将通过独立于血压和脑血流的机制来研究ACE2/Ang(1-7)/mAs在对抗ACE/Ang II/AT1在缺血性损伤中的作用。具体目标3将研究ACE2/Ang(1-7)/mas在对抗ACE/Ang II/AT1在缺血性卒中后脑修复中的作用。具体目标4将研究ACE2/Ang(1-7)/mas在抑制ACE/Ang II/AT1中的作用，以促进EPC移植促进缺血性卒中后的修复。 公共卫生相关性：在美国，缺血性中风是第三大死亡原因。目前，还没有有效的药物来减轻缺血性卒中后的脑损伤，促进修复。肾素-血管紧张素系统(RAS)是治疗缺血性卒中的一个可能靶点。RAS包括血管紧张素转换酶(ACE)、血管紧张素转换酶II和血管紧张素转换酶AT1受体(ACE/Ang II/AT1轴)和血管紧张素转换酶2(ACE2)、Ang(1-7)和Mas受体(ACE2/Ang(1-7)/Mas受体轴)。这将验证ACE2/Ang(1-7)/mas轴通过对抗ACE/Ang II/AT1在急性脑缺血损伤和修复中发挥重要作用的假说。本项目的研究结果将对Ang II/Ang(1-7)平衡在缺血性卒中的病理生理机制中的作用提供新的见解，从而促进缺血性卒中新的治疗靶点的开发。

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.