Nox 4 in Endothelial Cell ROS Production, Signaling and Motility

Nox 4 在内皮细胞 ROS 产生、信号传导和运动中的作用

• 批准号: 7136887
• 负责人: VISWANATHAN NATARAJAN
• 金额: $ 39.32万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-04 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7136887
• 关键词: NAD(P)H dehydrogenase; biological signal transduction; cell migration; disease /disorder model; enzyme activity; enzyme induction /repression; free radical oxygen; gene deletion mutation; genetic regulation; genetically modified animals; human tissue; hyperoxia; laboratory mouse; lung injury; mitogen activated protein kinase; phospholipase D; tumor necrosis factor alpha; vascular endothelium; western blottings

DESCRIPTION (provided by applicant): Increased generation of reactive oxygen species (ROS) in the vasculature has been linked to inflammation, vascular leakiness, atherosclerosis and other cardiovascular disorders. In the vascular endothelium, NADPH oxidase has been identified as a major source of ROS, which regulates signaling pathways involved in endothelial cell growth, migration, cytoskeletal organization and barrier function. The endothelial NADPH oxidase, similar to phagocytic oxidase, consists of cytosolic (p47phox, p67phox and Rac1) and membranebound (gp91phox and p22phox) components. Recently, several homologues of Nox 2 (gp91phox) namely Nox1, Nox3, Nox4 and Nox5 have been identified. We and others have found that Nox4 mRNA is expressed at much higher levels, compared to Nox2, in the endothelial cells from different vascular beds; however, the role of Nox4 in ROS production and endothelial cell functions is not well understood. In preliminary experiments, we have identified that Nox4 is not only involved in hyperoxia- and TNF-alpha-induced superoxide/ROS production but also in endothelial cell signal transduction regulating motility and capillary tube formation. The overall hypothesis of this proposal is that "Nox4 plays a key role in regulating endothelial signaling pathways involved in superoxide/ROS production, migration and capillary tube formation". This hypothesis will be tested in primary human lung endothelial cells and in genetically modified Nox4 and Nox2 (gp91phox) mice using hyperoxia- and TNF-alpha as modulators of NADPH oxidase. Specific Aim 1: To characterize expression of Nox 4 in HPAECs and mouse lung ECs and determine its role in ROS production; Specific Aim 2: To investigate molecular mechanisms of increased expression and activation of Nox4 by hyperoxia in human lung endothelial cells ; Specific Aim 3: To characterize signaling pathways that regulate Nox4 dependent endothelial cell migration and capillary tube formation in response to hyperoxia; Specific Aim 4: To investigate the role of Nox4 in ROS generation and vascular leakiness in an in vivo murine model of lung injury. These experiments will provide novel and new insights into the role of Nox4, as a component of NADPH oxidase, in regulating endothelial cell ROS production and function that may lead to development of therapeutic strategies minimizing lung injury.

描述（由申请人提供）：血管系统中活性氧（ROS）生成增加与炎症、血管渗漏、动脉粥样硬化和其他心血管疾病有关。在血管内皮中，NADPH氧化酶已被确定为ROS的主要来源，其调节参与内皮细胞生长、迁移、细胞骨架组织和屏障功能的信号通路。内皮细胞NADPH氧化酶与吞噬细胞氧化酶相似，由胞质（p47 phox、p67 phox和Rac 1）和膜结合（gp 91 phox和p22 phox）组分组成。最近，Nox 2（gp 91 phox）的几个同源物，即Nox 1、Nox 3、Nox 4和Nox 5已被鉴定。我们和其他人已经发现，与Nox 2相比，Nox 4 mRNA在来自不同血管床的内皮细胞中以高得多的水平表达;然而，Nox 4在ROS产生和内皮细胞功能中的作用尚不清楚。在初步的实验中，我们已经确定，Nox 4不仅参与高氧和TNF-α诱导的超氧化物/ROS的生产，但也在内皮细胞的信号转导调节运动和毛细血管的形成。该提议的总体假设是“Nox 4在调节涉及超氧化物/ROS产生、迁移和毛细管形成的内皮信号传导途径中起关键作用”。这一假设将在原代人肺内皮细胞和基因修饰的Nox 4和Nox 2（gp 91 phox）小鼠中使用高氧和TNF-α作为NADPH氧化酶的调节剂进行测试。具体目标1：表征HPAEC和小鼠肺EC中Nox 4的表达并确定其在ROS产生中的作用;具体目标2：研究人肺内皮细胞中高氧引起的Nox 4表达和活化增加的分子机制;具体目标3：表征响应于高氧调节Nox 4依赖性内皮细胞迁移和毛细血管形成的信号通路;具体目标4：研究Nox 4在体内小鼠肺损伤模型中ROS产生和血管渗漏中的作用。这些实验将为Nox 4作为NADPH氧化酶的一种组分在调节内皮细胞ROS产生和功能中的作用提供新颖和新的见解，这可能导致开发最小化肺损伤的治疗策略。