Regulation of the Nox1 NADPH Oxidase in Vascular Smooth Muscle Cells

血管平滑肌细胞中 Nox1 NADPH 氧化酶的调节

• 批准号: 8698326
• 负责人: FRANCIS J MILLER
• 金额: --
• 依托单位: IOWA CITY VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8698326
• 关键词: Address; Adverse effects; Affect; Amino Acids; Animals; Arterial Fatty Streak; Arteries; Atherosclerosis; Blood Vessels; Cardiovascular Diseases; Cardiovascular system; Catalytic Domain; Cell Migration Pathway; Cell membrane; Cells; Clinical; Complement Factor B; Complex; Cytokine Activation; Data; Development; Effectiveness; Endocytosis; Event; Family; Generations; Goals; Growth; Healthcare; Homologous Gene; Human; Hyperplasia; Immunologic Deficiency Syndromes; Inflammatory; Injury; Link; Mediating; Membrane Protein Traffic; Morbidity - disease rate; Mutate; NADPH Oxidase; Nuclear; Oxidation-Reduction; Pathogenesis; Pathology; Pathway interactions; Phosphorylation; Phosphorylation Site; Positioning Attribute; Prevention; Production; Protein Isoforms; Proteins; Reactive Oxygen Species; Regulation; Research; Role; Signal Pathway; Signal Transduction; Site-Directed Mutagenesis; Smooth Muscle Myocytes; Source; Specificity; Systems Biology; TNF gene; Testing; Therapeutic; United States; Vascular Diseases; Work; antioxidant therapy; burden of illness; care burden; cell growth; cell motility; clinical application; cytokine; in vivo Model; inhibitor/antagonist; insight; meetings; migration; mortality; mutant; neointima formation; new therapeutic target; novel; novel therapeutic intervention; novel therapeutics; prevent; programs; response; response to injury; therapeutic target; trafficking

DESCRIPTION (provided by applicant): Cardiovascular disease is characterized by increased generation of reactive oxygen species (ROS) in the vessel wall, which results in activation of signaling pathways that ultimately promote cell growth and neointimal formation. ROS derived from smooth muscle cells (SMCs) is a major contributing factor in the development of vascular disease, though antioxidant therapies have achieved only limited therapeutic benefit. Therefore, it is necessary to identify targeted approaches to prevent ROS generation. NADPH oxidases are the predominant source of ROS in the vasculature, with Nox1 being the primary catalytic NADPH oxidase expressed in SMCs. Nox1-derived ROS have been linked to atherosclerosis as well as neointimal formation and SMC migration following injury, but the precise mechanisms by which Nox1 activates redox- dependent signaling pathways remain incompletely defined. In order to develop targeted therapeutics against Nox1, it is first necessary to understand the parameters that must be met for Nox1 activation. Previous studies demonstrate that cytokine activation of the pro- inflammatory factor nuclear factor-¿B (NF-¿B) requires internalization of Nox1 via endocytosis. The objective of this proposal is to identify novel regions within Nox1 that regulate its activatio and redox signaling to mediate migration and neointimal formation. The hypothesis is that membrane trafficking and phosphorylation of Nox1 are necessary for cytokine-induced Nox1 activation in SMCs. The following aims are proposed to test the central hypothesis: 1) Examine the functional consequences of cytokine-induced Nox1 trafficking in SMCs. 2) Define how phosphorylation of Nox1 regulates its trafficking and activation. 3) Determine whether the inhibition of Nox1 phosphorylation or trafficking provides therapeutic benefit in the prevention of neointimal hyperplasia. Proposed studies for the first aim will utilize site-directed mutagenesis o canonical internalization motifs within Nox1 to define how cytokine stimulation affects Nox1 trafficking as well Nox1-dependent ROS generation, NF-¿B activation, and SMC migration. For the second aim, a systems biology approach will be applied to quantitate the dynamic changes in Nox1 phosphorylation at specific amino acid residues. Next, these phosphorylation sites will be mutated to evaluate the importance of phosphorylation in the mechanisms of Nox1 activation. The third aim will use an in vivo model of injury to examine the role of Nox1 internalization and/or phosphorylation in neointimal formation. These studies have the potential to identify additional signaling events and motifis within Nox1 that are necessary for activation i SMCs. An immediate clinical impact of these studies is the potential to uncover alternative approaches to generate Nox1-targeted therapeutics for vascular pathologies.

描述（由申请人提供）： 心血管疾病的特征是血管壁中活性氧（ROS）的产生增加，这导致信号传导途径的激活，最终促进细胞生长和新内膜形成。源自平滑肌细胞（SMC）的ROS是血管疾病发展的主要因素，尽管抗氧化剂疗法仅获得了有限的治疗益处。因此，有必要确定预防ROS产生的目标方法。 NADPH氧化物是脉管系统中ROS的主要来源，NOX1是在SMC中表达的一级催化NADPH氧化物。 NOX1衍生的ROS已与动脉粥样硬化以及损伤后的新内膜形成和SMC迁移有关，但是NOX1激活氧化还原依赖的信号通路的精确机制仍未完全定义。为了开发针对NOX1的靶向治疗，首先需要了解必须满足NOX1激活的参数。先前的研究表明，促炎性因子核因子 - B（NF-€）的细胞因子激活需要通过内吞作用将NOX1内在化。该提案的目的是确定NOX1中的新区域，这些区域调节其活化和氧化还原信号传导以介导迁移和新内膜形成。假设是，NOX1的膜运输和光术对于SMC中的细胞因子诱导的NOX1激活是必需的。提出了以下目的来检验中心假设：1）检查SMC中细胞因子诱导的NOX1运输的功能后果。 2）定义NOX1的磷酸化如何调节其运输和激活。 3）确定抑制NOX1磷酸化或运输是否为预防新内膜增生提供治疗益处。提议的第一个目标研究将利用NOX1中的位置定向诱变o典型的内在化基序来定义细胞因子刺激如何影响NOX1运输以及NOX1依赖性ROS的产生，NF-€b激活和SMC迁移。对于第二个目标，将采用系统生物学方法来定量特定氨基酸保留下NOX1磷酸化的动态变化。接下来，这些磷酸化位点将被突变，以评估NOX1激活机理中磷酸化的重要性。第三个目标将使用体内损伤模型来检查NOX1内在化和/或磷酸化在新内膜形成中的作用。这些研究有可能在NOX1中识别激活I SMC所必需的其他信号事件和基序。这些研究的直接临床影响是发现为血管病理产生NOX1靶向疗法的替代方法。