Mechanism of Neutrophil Activation

中性粒细胞激活机制

• 批准号: 8762444
• 负责人: Kenneth R MCLEISH
• 金额: --
• 依托单位: LOUISVILLE VA MEDICAL 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: 2012-10-01 至 2016-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8762444
• 关键词: Acute; Acute Lung Injury; Address; Adult Respiratory Distress Syndrome; Affect; Aging; Animal Model; Antineutrophil Cytoplasmic Antibodies; Arteriosclerotic Cardiovascular Disease; Atherosclerosis; Bacteria; Biochemical Genetics; Biology; Cell membrane; Chemicals; Chronic; Cytoplasmic Granules; Data; Development; Disease; Event; Exocytosis; Exposure to; Gelatinases; Goals; Health; Human; In Vitro; Infection; Infection Control; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Arthritis; Injury; Intervention; Invaded; Knowledge; Laboratories; Lead; Leukocytes; MAP Kinase Gene; MAPK14 gene; Mediating; Membrane; Molecular; Molecular Target; NADPH Oxidase; Neutrophil Activation; Normal Cell; Oxidases; Patients; Peptidylprolyl Isomerase; Phosphorylation; Play; Poison; Population; Positioning Attribute; Process; Production; Proteins; Proteomics; Reagent; Relative (related person); Reperfusion Injury; Reporting; Rest; Rheumatoid Arthritis; Role; Secretory Vesicles; Signal Transduction Pathway; Testing; Therapeutic; Tissues; Toxin; Vasculitis; Veterans; Work; acute liver injury; adaptive immunity; bactericide; base; experience; extracellular; human CYBA protein; human disease; in vivo; killings; liver injury; lung vascular injury; microbial; neutrophil; novel; novel strategies; reactive oxygen intermediate; response; sound; therapeutic target; tool

DESCRIPTION (provided by applicant): Neutrophils are a critical component of both the innate and adaptive immune responses that control microbial invasion. Priming is an intermediate step by which resting neutrophils enhance their production of reactive oxygen intermediates (ROI) upon activation, and priming is necessary for optimal microbial killing. Although priming greatly enhances neutrophil bactericidal capability, primed neutrophils are also more dangerous to normal cells. Primed neutrophils participate in a number of diseases prominent in the aging Veteran population, including ANCA-associated vasculitis, ischemia- reperfusion injury, acute respiratory distress syndrome, rheumatoid arthritis, peridontitis, atherosclerosis, and acute inflammatory liver injury. Thus, manipulation of neutrophil priming is a potential therapeutic strategy for pharmacological intervention to restrain inflammation. Recent studies identified granule exocytosis and enhanced translocation of cytosolic components of the NADPH oxidase to the plasma membrane as possible mechanisms of neutrophil priming. However, many gaps in knowledge remain. The relative contribution of each of those events to priming has not been examined and whether both are necessary, or sufficient, for priming is unknown. The molecular events by which granule exocytosis contributes to priming and the molecular events that control enhanced translocation of cytosolic NADPH oxidase components are incompletely understood. Finally, the ability to manipulate neutrophil priming in vivo has not been established. The McLeish laboratory is in a unique position to address each of these gaps based on our extensive experience examining signal transduction pathways, our application of biochemical, genetic, and proteomic approaches to understand neutrophil biology, and our recent development of novel reagents that inhibit exocytosis in vitro and in vivo. The current proposal will address the central hypothesis that neutrophil priming results from the convergence of signal transduction pathways into two responses, an exocytose-dependent increase in plasma membrane expression of membrane components of NADPH oxidase and prolyl isomerase-dependent conformational changes in cytosolic components of NADPH oxidase. The following four specific objectives will be accomplished. Specific Objective 1 will test the working hypothesis that exocytosis of secretory vesicles and gelatinase granules results in increased plasma membrane expression of p91phox and p22phox, which is necessary, but not sufficient, for priming. Specific Objective 2 will test the working hypothesis that activation of the prolyl isomerase Pin1 contributes to priming of human neutrophils by multiple mechanisms, including enhanced p47phox translocation and increased granule exocytosis. Specific Objective 3 will test the working hypothesis that p38 MAPK phosphorylates multiple proteins that mediate events necessary for priming, including exocytosis and Pin1 activation. Specific Objective 4 will test the working hypothesis that neutrophil priming is a necessary component for neutrophil participation in inflammatory diseases and represents a viable therapeutic strategy, using an animal model of acute lung injury (ALI). The proposed work is expected to establish the molecular basis for neutrophil priming. The impact of the project is that molecular targets identified will permit therapeutic manipulation of neutrophil priming in inflammatory diseases. Thus, new approaches to treating Veteran patients with a number of acute and chronic inflammatory diseases are anticipated.

描述（由申请人提供）： 中性粒细胞是控制微生物侵袭的先天和适应性免疫反应的关键组成部分。启动是一个中间的步骤，静息中性粒细胞在激活后增强其活性氧中间体（ROI）的产生，而启动对于最佳的微生物杀死是必需的。尽管启动极大地增强了中性粒细胞的杀菌能力，但灌注的嗜中性粒细胞对正常细胞也更加危险。引发性嗜中性粒细胞参与了老年退伍军人人群中突出的多种疾病，包括与ANCA相关的血管炎，缺血性再灌注损伤，急性呼吸窘迫综合征，类风湿关节炎，围产期炎，动脉粥样硬化和急性炎性肝损伤。因此，对中性粒细胞启动的操纵是抑制炎症的药理干预的潜在治疗策略。最近的研究确定了NADPH氧化酶对质膜的颗粒胞吐作用和增强的胞质成分的易位，作为中性粒细胞启动的可能机制。但是，知识的许多差距仍然存在。这些事件中每个事件对启动的相对贡献尚未进行检查，而两者都是必要的，是否足以使启动是未知的。颗粒胞吐作用的分子事件有助于启动和控制胞质NADPH氧化酶成分易位的分子事件，这是不完全了解的。最后，尚未确定在体内操纵嗜中性粒细胞启动的能力。 McLeish实验室的独特位置可以根据我们的丰富经验来解决这些差距，从而研究信号转导途径，我们对了解嗜中性粒细胞生物学的生化，遗传和蛋白质组学方法的应用以及我们最近开发的新型试剂的发展，这些试剂抑制了体外和体内的胞外增生。当前的提案将解决以下中心假设：中性粒细胞启动是由信号转导途径转化为两个反应的收敛性，这是NADPH氧化酶和丙糖基异构酶依赖性构型变化的NADPH氧化酶成分的膜膜表达的胞外膜表达的增加。将实现以下四个特定目标。 具体目标1将检验以下假设：分泌囊泡和明胶酶颗粒的胞吐作用会导致p91phox和p22phox的质膜表达增加，这是必要但不足以进行启动。具体目标2将检验以下假设：丙酰异构酶PIN1的激活通过多种机制有助于人类嗜中性粒细胞的启动，包括增强的P47phox易位和增加的颗粒外胞毒性。特定目标3将检验p38 MAPK磷酸化的多种蛋白质的工作假设，这些蛋白质介导了启动所需的事件，包括胞吐作用和PIN1激活。具体目标4将测试 使用急性肺损伤动物模型（ALI），中性粒细胞启动是中性粒细胞参与炎症性疾病的必要组成部分，代表了可行的治疗策略。提出的工作有望建立中性粒细胞启动的分子基础。该项目的影响是，所鉴定的分子靶标将允许对炎症性疾病中嗜中性粒细胞启动的治疗操纵。因此，预计治疗患有许多急性和慢性炎症性疾病的老将患者的新方法。