Legionella-Polymorphonuclear Leukocytes Interaction

军团菌-多形核白细胞相互作用

• 批准号: 10057609
• 负责人: Yousef A Abu Kwaik
• 金额: $ 23.4万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-17 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10057609
• 关键词: Alveolar Macrophages; Amylases; Animals; Bacteria; Bacterial Infections; Biopsy; Cells; Cytoplasmic Granules; Cytosol; Data; Exposure to; Generations; Glucose; Glycogen; Growth; Host Defense; Human; Hyperglycemia; Immune; In Vitro; Infection; Infection Control; Inflammation; Inflammatory; Inflammatory Response; Innate Immune Response; Knowledge; Learning; Legionella; Legionella pneumophila; Leukocytes; Mediating; Metabolic; Metabolism; Molecular; NADPH Oxidase; Natural Immunity; Patients; Phagolysosome; Phagosomes; Pneumonia; Production; Proliferating; Proteins; Pulmonary alveolar structure; Reactive Oxygen Species; Reporting; Syringes; System; Testing; Up-Regulation; aerobic glycolysis; antimicrobial; antimicrobial drug; cytokine; diabetic; diabetic patient; glycogenolysis; in vivo; innate immune mechanisms; macrophage; microbial; monocyte; mutant; neutrophil; novel; pathogen; pathogenic bacteria; peripheral blood; response

Project Summary Polymorphonuclear leukocytes (PMNs)/Neutrophils account for 50-60% of peripheral blood leukocytes in humans, and are an essential part of the innate host defense against microbial infections. Quiescent PMNs respond to high levels of glucose in vitro through a pro-inflammatory response mediated by up-regulation of aerobic glycolysis. Importantly, PMNs respond to hyperglycemia in diabetic patients by releasing pro- inflammatory cytokines. PMNs restrict intracellular growth of Legionella pneumophila (Lp), whose ability to cause pneumonia is dependent on pathogen proliferation within alveolar macrophages. The Dot/Icm type IV translocation system of Lp, which functions as a molecular syringe, injects protein effectors into the macrophage cytosol to evade lysosomal fusion and macrophage restriction of pathogen proliferation. Despite the Dot/Icm-mediated translocation of effectors into the PMNs cytosol, the cells restrict Lp proliferation. The two main powerful antimicrobial machineries of PMNs to control microbial infections are the generation of reactive oxygen species (ROS) and the antimicrobial agents within the azurophilic and specific granules. It is not known whether any of the two major PMNs antimicrobial machineries are involved in Lp restriction. The PMNs mount a pro-inflammatory response to Lp in vivo, but the mechanism is not known. In contrast to macrophages, our preliminary data show that PMNs rapidly degrade the WT strain of Lp, while the Dot/Icm translocation-defective mutant survives for at least 4h. We discovered that one of the Dot/Icm-translocated effectors of Lp into the cytosol of human PMNs is a Legionella amylase (LamA), which degrades the PMNs glycogen within 30-60 min of infection. Interestingly, PMNs fail to degrade the lamA-deficient mutant, which does not degrade PMNs glycogen, similar to the Dot/Icm translocation-defective mutant. We will test the hypothesis that human PMNs respond to the effect of the translocated LamA effector by engaging their antimicrobial machineries in response to the cytosolic hyper-glucose generated by the LamA-mediated degradation of glycogen. To test the hypothesis, our specific aims are: Specific Aim I: Engagement of the PMNs antimicrobial machineries in response to the effect of LamA; and Specific Aim II: Pro-inflammatory activation of PMNs in response to cytosolic hyper-glucose generated through glycogen degradation by LamA. Upon completion of the proposed studies, we will learn the mechanism of the innate immune response of human PMNs to Lp in response to abnormally high levels of cytosolic glucose. Importantly, our studies have broad significance as they will be relevant to mechanisms of innate immunity under diabetic conditions where dysregulated inflammation is a common persistent occurrence. Our proposed studies are novel as well as mechanistic, and would generate a new paradigm in our knowledge of PMNs innate immune responses to bacterial pathogens.

项目摘要 多形核白细胞（PMNS）/中性粒细胞占外周血白细胞50-60％ 人类，是对微生物感染的先天宿主防御的重要组成部分。静止的PMN 通过上调介导的促炎反应来应对体外葡萄糖的高水平 有氧糖酵解。重要的是，PMN通过释放糖尿病患者的高血糖反应 炎症细胞因子。 PMN限制了肺炎军团菌（LP）的细胞内生长，其能力 引起肺炎，取决于肺泡巨噬细胞中的病原体增殖。点/ICM类型 LP的IV易位系统，该系统充当分子注射器，将蛋白质效应子注入 巨噬细胞胞质溶胶以逃避病原体增殖的溶酶体融合和巨噬细胞限制。尽管 DOT/ICM介导的效应子转移到PMNS细胞质中，细胞限制了LP增殖。这 PMN的两个主要强大抗菌机器以控制微生物感染 活性氧（ROS）和硫醇和特定颗粒中的抗菌剂。它 尚不清楚两个主要的PMN抗菌机器中是否涉及LP限制。 PMNS对体内LP进行了促炎反应，但该机制尚不清楚。与 巨噬细胞，我们的初步数据表明，PMN迅速降解了LP的WT菌株，而点/ICM 易位缺陷突变体至少生存4H。我们发现其中一个点/ICM转换为 LP进入人类PMN的胞质溶胶的效应子是军团甲基酶（LAMA），它降解了PMNS 糖原感染30-60分钟内。有趣的是，PMN无法降解缺乏喇嘛的突变体，该突变体 不会降解PMNS糖原，类似于点/ICM易位缺陷突变体。我们将测试 假设人PMN通过参与易位的喇嘛效应子的影响 响应喇嘛介导的胞质高葡萄糖的抗菌机器 糖原的降解。为了检验假设，我们的具体目的是：具体目的I：参与 PMNS抗菌机器响应喇嘛的作用；和特定的目标II：促炎性 PMN响应于喇嘛通过糖原降解产生的胞质高葡萄糖的激活。 拟议的研究完成后，我们将学习先天免疫反应的机制 人类PMN响应异常高的胞质葡萄糖而响应LP。重要的是，我们的研究有 广泛的意义，因为它们将与糖尿病条件下先天免疫的机制有关 炎症失调是常见的持续发生。我们提出的研究是新颖的 机理，并将在我们对PMN先天免疫反应的了解中产生一个新的范式 细菌病原体。