Role of Macrophage Efferocytosis in Ozone-Induced ARDS

项目摘要/摘要 急性呼吸窘迫综合征(ARDS)在一些个体中发展为间接应激的后遗症 全身感染(败血症/内毒素血症)对肺部的影响。然而，目前尚不清楚为什么只有一些患者患有 败血症会发展成ARDS。导致败血症患者发生ARDS的一个可能的危险因素是暴露在空气中 臭氧等污染物。最近，FDA可接受的臭氧暴露环境水平已经达到 与急性呼吸窘迫综合征的发展直接相关。我们的总体目标是阐明 暴露在臭氧等氧化剂中后，患ARDS的风险增加。ARDS在一定程度上发展了 由于死亡和濒临死亡的中性粒细胞和中性粒细胞衍生的促炎细胞凋亡小体积聚 在肺里。在动态平衡条件下，巨噬细胞通过一种称为 泡泡细胞增多症。我们假设暴露在臭氧下的ARDS风险增加是由于受损。 泡泡细胞增多症。此外，在肺集合素基因缺陷的个体中，这种情况会加剧， 表面活性蛋白D(SPD)，它控制巨噬细胞的泡腾作用。为了测试这一点，我们开发了一部小说 小鼠吸入臭氧后静脉注射的实验模型。 脂多糖(LPS)，一种细菌衍生的毒素，在败血症(内毒素血症)时释放到血液中。我们的 目的是(1)确定臭氧暴露和SPD活性降低是否会加剧炎症和急性 肺损伤(ALI)通过损害巨噬细胞泡出功能和(2)确定SPD活性是否降低 加剧臭氧对人类巨噬细胞泡出功能的损害。野生型和肺特异性 条件性SPD基因敲除的小鼠将接受臭氧和内毒素治疗。巨噬细胞泡沫化会 用流式细胞仪检测。与氧化应激相关的机械途径，这在 臭氧毒性，将使用RNA测序(RNAseq)进行鉴定。我们将分析肺部炎症和 受试者的巨噬细胞泡出作用，根据单核苷酸多态进行分层 Spd基因，在受控臭氧暴露后。这些实验的结果将提供新的 臭氧暴露、巨噬细胞功能、SPD变异和 对ARDS的易感性。这些研究意义重大，因为臭氧等氧化剂被认为是 危险因素是ARDS发生的危险因素。中提出的实验、课程作业和结构化指导 这份申请将为NIH R01拨款提供基础，并开启PI在独立学院的职业生涯 翻译研究。

Project Summary/Abstract Acute respiratory distress syndrome (ARDS) develops in some individuals as a sequela to indirect stress on the lung from systemic infection (sepsis/endotoxemia). However, it is unclear why only some patients with sepsis develop ARDS. One possible risk factor leading to ARDS in patients with sepsis is exposure to air pollutants such as ozone. Recently, FDA acceptable environmental levels of ozone exposure have been directly linked to the development of ARDS. Our overall goal is to elucidate the mechanisms underlying the increased risk of developing ARDS following exposure to oxidants such as ozone. ARDS develops, in part, due to an accumulation of dead and dying neutrophils and neutrophil-derived proinflammatory apoptotic bodies in the lung. Under homeostatic conditions, these are removed by macrophages via a process known as efferocytosis. We hypothesize that the increased risk of ARDS following ozone exposure is due impaired efferocytosis. Moreover, this is exacerbated in individuals with genetic deficits in the pulmonary collectin, surfactant protein D (SPD), which controls macrophage efferocytosis. To test this, we developed a novel experimental model in which mice are exposed to inhaled ozone followed by intravenous (i.v.) lipopolysaccharide (LPS), a bacterial-derived toxin released into the blood during sepsis (endotoxemia). Our aims are to (1) Determine if ozone exposure and decreased SPD activity exacerbate inflammation and acute lung injury (ALI) by impairing macrophage efferocytosis and (2) Determine if decreased SPD activity exacerbates ozone-induced impairment of macrophage efferocytosis in humans. Wild type and lung-specific conditional SPD knock out mice will be treated with ozone followed by LPS. Macrophage efferocytosis will be measured by flow cytometry. The mechanistic pathways associated with oxidative stress, which is important in ozone toxicity, will be identified using RNA sequencing (RNAseq). We will analyze lung inflammation and macrophage efferocytosis in human subjects, stratified according to single nucleotide polymorphisms within the SPD gene, following controlled ozone exposure. The results of these experiments will provide novel mechanistic insights into the relationship between ozone exposure, macrophage function, SPD variation, and susceptibility to ARDS. These studies are significant, as oxidants such as ozone have been implicated as a risk factor the development of ARDS. The experiments, coursework, and structured mentorship proposed in this application will provide the basis for an NIH R01 grant and initiate the PI's career in independent translational research.