Low Density Neutrophils Link Inflammation and Coagulopathy in COVID-19

低密度中性粒细胞与 COVID-19 中的炎症和凝血病有关

基本信息

批准号：
10276657
负责人：
Jiapeng Huang
金额：
$ 76.14万
依托单位：
UNIVERSITY OF LOUISVILLE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-23 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10276657
关键词：
2019-nCoV Acute Lung Injury Acute Respiratory Distress Syndrome Alpha Granule Antibodies Apoptosis Bilateral Blood Circulation Blood Coagulation Disorders Blood capillaries Blood specimen Bronchoalveolar Lavage Fluid COVID-19 COVID-19 patient COVID-19 severity COVID-19/ARDS Cells Chemotaxis Chimeric Proteins Clinical Coagulation Process Custom Cytoplasmic Granules Data Diffuse Disease Disease Outcome Disease Progression Economics Endothelial Cells Exocytosis Exocytosis Inhibition FCGR3B gene Fibrin fragment D Fibrinogen Functional disorder Gene Expression Hemorrhage Histologic Hypoxia Immune Impairment In Vitro Inflammation Inflammation Mediators Inflammatory Inflammatory Response Interleukin-6 Life Link Lower respiratory tract structure Lung Lymphocyte Measures Mediating Modeling Neutrophil Infiltration Outcome Patients Pattern Perfusion Phagocytosis Plasma Platelet Activation Platelet Count measurement Play Pneumonia Population Predisposing Factor Proteome Proteomics Pulmonary Inflammation Reactive Oxygen Species Respiratory Burst Role SARS-CoV-2 negative Sampling Severity of illness Structure of parenchyma of lung Surface T-Lymphocyte TNF gene Therapeutic Therapeutic Intervention Thrombus Time Venous Viral antimicrobial base cell injury cytokine cytokine release syndrome cytotoxic density design exhaustion experience extracellular immune activation immunoregulation immunothrombosis in vivo inhibitor insight lung injury mouse model neutrophil novel novel marker novel therapeutic intervention pandemic disease pathogen pathogenic virus peripheral blood prevent response therapeutic target therapeutically effective transcriptome sequencing transcriptomics treatment effect ventilation

项目摘要

Project Summary Coronavirus disease 2019 (COVID-19) is a potentially life threatening disease caused by the novel viral pathogen, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Approximately 20% of COVID-19 patients experience severe disease, typically presenting with bilateral pneumonia, and about 5% progress to acute respiratory distress syndrome (ARDS). ARDS results from a combination of virally induced lung injury and the rapid influx of immune cells that release inflammatory mediators leading to a hyper-activated state known as cytokine storm. COVID-19 ARDS is further exacerbated by a unique diffuse coagulopathy leading to thrombus formation in the venous and arterial circulations and microthrombi in capillaries of the lungs. Predisposing factors for this coagulopathy include increased fibrinogen, activated coagulation cascade, platelet activation, hyper- inflammation, neutrophil extracellular trap (NET) formation, and endothelial cell damage. Understanding the pathophysiology of COVID-19 coagulopathy and ARDS is critical to finding effective therapeutic interventions. Accumulating evidence indicates critical roles of neutrophils in both ARDS and immunothrombosis in COVID-19. Our preliminary studies identified a novel population of low-density neutrophils (LDN) which expresses intermediate levels of CD16 (CD16Int LDN) in COVID-19 patients. The number of CD16Int LDN correlated with disease severity, levels of inflammatory cytokines IL-6/TNF-a, D-dimer levels, and clinical outcomes. In addition, CD16Int LDN showed spontaneous NET formation and evidence of in vivo platelet activation and granule exocytosis. Based on these findings, we postulate that CD16Int LDN play a critical role in the induction of coagulopathy and pulmonary inflammation in severe and critical COVID-19 patients. Three specific Aims are proposed to further dissect the underlying mechanisms. Aim 1 will comprehensively characterize LDN subsets using proteomics and transcriptomics approaches. The information gained from those studies will be used to refine our CyTOF antibody panel. We will use this panel to track differential neutrophil clusters in longitudinal patient samples. Aim 2 will determine LDN subsets functional changes during disease progression and their contributions to dysregulated inflammatory response and coagulopathy in severe and critical COVID- 19 patients. Neutrophil degranulation, NET formation, phagocytosis, chemotaxis, apoptosis, and cytokine release will be examined. We will also determine if LDN promote coagulopathy in COVID-19 patients. Aim 3 will determine whether inhibition of neutrophil granule exocytosis using our novel TAT-fusion protein inhibitors prevents activated neutrophil-mediated functional changes and hypercoagulation. We will also use a hACE2 Tg mouse model to determine the in vivo efficacy of TAT-fusion proteins on lung inflammation and impaired function. Successful completion of this proposal will provide novel insights into COVID-19 pathophysiology by defining the role of a unique subset of neutrophils and by establishing neutrophil degranulation as a therapeutic target for inhibiting inflammatory lung injury and immunothrombosis in COVID-19.

项目概要 2019 年冠状病毒病 (COVID-19) 是一种由新型病毒引起的可能危及生命的疾病病原体，严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2)。大约 20% 的 COVID-19 患者病情严重，通常表现为双侧肺炎，约 5% 进展为急性呼吸窘迫综合征（ARDS）。 ARDS 由病毒引起的肺损伤和免疫细胞快速涌入，释放炎症介质，导致过度激活状态，称为细胞因子风暴。独特的弥漫性凝血病导致血栓，进一步加剧了 COVID-19 ARDS 静脉和动脉循环中的形成以及肺部毛细血管中的微血栓。诱发因素对于这种凝血病，包括纤维蛋白原增加、凝血级联激活、血小板活化、高凝血炎症、中性粒细胞胞外陷阱（NET）形成和内皮细胞损伤。了解 COVID-19 凝血病和 ARDS 的病理生理学对于寻找有效的治疗干预措施至关重要。越来越多的证据表明，中性粒细胞在 ARDS 和 COVID-19 免疫血栓形成中发挥着关键作用。我们的初步研究发现了一个新的低密度中性粒细胞 (LDN) 群体，其表达 COVID-19 患者的 CD16 (CD16Int LDN) 处于中等水平。 CD16Int LDN 的数量与疾病严重程度、炎症细胞因子 IL-6/TNF-a 水平、D-二聚体水平和临床结果。此外， CD16Int LDN 显示自发 NET 形成以及体内血小板活化和颗粒的证据胞吐作用。基于这些发现，我们假设 CD16Int LDN 在诱导重症和危重症 COVID-19 患者的凝血功能障碍和肺部炎症。三个具体目标建议进一步剖析潜在机制。目标 1 将全面表征 LDN 使用蛋白质组学和转录组学方法的子集。从这些研究中获得的信息将是用于完善我们的 CyTOF 抗体组。我们将使用此面板来跟踪中性粒细胞簇的差异纵向患者样本。目标 2 将确定疾病进展期间 LDN 子集的功能变化及其对严重和危重新冠肺炎患者炎症反应失调和凝血病的贡献 19名患者。中性粒细胞脱颗粒、NET 形成、吞噬作用、趋化性、细胞凋亡和细胞因子释放将被检查。我们还将确定 LDN 是否会促进 COVID-19 患者的凝血功能障碍。目标3将确定使用我们的新型 TAT 融合蛋白抑制剂是否抑制中性粒细胞颗粒胞吐作用防止激活的中性粒细胞介导的功能变化和高凝状态。我们还将使用 hACE2 Tg 小鼠模型以确定 TAT 融合蛋白对肺部炎症和功能受损的体内功效。该提案的成功完成将为 COVID-19 病理生理学定义提供新的见解中性粒细胞的独特亚群的作用，并建立中性粒细胞脱颗粒作为治疗靶点抑制 COVID-19 中的炎症性肺损伤和免疫血栓形成。