The Impact of SARS-CoV-2 Immune Dysregulation on Antifungal Immunity

SARS-CoV-2 免疫失调对抗真菌免疫的影响

基本信息

批准号：
10658355
负责人：
Sixto Manuel Leal
金额：
$ 74.25万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-04 至 2027-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10658355
关键词：
2019-nCoV Acceleration Affect Air Allergic Bronchopulmonary Aspergillosis Anabolism Antifungal Agents Apoptosis Aspergillus Aspergillus fumigatus Bacterial Pneumonia Biological Biological Availability Biological Models Biology Bronchioles Bronchoalveolar Lavage Bronchoalveolar Lavage Fluid Cell Culture Techniques Cell Line Cells Clinical Clustered Regularly Interspaced Short Palindromic Repeats Critical Pathways Cytolysis Development Divalent Cations Epithelial Cells Epithelium Erythrocytes Evaluation Exhibits Exposure to FDA approved Flow Cytometry Functional disorder Gene Expression Germination Growth Heme Hemolysis Human Hyphae Immune Immune response Immune system Immunity Immunocompetent Immunofluorescence Immunologic Impairment In Situ In Vitro Individual Infection Infection Control Influenza Inhalation Therapy Intensive Care Units Interferons Iron Iron Chelation Knock-out Knockout Mice Laboratories Liquid substance Lung Lytic Macrophage Mediating Meta-Analysis Metals Microbe Modeling Molds Molecular Mus Mycoses Nutrient availability Outcome Pathway interactions Patients Predisposition Production Proteins Publications Publishing Pulmonary Pathology Regimen Reporter Reproduction spores Role SARS-CoV-2 antiviral SARS-CoV-2 infection SARS-CoV-2 variant Sampling Siderophores Signal Transduction Stimulus Structure of parenchyma of lung Surface Syndrome Testing Therapeutic Therapeutic Intervention Toxic effect Viral Viral Pneumonia Virus Diseases chemokine cohort conditional knockout coronavirus disease cytokine fungus improved in vivo Model inhibitor innovation metal chelator microbial migration mortality mouse model mutant neutrophil novel pathogenic virus pharmacologic pre-clinical prevent recruit risk mitigation severe COVID-19 single cell sequencing small molecule inhibitor transcriptomics uptake

项目摘要

Project Summary Despite hundreds of published articles, meta-analyses, and reviews describing the clinical syndrome of COVID-Associated Pulmonary Aspergillosis (CAPA), there have been zero publications to date exploring the mechanism by which individuals with severe SARS-2 infection succumb to 2° mold infection. This proposal seeks to evaluate the novel conceptual advancement that immune responses targeting intracellular viral pathogens, like SARS-2, promote 2° mold infection via release of bioavailable iron, delayed neutrophil (PMN) recruitment, and decreased antifungal effector (αFE) expression. In Aims 1-3, we explore the hypotheses that lytic programmed cell death (PCD) in pulmonary epithelial cells (PEC) and red blood cell hemolysis increase the availability of nutrients including iron/heme which drive siderophore- dependent acceleration in mold growth. Concurrently, SARS-2 and the antiviral cytokine milieu mitigate PMN recruitment and activation resulting in spore germination into large invasive hyphae that overwhelm αFE mechanisms enabling 2° mold infection in an otherwise immunocompetent host. In Aim1, we will quantify iron, heme, divalent cations, host metal sequestration proteins (MSP), antiviral cytokines, and PMN-recruiting chemokines in SARS-2 BALs vs control infection cohorts (IC). We will also use immunofluorescence (IF) and spatial transcriptomics to characterize lytic PCD, viral ORF3a-mediated cell lysis, and αFE expression in FFPE lung tissues and utilize isogenic mutant fungi to identify critical host and microbial factors mediating mold growth in BALs. In Aim2, we will utilize novel Calu-3 knockout (KO) cell lines and 1°normal human bronchiole epithelial (NHBE) air liquid interface cell cultures to study the impact of the lower airway milieu and direct SARS-2 infection on PEC: lytic PCD, IFN synthesis, αFE secretion, metal sequestration, and ability to respond to 2° fungal stimuli. We will also utilize isogenic mutant fungi to identify critical pathways mediating mold growth and an innovative PMN airway transmigration model to determine the impact of Type I and III IFNs, iron/heme toxicity, and SARS-2 uptake on PMN recruitment, activation, and fungal killing. In Aim3, we will utilize novel conditional KO mice, fluorescent viral and fungal reporter strains, IF, flow cytometry, and single cell sequencing to assess the potential of SARS-2 variants to induce lytic PCD, define the role of lytic PCD and IFN signaling in PECs, macrophages (Mφ), and PMNs, and evaluate their impact on lung pathology, metal release, PMN- recruitment, spore viability, fungal growth, and infection outcome. We will also dissect key host MSP and fungal pathways mediating mold growth, evaluate the role of Mφ and PMN hACE2 on viral and fungal clearance and determine if small molecule inhibitors of lytic PCD, iron chelators, and siderophore biosynthesis inhibitors prevent or mitigate the development of 2° mold infection. We believe that the results of the proposed study will shed new light on the fundamental biology mediating CAPA and may enable the development of improved therapeutic regimens that mitigate the risk of acquiring and succumbing to this devastating infection.

项目概要尽管有数百篇发表的文章、荟萃分析和评论描述了临床综合征与新冠肺炎相关的肺曲霉病 (CAPA)，迄今为止探索该疾病的出版物为零严重 SARS-2 感染者死于 2° 霉菌感染的机制。旨在评估针对细胞内的免疫反应的新概念进展病毒病原体，如 SARS-2，通过释放生物可利用的铁促进 2° 霉菌感染，延迟在目标 1-3 中，我们发现中性粒细胞 (PMN) 募集和抗真菌效应子 (αFE) 表达减少。探索肺上皮细胞（PEC）中的裂解性程序性细胞死亡（PCD）和红细胞的假设血细胞溶血增加了营养物质的可用性，包括驱动铁载体的铁/血红素同时，SARS-2 和抗病毒细胞因子环境可减轻 PMN。招募和激活导致孢子萌发成大型侵入性菌丝，压倒αFE 在具有免疫能力的宿主中实现 2° 霉菌感染的机制在 Aim1 中，我们将量化铁，血红素、二价阳离子、宿主金属螯合蛋白 (MSP)、抗病毒细胞因子和 PMN 募集 SARS-2 BAL 中的趋化因子与对照感染队列 (IC) 我们还将使用免疫荧光 (IF) 和空间转录组学表征裂解性 PCD、病毒 ORF3a 介导的细胞裂解和 FFPE 中的 αFE 表达肺组织并利用同基因突变真菌来识别介导霉菌的关键宿主和微生物因素在 Aim2 中，我们将利用新型 Calu-3 敲除 (KO) 细胞系和 1° 正常人细支气管。上皮 (NHBE) 气液界面细胞培养物，用于研究下呼吸道环境和直接影响 PEC 上的 SARS-2 感染：裂解 PCD、IFN 合成、αFE 分泌、金属螯合和反应能力至 2° 真菌刺激我们还将利用同基因突变真菌来确定介导霉菌生长的关键途径。以及创新的 PMN 气道迁移模型，以确定 I 型和 III 型干扰素、铁/血红素的影响在 Aim3 中，我们将利用新颖的方法来研究 SARS-2 的毒性和 SARS-2 摄取对 PMN 募集、激活和杀灭的影响。条件 KO 小鼠、荧光病毒和真菌报告菌株、IF、流式细胞术和单细胞测序评估 SARS-2 变异体诱导裂解性 PCD 的潜力，定义裂解性 PCD 和 IFN 信号传导的作用中性粒细胞 (PEC)、巨噬细胞 (Mφ) 和中性粒细胞 (PMN)，并评估它们对肺病理学、金属释放、中性粒细胞 (PMN) 的影响我们还将剖析关键宿主 MSP 和真菌。介导霉菌生长的途径，评估 Mφ 和 PMN hACE2 对病毒和真菌清除的作用，以及确定是否是裂解性 PCD 的小分子抑制剂、铁螯合剂和铁载体生物合成抑制剂我们相信所提出的研究结果可以预防或减轻 2° 霉菌感染的发展。将为介导CAPA的基础生物学提供新的线索，并可能促进改进的开发减轻感染和死于这种毁灭性感染的风险的治疗方案。