COVID-19: Understanding The Role of Corona Virus InducedDisruption Of Alveolar Type 2 Cell Function And SurfactantHomeostasis In The Pathogenesis Of COVID-19 AcuteRespiratory Distress Syndrome

COVID-19：了解冠状病毒引起的肺泡 2 型细胞功能和表面活性剂稳态破坏在 COVID-19 急性呼吸窘迫综合征发病机制中的作用

• 批准号: 10744174
• 负责人: MICHAEL FRANCIS BEERS
• 金额: --
• 依托单位: PHILADELPHIA VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2023-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10744174
• 关键词: 2019-nCoV; ACE2; Acceleration; Acute Lung Injury; Acute Respiratory Distress Syndrome; Address; Affect; Alveolar; Anabolism; Apoptosis; Autophagocytosis; Behavior; Berlin; Big Data; Biochemistry; Bioenergetics; Biology; Biophysics; COVID-19; COVID-19 pandemic; COVID-19 pathogenesis; Caring; Catalogs; Cell Communication; Cell Ontogeny; Cell model; Cell physiology; Cells; Cellular Stress; Cellular biology; Characteristics; Cicatrix; Clinical; Coronavirus; Coronavirus Infections; Coupled; Data; Defect; Development; Disease; Dissociation; Distal; Distress; Economics; Epidemic; Epithelial Cells; Epithelium; Evaluation; Foundations; Functional disorder; Funding; Future; Genetic; Goals; Growth; Hand; Health; Homeostasis; Human; Hypoxemia; Hypoxemic Respiratory Failure; Impairment; In Vitro; Infection; Inflammatory; Innate Immune Response; International; Investigation; Knowledge; Lead; Link; Lung; Lung Capacity; Lung infections; Maintenance; Maps; Mediating; Metabolism; Middle East Respiratory Syndrome Coronavirus; Mitochondria; Modeling; Molecular; Molecular Target; Morbidity - disease rate; Motivation; Mus; Natural Immunity; Organ; Organoids; Outcome; Pathogenesis; Pathway interactions; Peripheral; Phenotype; Physiological; Population; Positioning Attribute; Pre-Clinical Model; Program Reviews; Proteins; Pulmonary Inflammation; Pulmonary Surfactants; Quality Control; Radiology Specialty; Reagent; Recovery; Recurrence; Refractory; Reporting; Research Design; Residual state; Respiratory Disease; Respiratory Failure; Respiratory Mechanics; Role; SARS coronavirus; SARS-CoV-2 infection; Scientist; Severities; Signal Pathway; Slice; Survivors; Syndrome; System; Techniques; Technology; Testing; Therapeutic; Translating; Ubiquitin; Veterans; Viral; Virus; Virus Diseases; Work; alveolar epithelium; arm; biological adaptation to stress; coronavirus pandemic; cytokine; effective therapy; endophenotype; endoplasmic reticulum stress; epithelial injury; epithelial repair; experience; fibrotic lung; immunopathology; improved; in vivo; induced pluripotent stem cell; lipid metabolism; lung injury; lung repair; mortality; mouse model; multicatalytic endopeptidase complex; new therapeutic target; novel; novel coronavirus; novel strategies; pandemic disease; pharmacologic; pre-clinical; progenitor; programs; protein kinase R; protein metabolism; receptor; repaired; respiratory; response; senescence; stem cell function; stem cells; surfactant; tool; transcriptomic profiling; transcriptomics; virology

ABSTRACT The Severe Acute Respiratory Syndrome (SARS)-associated coronavirus 2 (SCoV2) is the cause of COVID-19 syndrome which is marked by a refractory acute lung injury that results in dramatic hypoxic respiratory failure and high mortality. Despite the recurrent health and economic devastation produced by novel coronaviruses (nCoV) over the past 20 years including SCoV2 as well as its predecessors SCoV1, and MERS-CoV, there remains a significant unmet need both for a clearer understanding of virus-host cell interactions as well as identification of new therapeutic targets. To address these issues with the ultimate goal of improving the health and outcomes of veterans with COVID-19, we have assembled a team of internationally recognized scientists with expertise in coronavirus virology and in lung biology coupled with a strong foundation of our own prior work on surfactant biology, lung injury, and fibrotic repair funded, in part, by the VA Merit Review program. Utilizing this expertise, this proposal is directed at filling in large knowledge gaps that exist in the pathogenesis of nCoV induced respiratory failure. The motivation for this investigation has been fueled by the recent recognition that the alveolar type 2 (AT2) epithelial cell of the distal lung has emerged as an important portal of entry for SCoV- 2. The central hypothesis of this application is that AT2 cells infected with nCoV acquire defects in surfactant biosynthesis/metabolism, activate cellular stress pathways, and develop alterations in progenitor cell function all of which promote hypoxic respiratory failure, persistent lung inflammation and injury, and impact recovery through effects on epithelial repair capacity. To test this, we will leverage an established murine model of CoV infection (MHV-1) with a pulmonary phenotype combined with reductionist studies supported by ex vivo infection of primary human AT2 cells obtained from a robust human lung pipeline with an already in hand SARS-CoV-2 isolate. Our experimental approach will interrogate these preclinical models using tools and reagents available in our program, to map the effect of CoV infection on distal lung cell populations with a focus on identifying and translating molecular mechanisms linking the disrupted AT2 function with altered surfactant biology and proinflammatory/profibrotic cell cross talk in the alveoilar niche. In Specific Aim 1, we will first define temporal changes in distal lung epithelial endophenotypes focusing on the ontogeny of the disruption of AT2 homeostasis by viral infection using a well characterized mouse model of MHV-1 lung infection. Using both unbiased approaches such as transcriptomic profiling as well as classical cell biology and biochemistry this aim will investigate CoV induced changes in surfactant metabolism/biophysical activity, AT2 cell stress (i.e. ER stress, proteasome dysfunction, autophagy malfunction, changes in mitochondrial dynamics/ / bioenergetics) and AT2 progenitor cell function. In Specific Aim 2, armed with this “functional map” of the mouse CoV-lung and aberrant AT2 behaviors, we will translate the identified lead targets and mechanisms to human AT2 cells utilizing both primary AT2 cultures as well as precision cut lung slices (PCLS) infected with SARS-CoV-2. The research design involves the analysis of virus-dependent AT2 surfactant protein and lipid metabolism, interrogation of AT2 cell quality control pathways, and functional evaluation of AT2 progenitor function phenotypes in the SCoV-2 infected human epithelial cells. By understanding the path to epithelial injury / dysfunction from nCoV, the mechanisms and affected lung cell populations identified using these models will improve the understanding of sCOV2/COVID- 19 syndrome, promote identification of new target pathways, and can be cross-purposed to test emerging therapies for both the current and future nCoV pandemics.

摘要 严重急性呼吸综合征（SARS）相关冠状病毒2（SCoV 2）是COVID-19的病因 一种以难治性急性肺损伤为特征的综合征，导致严重的缺氧性呼吸衰竭 和高死亡率。尽管新型冠状病毒一再造成健康和经济破坏， 在过去的20年里，包括SCoV 2及其前身SCoV 1和MERS-CoV， 对于更清楚地了解病毒-宿主细胞相互作用以及 确定新的治疗靶点。为了解决这些问题，最终目标是改善健康状况， 和退伍军人与COVID-19的结果，我们组建了一个国际公认的科学家团队， 在冠状病毒学和肺部生物学方面的专业知识，加上我们自己先前工作的坚实基础， 表面活性剂生物学，肺损伤和纤维化修复，部分由VA Merit Review计划资助。利用 这项建议旨在填补nCoV发病机制中存在的巨大知识空白 导致呼吸衰竭。最近认识到， 远端肺的肺泡2型（AT 2）上皮细胞已成为SCoV进入的重要门户， 2.本申请的中心假设是感染nCoV的AT 2细胞获得表面活性剂缺陷， 生物合成/代谢，激活细胞应激途径，并发展祖细胞功能的改变， 其中促进缺氧性呼吸衰竭、持续性肺部炎症和损伤，并影响恢复 通过影响上皮修复能力。为了验证这一点，我们将利用已建立的CoV小鼠模型 感染（MHV-1）与肺部表型结合的还原论研究支持离体感染 原代人AT 2细胞从一个强大的人肺管道获得，已经在手中的SARS-CoV-2 孤立。我们的实验方法将使用现有的工具和试剂来询问这些临床前模型 在我们的项目中，绘制CoV感染对远端肺细胞群的影响，重点是识别和 翻译分子机制将破坏的AT 2功能与改变的表面活性剂生物学联系起来， 肺泡龛中的促炎症/促纤维化细胞串扰。在具体目标1中，我们将首先定义时间 远端肺上皮细胞内表型的变化，重点关注AT 2稳态破坏的个体发生 使用充分表征的MHV-1肺部感染小鼠模型通过病毒感染。使用无偏 例如转录组学分析以及经典细胞生物学和生物化学的方法， 研究CoV诱导的表面活性剂代谢/生物物理活性、AT 2细胞应激（即ER应激， 蛋白酶体功能障碍、自噬功能障碍、线粒体动力学/ /生物能量学的变化）和AT 2 祖细胞功能在特异性目标2中，配备了小鼠CoV肺和异常肺的“功能图”， AT 2的行为，我们将利用这两种方法将所确定的先导靶标和机制翻译到人类AT 2细胞中。 原代AT 2培养物以及感染SARS-CoV-2的精密切割肺切片（PCLS）。研究设计 包括分析病毒依赖的AT 2表面活性蛋白和脂质代谢，询问AT 2细胞 质量控制途径，以及在SCoV-2感染中AT 2祖细胞功能表型的功能评价 人类上皮细胞通过了解nCoV导致上皮损伤/功能障碍的途径， 使用这些模型识别的受影响的肺细胞群将提高对sCOV 2/COVID的理解， 19综合征，促进识别新的目标途径，并可以交叉目的测试新兴 治疗当前和未来的nCoV大流行。