Alveolar responses to viral lung infection

肺泡对病毒性肺部感染的反应

基本信息

批准号：
10650415
负责人：
Jaime Lynn Hook
金额：
$ 48.59万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
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：
2022-06-20 至 2027-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10650415
关键词：
Accounting Acute Lung Injury Address Adult Air Alveolar Alveolar wall Alveolus Anatomy Apical Bacteria Bioenergetics COVID-19 pandemic Catalytic Domain Cell Culture Techniques Cells Cessation of life Child Complex Confocal Microscopy Critical Illness Cultured Cells Cystic Fibrosis Transmembrane Conductance Regulator Data Defense Mechanisms Edema Enzymes Epithelium Excess Mortality Film Goals Hemolysin Hospitalization Hour Human Image In Situ In Vitro Influenza Influenza A virus Inhalation Ion Channel Leucine Lipids Liquid substance Lung Lung infections Measures Mediating Membrane Membrane Proteins Methods Methylation Methyltransferase Mission Modeling Molecular Mus National Heart, Lung, and Blood Institute Particle Size Pathogenesis Pathologic Patients Persons Pharmaceutical Preparations Phosphorylation Physiology Population Positioning Attribute Protein Dephosphorylation Protein Methylation Protein phosphatase Proteins Pulmonary Edema Pulmonary alveolar structure Research Resistance Respiratory Failure Role Secondary to Small Interfering RNA Staphylococcus aureus Staphylococcus aureus infection Structure Surface Testing Time Toxin Transgenic Mice United States Viral Viral Interference Virus Wild Type Mouse alveolar epithelium aqueous co-infection future pandemic influenza infection inhibitor insight knock-down lung injury mortality mouse model mutant new therapeutic target novel therapeutic intervention pandemic virus particle plasmid DNA prevent protective effect protein expression real-time images response restoration secondary infection success therapeutic evaluation

项目摘要

PROJECT SUMMARY / ABSTRACT Significance. One-third of patients with severe lung infection by influenza A virus (IAV) develop secondary infection by inhaled Staphylococcus aureus (SA). Coinfection by IAV and SA causes about 30% mortality despite therapy. It remains unclear how IAV promotes secondary SA infection, particularly in lung alveoli. This issue is important because alveoli are the anatomical site of fatal SA-induced Acute Lung Injury (ALI), but alveolar defense mechanisms, including alveolar wall liquid (AWL) secretion, should prevent SA stabilization and coinfection initiation. The long-term objective of this proposal is to determine alveolar responses to IAV that promote secondary SA infection in alveoli, resulting in SA-induced alveolar damage, ALI, and mortality. The hypothesis is IAV lung infection inhibits AWL secretion, a homeostatic mechanism by which alveoli clear inhaled particles. The inhibition causes alveolar retention of SA and the secreted SA toxin, alpha hemolysin (Hla). The retention enhances alveolar contact with SA and Hla, promoting SA stabilization against the alveolar wall and Hla-induced alveolar fluid barrier loss, leading to alveolar edema and fatal ALI. In addition to directly supporting the hypothesis, preliminary data indicate IAV lung infection caused: (A) dephosphorylation, hence inactivation of the alveolar cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel, the critical protein for AWL secretion; and (B) methylation of the CFTR-dephosphorylating protein, protein phosphatase 2A (PP2A) catalytic subunit (PP2Ac), which may promote PP2Ac-CFTR interactions. Specific Aims are as follows. Aim 1 will define the role of CFTR in alveolar retention of inhaled SA. Aim 2 will define the role of PP2Ac methylation in IAV-induced inhibition of AWL secretion. Since our preliminary data suggest CFTR and the PP2Ac- methylating enzyme, leucine carboxyl methyltransferase 1 (LCMT1) may represent new therapeutic targets to restore AWL secretion in IAV-infected lungs, Aim 3 will test the therapeutic potential of CFTR- and LCMT1- targeted approaches to protect against coinfection-induced alveolar damage, ALI, and mortality. These Aims will be achieved using our established methods, which include cell culture, mouse models of ALI, and real-time confocal microscopy of live, intact mouse and human lungs. Determinations in IAV-infected mice will include measures of: (1) alveolar retention of SA; (2) AWL secretion; (3) alveolar CFTR phosphorylation status; (4) alveolar PP2Ac methylation status; (5) SA- and Hla- alveolar epithelial damage and alveolar barrier loss; and (6) SA-induced pulmonary edema and mortality. We will use: (i) wild type mice treated with inhibitors of alveolar PP2Ac-CFTR and PP2Ac-LCMT1 interactions, including drug inhibitors, plasmid DNA encoding mutant proteins, and siRNA; and (ii) transgenic mice lacking alveolar epithelial CFTR and LCMT1 expression. This proposal is expected to achieve new insights into the molecular mechanisms by which IAV disrupts critical alveolar function leading to fatal ALI, and to establish restoration of AWL secretion – that is, “AWL rescue” – as a new therapeutic approach for ALI caused by IAV-SA coinfection. Therefore, this proposal addresses the NHLBI mission.

项目摘要 /摘要意义。因影响Za A病毒（IAV）发育次级患有严重肺部感染的患者中有三分之一吸入金黄色葡萄球菌（SA）感染。 IAV和SA的共感染导致约30％的死亡率需求治疗。目前尚不清楚IAV如何促进继发性SA感染，尤其是在肺肺泡中。这个问题是重要的是因为肺泡是致命SA诱导的急性肺损伤（ALI）的解剖部位，但肺泡包括肺泡壁液（AWL）分泌在内的防御机制应防止SA稳定和共同感染计划。该提案的长期目标是确定对IAV的肺泡反应促进肺泡中的SA SA感染，从而导致SA诱导的肺泡损伤，ALI和死亡率。假设是IAV肺感染抑制了AWL分泌，AWL分泌是一种稳态遗传的稳态机制颗粒。抑制作用会导致SA和分泌的SA毒素α溶血素（HLA）的肺泡保留。保留增强了与SA和HLA的肺泡接触，从而促进SA稳定在肺泡壁上， HLA诱导的肺泡液屏障损失，导致肺泡水肿和致命的ALI。除了直接支持指示IAV肺部感染的假设，初步数据引起的：（a）去磷酸化，因此失活肺泡囊性纤维化跨膜电导调节剂（CFTR）CL-通道，临界蛋白锥子分泌；（b）CFTR-脱磷酸蛋白，蛋白磷酸酶2a（PP2A）的甲基化催化亚基（PP2AC），它可能促进PP2AC-CFTR相互作用。具体目标如下。目标1 将定义CFTR在遗传SA的肺泡保留中的作用。 AIM 2将定义PP2AC甲基化的作用在IAV诱导的抑制锥度分泌中。由于我们的初步数据表明CFTR和PP2AC- 甲基化酶，亮氨酸羧基甲基转移酶1（LCMT1）可能代表新的治疗靶标恢复感染IAV的肺中的锥子分泌，AIM 3将测试CFTR-和LCMT1-的治疗潜力针对防止共感染引起的肺泡损伤，ALI和死亡率的靶向方法。这些目标将使用我们已建立的方法来实现，其中包括细胞培养，ALI的小鼠模型和实时活的小鼠和人肺的共聚焦显微镜。 IAV感染的小鼠的测定将包括措施：（1）SA的肺泡保留；（2）锥子分泌物；（3）肺泡CFTR磷酸化状态；（4）肺泡PP2AC甲基化状态；（5）SA-和HLA肺泡上皮损伤和肺泡屏障损失；和（6）SA诱导的肺水肿和死亡率。我们将使用：（i）用肺泡抑制剂治疗的野生型小鼠 PP2AC-CFTR和PP2AC-LCMT1相互作用，包括药物抑制剂，编码突变蛋白的质粒DNA，和sirna; （ii）缺乏肺泡上皮CFTR和LCMT1表达的转基因小鼠。该提议是预计将对IAV破坏临界肺泡功能的分子机制实现新的见解导致致命的Ali，并建立awl分泌的恢复，即“ awl救援” - 作为一种新疗法 IAV-SA共感染引起的ALI方法。因此，该提案解决了NHLBI任务。