Mitigating Acute Lung Injury by Cell-specific Targeting of MTOR

通过细胞特异性靶向 MTOR 减轻急性肺损伤

• 批准号: 10056811
• 负责人: David A Dean
• 金额: $ 58.94万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-15 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10056811
• 关键词: Acute; Acute Lung Injury; Address; Adult Respiratory Distress Syndrome; Alveolar Cell; Animal Model; Anti-Inflammatory Agents; Blood Vessels; Bronchopulmonary Dysplasia; CMV promoter; Case Study; Cell Nucleus; Cells; Cellular Metabolic Process; Cessation of life; Chronic Obstructive Airway Disease; Clinical; Complementary DNA; Critical Illness; DNA; DNA Polymerase III; Data; Development; Disease; Drug Targeting; Effectiveness; Electroporation; Endothelial Cells; Endothelium; Epithelial; Epithelial Cells; Epithelium; FRAP1 gene; Functional disorder; Gases; Gene Delivery; Gene Expression; Gene Transfer; Genes; In Vitro; Incidence; Infiltration; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Injury; Knock-out; Knockout Mice; Lead; Length of Stay; Lung; Lung Inflammation; Mediating; Modeling; Monitor; Mus; Nuclear; Outcome; Pathogenesis; Patients; Phenotype; Plasmids; Preventive; Public Health; Publishing; Pulmonary Edema; Reporting; Resolution; Respiratory Failure; Role; Sepsis; Signal Transduction; Sirolimus; Structure; Supportive care; Testing; Therapeutic; Treatment Efficacy; Tumor Angiogenesis; United States; Work; alveolar epithelium; base; cDNA Expression; cecal ligation puncture; cell growth; cell injury; cell type; design; effective therapy; effectiveness evaluation; experimental study; gene therapy; improved; improved outcome; innovation; insight; knock-down; lung development; lung injury; mortality; mouse model; novel; novel strategies; novel therapeutic intervention; prevent; promoter; response; small hairpin RNA; transgene expression; treatment strategy; uptake; vascular injury

Acute lung Injury (ALI) is a common cause of respiratory failure in critically ill patients. It has an incidence of ~ 200,000 cases each year in the United States alone and is associated with an unacceptably high mortality rate of 25-40% and 3.6 million hospital days in reported cases. Although, our understanding of the mechanisms relevant to the pathogenesis and the resolution of ALI and Acute Respiratory Distress Syndrome (ARDS) has increased during the past four decades, all current therapies for ALI/ARDS still rely on supportive care and no effective therapeutic options are available to improve clinical outcome. Thus, the development of new treatment strategies for ALI/ARDS that are safe, effective, and based on deeper understanding of the mechanisms involved in ALI pathogenesis is warranted. This proposal aims to clarify the cell type-specific role of MTOR (mechanistic [formerly mammalian] target of rapamycin) in ALI and test the utility of simultaneous but differential cell-specific targeting of MTOR to control ALI. The proposal is based on our published and on-going work that implicates a cell type-specific role for MTOR in inflammation; it mediates inflammation in epithelial cells whereas it serves to limit endothelial cell inflammation. Intriguingly, however, the “net effect” of MTOR signaling results in a proinflammatory phenotype in the lung. The proposal will address the following three inter-related, but independent, aims. Aim 1 will test the hypothesis that MTOR limits ALI by serving an anti-inflammatory function in pulmonary endothelium. Studies in this aim will ascertain the role of endothelial MTOR in moderating ALI by determining the effects of modulating MTOR signaling in pulmonary endothelium on lung inflammation and injury. Aim 2 will investigate the possibility that MTOR promotes ALI by exerting a proinflammatory function in alveolar epithelium. Aim 2 studies will determine the role of epithelial MTOR in augmenting ALI by monitoring the effects of modulating MTOR signaling in alveolar epithelium on lung inflammation and injury. Aim 3 will test the hypothesis that targeting MTOR simultaneously but differentially in a cell type-specific manner (increasing it in pulmonary endothelium but decreasing it in alveolar epithelium) will yield a superior protective and therapeutic benefit against ALI. The proposed studies will be carried out using established mouse models of ALI and will utilize a very new and exciting approach that uses unique cell-specific DNA nuclear targeting sequences (DTSs) in the plasmid to direct cell-specific plasmid nuclear uptake and gene (shRNA or cDNA) expression in the desired cell type. The plasmids will be delivered into the lungs of mice via electroporation, which yields high level of gene expression without inducing inflammation or any cell damage to the epithelial or endothelial cell layer. The creative use of cell-specific DTS carrying plasmid and electroporation will provide valuable insight into the cell- specific role of MTOR in ALI and the basis for novel therapeutic approaches involving cell-specific modulation of MTOR to control ALI.

急性肺损伤（ALI）是危重患者呼吸衰竭的常见原因。它的发病率为~ 仅在美国，每年就有20万例，死亡率高得令人无法接受 25-40%和360万个住院日。尽管我们对这些机制的理解 与ALI和急性呼吸窘迫综合征（ARDS）的发病机制和解决方案相关的 在过去的四十年中，所有目前的ALI/ARDS治疗仍然依赖于支持性护理， 有效的治疗选择可用于改善临床结果。因此，开发新的治疗方法 安全、有效、基于对相关机制更深入理解的ALI/ARDS治疗策略 在ALI发病机制中的作用。该建议旨在阐明MTOR的细胞类型特异性作用（机制 [原哺乳动物]雷帕霉素的靶点）在ALI中的作用，并测试同时但不同的细胞特异性 以MTOR为目标控制ALI。该提案是基于我们已发表和正在进行的工作，其中涉及 MTOR在炎症中的细胞类型特异性作用;它介导上皮细胞中的炎症， 限制内皮细胞炎症。然而，有趣的是，MTOR信号传导的“净效应”导致 肺中的促炎表型。该提案将解决以下三个相互关联的问题，但 独立，目标。目的1将检验MTOR通过提供抗炎功能限制ALI的假设 在肺内皮中。这方面的研究将通过以下方法确定内皮MTOR在减轻ALI中的作用： 确定调节肺内皮中的MTOR信号传导对肺炎症和损伤的影响。 目的2探讨MTOR是否通过在肺泡内发挥促炎作用而促进ALI的发生 上皮目的2研究通过监测上皮细胞MTOR在加重ALI中的作用， 调节肺泡上皮细胞中的MTOR信号传导对肺炎症和损伤的影响。目标3将测试 假设同时但以细胞类型特异性方式差异地靶向MTOR（在细胞中增加它）， 肺内皮细胞，但减少肺泡上皮细胞）将产生优异的上级保护和治疗 对ALI有好处。拟议的研究将使用已建立的ALI小鼠模型进行， 利用一种非常新的和令人兴奋的方法，使用独特的细胞特异性DNA核靶向序列（DTS） 在质粒中引导细胞特异性质粒核摄取和基因（shRNA或cDNA）在所需细胞中的表达。 细胞类型。质粒将通过电穿孔被递送到小鼠的肺中，其产生高水平的基因。 表达而不诱导炎症或对上皮或内皮细胞层的任何细胞损伤。的 创造性地使用携带质粒的细胞特异性引物和电穿孔将提供对细胞的有价值的了解， MTOR在ALI中的特定作用以及涉及细胞特异性调节MTOR的新治疗方法的基础。 MTOR控制ALI。