Src Regulation of Lung Endothelial Barrier Function

Src 对肺内皮屏障功能的调节

基本信息

批准号：
7843674
负责人：
RICHARD D MINSHALL
金额：
$ 31.35万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7843674
关键词：
Achievement Acute Lung Injury Address Adrenergic Receptor Albumins Aleurites Apical Binding Binding Proteins Biochemical Blood Vessels Caveolae Caveolins Cell membrane Cell surface Cells Characteristics Cholera Toxin Cholera Toxin Protomer B Co-Immunoprecipitations Coupled Data Dissociation Dominant-Negative Mutation Dynamin Dynamin 2 Electron Microscopy Electrons Endocytosis Endothelial Cells Event Fluorescent Probes Funding GTP-Binding Proteins Goals Gold Guanosine Triphosphate Guanosine Triphosphate Phosphohydrolases Heterotrimeric GTP-Binding Proteins Image Immunoblotting In Situ Label Lead Liposomes Liquid substance Lung Measures Mediating Membrane Microscopic Modeling Motion Mus Neck PAR-1 Receptor Pathway interactions Peptides Permeability Pertussis Toxin Phosphorylation Phosphotransferases Physiology Principal Investigator Protein Isoforms Proteins Pulmonary Edema Regulation Retroviral Vector Role Side Signal Pathway Signal Transduction Signaling Molecule Small Interfering RNA Techniques Tertiary Protein Structure Testing Therapeutic Thrombin Thrombin Receptor Toxin Tracer Transfection Tyrosine Tyrosine Phosphorylation Vascular Endothelial Cell Vascular Permeabilities Vesicle base caveolin 1 in vivo inhibitor/antagonist knock-down lung injury macromolecule migration monolayer mutant novel plasmalemmal vesicle programs protein protein interaction receptor coupling research study response scaffold src-Family Kinases trafficking transcytosis uptake

项目摘要

Loss of endothelial barrier function is an important characteristic of Acute Lung Injury (ALl). The transcellular transport of albumin and other macromolecules via endothelial caveolae is a factor contributing to endothelial barrier function. We have identified specific interactions between caveolin-1 (a caveolar protein), the heterotrimeric G protein Gi, and Src kinase in the mechanism of caveolae-mediated endocytosis. The goals of Project 4 are to define the role of caveolin-1 as an organizer and regulator of signal transduction cascades essential for plasmatemmal vesicle trafficking and the protein-protein interactions that regulate albumin permeability via transcytosis. The studies in Project 4 will address the following specific aims. Specific Aim #1: To determine the role of the heterotrimeric G protein, Gi, in signaling ceaveolae-mediated endocytosis and transendothelial albumin permeability in endothelial monolayers; Specific Aim #2: To determine the role of Src activation of the GTPase, dynamin-2, in signaling caveolae-mediated endocytosis and transendothelial albumin permeability; Specific Aim #3:To address the component of thrombin/Protease Activated Receptor-1-induced increase in lung vascular permeability resulting from internalization of caveolae and transcelinlar albumin transport. Thus, Project 4 will identify the receptor-coupled signals activating Src, the phosphorylation targets of Src signaling caveolae fission (specifically, caveolin-1 and dynamin-2), and the role of Src activation in regulating transcellular permeability. To address the in vivo relevance and functional significance of these studies in pulmonary microvascular endothelial cells, experiments will also be made, wherever possible, in intact mouse lung models. Studies will employ approaches in both imaging (i.e., using fluorescent probes and electron microscopic assessment) and physiology (i.e., determination of endothelial permeability in monoayers and mouse lung models) to address the role of caveolae-mediated endocytosis in activating increased albumin permeability. Thus, these studies will elucidate the signaling mechanisms that regulate caveolae internalization and plasmalemmal vesicle trafficking, and thus contribute to the mechanism of transendothelial albumin permeability in lungs. The achievement of these objectives will lead to tthe elucidation of the signals regulating caveolae-mediated endocytosis and its role in contributing to the thrombin-induced increase in lung vascular permeability. With the identification of novel signaling pathways, it may be possible to develop therapeutic strategies that specifically target signals leading to inappropriate increase in lung vascular permeability.

内皮屏障功能的丧失是急性肺损伤的重要特征（ALL）。白蛋白和其他大分子通过内皮小窝的跨细胞运输是导致内皮屏障功能的因素。我们已经在小窝介导的内吞作用机理中鉴定了小窝蛋白-1（小窝蛋白），异三聚体G蛋白GI和SRC激酶之间的特定相互作用。项目4的目标是定义 Caveolin-1是信号转导的组织者和调节剂，这对于浆膜囊泡运输至关重要，以及通过跨胞菌病调节白蛋白渗透率的蛋白质 - 蛋白质相互作用。项目4中的研究将解决以下特定目标。具体目的＃1：确定异三聚体G蛋白GI在信号ceaveolae介导的内吞作用和跨内皮白蛋白渗透性在内皮单层中的作用；特定目的＃2：确定GTPase（Dynamin-2）在信号传导介导的内吞作用和跨内皮白蛋白渗透性中的SRC激活的作用；特定目的＃3：解决凝血酶/蛋白酶激活受体-1诱导的肺血管通透性增加，这是由于小窝和Transcelinlar白蛋白转运的内在化而导致的。因此，项目4将识别激活SRC的受体偶联信号，SRC信号Caveolae裂变的磷酸化靶标（具体是Caveolin-1和 Dynamin-2）以及SRC激活在调节跨细胞渗透性中的作用。为了解决这些研究在肺微血管内皮细胞中的体内相关性和功能意义，在完整的小鼠肺模型中，还将进行实验。研究将在成像（即使用荧光探针和电子显微镜评估）和生理学（即确定内皮通透性的测定）中采用方法。在单层和小鼠肺模型中），以解决小窝介导的内吞作用在激活升高白蛋白渗透性中的作用。因此，这些研究将阐明调节口腔内部化和浆膜囊泡运输的信号传导机制，从而有助于肺中跨内皮白蛋白渗透率的机制。这些目标的实现将导致阐明调节小窝介导的信号内吞作用及其在促进凝血酶引起的肺血管通透性增加中的作用。通过鉴定新的信号通路，可能有可能制定治疗策略，这些治疗策略专门针对信号，从而导致肺血管渗透性不当。