Regulation of alveolar barrier function by claudins

描述（由申请人提供）：急性呼吸窘迫综合征（ARDS）是一种毁灭性的呼吸衰竭综合征，在美国每年导致近75,000人死亡。急性肺损伤的治疗主要是支持性的，目前还没有药物治疗。急性肺损伤的病理生理特征是炎症和肺泡-毛细血管屏障的破坏。急性肺损伤的恢复需要肺泡上皮屏障的重建，包括上皮细胞间低通透性紧密连接的形成。为了促进这一领域的发展，需要对肺泡上皮屏障的调节和修复有更全面的了解。克劳丁蛋白是屏障功能的核心。claudin是紧密连接形成所必需的跨膜蛋白，但可能参与其他重要的细胞修复过程。因此，我们研究了claudin在肺泡上皮中的功能和调控。我们的工作表明，肺泡上皮中两个最丰富表达的claudin在肺损伤期间被选择性调节。本提案将探讨其具体功能

DESCRIPTION (provided by applicant): Acute respiratory distress syndrome (ARDS), is a devastating syndrome of respiratory failure that contributes to nearly 75,000 deaths annually in the United States. Therapy for acute lung injury is primarily supportive and currently no pharmacologic therapies are available. The pathophysiology of acute lung injury is characterized by inflammation and disruption of alveolar-capillary barrier. Recovery from acute lung injury requires re- establishment of the alveolar epithelial barrier, including the formation f low-permeability tight junctions between epithelial cells. To advance the field, a more complete understanding of alveolar epithelial barrier regulation and repair is needed. Claudin proteins are central to barrier function. Claudins are transmembrane proteins that are required for tight junction formation, but likely participate in other cellular processes important to repair. Therefoe we have investigated the functions and regulation of claudins in the alveolar epithelium. Our work has shown that two of the most abundantly expressed claudins in the alveolar epithelium are selectively regulated during lung injury. This proposal will investigate the specific functions and regulation of claudins-18 and -4 in mechanistic detail using an integrative approach that includes experimental models of lung injury, primary cell culture, and studies in ex vivo perfused human lungs. In Aim 1 we will determine the unique functions of lung-specific claudin-18.1, and the mechanisms for its selective down regulation in acute lung injury. Will we test the hypothesis that claudin-18 is required for the macromolecule permeability barrier in the lung and that the loss of claudin-18 contributes to more severe lung injury. We propose that claudin-18 forms homotypic tight junction strands that uniquely limit macromolecule permeability. During injury, TNF-¿ mediates the selective loss of claudin-18 from intact tight junctions, in part through effects on the cytoskeleton. In Aim 2 we will follow up on our previous finding that claudin-4 is consistently induced in acute lung injury. We will determine if claudin-4 is required for EGFR-mediated epithelial repair in lung injury. We hypothesize that claudin-4 is induced by EGFR in response to epithelial cell injury and the loss of cell-cell contact. We propose claudin-4 acts to accelerate repair through a previously unknown interaction with active Rap1 that speeds tight junction sealing and promotes cell spreading. Preliminary data show that claudin-4 is consistently induced in several models of lung injury and in human lungs in association with more preserved barrier function. Claudin- 4 is required for repair and preferentially associates with active Rap1. In Aim 3 we will determine the clinical significance of differences in claudin expression to epithelial barrier function using the ex vivo perfused human lung model. We hypothesize that claudin-18 is the predominant claudin in human type 1 cells and the specific loss of claudin-18 results in increased alveolar epithelial permeability in the ex vivo model and greater lung injury in donors. We will investigate the mechanism for the regulation of claudin-18 and claudin-4 using a bacterial pneumonia model in the ex vivo system. These studies will provide novel insights into the contributions of claudin proteins to the loss and recovery of alveolar epithelial barrier function in acute lung injury.