Hypoxia Disrupts the Alveolar Barrier: Role of Claudins

描述（由申请人提供）： 肺泡上皮屏障需要被破坏以发展肺泡水肿。这种屏障的完整性取决于紧密连接（TJ）的存在;这有助于保持细胞相互作用的完整性。因此，它们的破坏可以表现为肺泡水肿和不稳定气体交换的发展。在肺损伤和肺水肿期间，肺泡处存在低氧环境，这可能有助于进一步破坏肺泡上皮屏障，特别是紧密连接。这些细胞结构由多种蛋白质形成，其中的claudins被认为是紧密连接的骨架。缺氧对肺泡上皮细胞紧密连接的影响尚未阐明。我们建议确定缺氧对肺泡上皮屏障的影响，特别是对claudin蛋白。我们提出了三个相互关联的具体目标：1）确定缺氧是否通过将密蛋白3和4从质膜内吞到肺泡上皮细胞的细胞内区室中而破坏肺泡屏障。2)确定缺氧是否通过线粒体活性氧刺激蛋白激酶C（PKC），从而增加密蛋白3和4的磷酸化并促进其内吞作用。3)为了确定在缺氧期间，泛素缀合系统是否是密蛋白3和4的运输和/或降解所必需的。我们期望从这些实验产生的数据将最终有助于理解肺泡上皮受缺氧影响的机制，并设计新的策略来治疗处于肺泡上皮屏障破坏风险中的患者，例如在ARDS中本研究与公共卫生的相关性：肺泡上皮屏障的破坏导致肺水肿。了解如何阻止其中断可能会提高疾病状态（如ARDS和充血性心力衰竭）的生存率。

DESCRIPTION (provided by applicant): The alveolar epithelial barrier needs to be disrupted for developing alveolar edema. The integrity of this barrier is dependent on the presence of tight junctions (TJs); which contribute to keep cell interactions intact. Accordingly their disruption could manifest in the development of alveolar edema and impared gas exchange. During lung injury and pulmonary edema a hypoxic environment is present at the alveolus which could contribute to further disruption of the alveolar epithalial barrier, specifically the tight junctions. These cell structures are formed by multiple proteins, among them the claudins which are considered the backbones of the tight junctions. The effect of hypoxia on alveolar epithelial cells (AECs) tight junctions have not been elucidated. We propose to determine the effect of hypoxia on the alveolar epithelial barrier specifically on the claudin proteins. We propose three inter-related specific aims: 1) to determine whether hypoxia disrupts the alveolar barrier via endocytosis of claudin 3 and 4 from the plasma membrane into intracellular compartments of alveolar epithelial cells. 2) To determine whether hypoxia, via mitochondrial reactive oxygen species, stimulates Protein Kinase C (PKC), thus increasing the phosphorylation of claudin- 3 and 4 and promoting their endocytosis. 3) To determine whether during hypoxia the ubiquitin-conjugating system is necessary for the trafficking and /or degradation of claudin 3 and 4.We expect that the data generated from these experiments will ultimately help with the understanding of mechanisms by which the alveolar epithelium is affected by hypoxia and the design of novel strategies to treat patients who are at risk to develop disruption of the alveolar epithelial barrier, such as in ARDS Relevance of this research to public health: Disruption of the alveolar epithelium barrier causes pulmonary edema. Understanding how to block its disruption can potentially improve survival in disease states such as ARDS and congestive heart failure.