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形成肺泡性水肿。这种屏障的完整性依赖于紧密连接(TJ)的存在，TJ有助于保持细胞相互作用的完整性。因此，它们的破坏可能表现为肺泡水肿和气体交换障碍的发展。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。这些细胞结构是由多种蛋白质组成的，其中包括被认为是紧密连接的骨架的Claudins。低氧对肺泡上皮细胞(AEC)紧密连接的影响尚未阐明。我们建议确定低氧对肺泡上皮屏障的影响，特别是对Claudin蛋白的影响。我们提出了三个相互关联的特定目标：1)确定低氧是否通过将Claudin 3和4从质膜内吞到肺泡上皮细胞的细胞内而破坏肺泡屏障。2)探讨低氧是否通过线粒体活性氧刺激蛋白激酶C(PKC)，从而增加Claudin-3和Claudin-4的磷酸化，促进其内吞作用。3)确定在低氧期间泛素结合系统是否对claudin 3和4的运输和/或降解是必需的。我们期望这些实验产生的数据最终将有助于理解低氧影响肺泡上皮细胞的机制，以及设计新的策略来治疗有可能发生肺泡上皮屏障破坏的患者，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ARDS和充血性心力衰竭等疾病状态下的存活率。

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.