Effects of hypoxia on the alveolar epthelium

缺氧对肺泡上皮的影响

• 批准号: 7435394
• 负责人: Jacob I Sznajder
• 金额: $ 39.92万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-06-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7435394
• 关键词: Acute Lung Injury; Adult Respiratory Distress Syndrome; Affect; Agonist; Altitude; Alveolar; Animal Model; Apical; Back; Blood Circulation; Blood capillaries; Cell membrane; Cells; Cessation of life; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Down-Regulation; Edema; Endocytosis; Epithelial; Epithelial Cells; Event; Exposure to; Floods; Forskolin; Functional disorder; Gases; Hour; Human; Hypercapnia; Hypoxemia; Hypoxia; Impairment; Liquid substance; Lung; Measures; Mediating; Mitochondria; Morbidity - disease rate; Na(+)-K(+)-Exchanging ATPase; Oxygen; Pathogenesis; Pathway interactions; Patients; Phosphorylation; Protein Kinase C; Proteins; Pulmonary Edema; Pump; Reactive Oxygen Species; Recovery; Regulation; Reporting; Role; Signaling Molecule; Terbutaline; Ubiquitination; Water; alveolar epithelium; basolateral membrane; capillary; clinically relevant; design; insight; mortality; multicatalytic endopeptidase complex; novel; novel strategies; prevent; protein function; research study

Patients with acute lung injury develop hypoxemia and gas exchange impairment which results in significant morbidity and mortality. It has been reported that hypoxia may impair the lung's ability to clear edema by inhibiting Na+ channels and the Na,K-ATPase in the alveolar epithelium. The focus of this application is to determine whether severe hypoxia of 1.5% or 3% (about 10 or 20 mm Hg) for 1 to 4 hours inhibits Na,K-ATPase by causing endocytosis of the Na+ pump into intracellular compartments via specific pathways activated by reactive oxygen species (ROS) and protein kinase C (PKC) signaling molecules. We will determine whether after exposure to hypoxia, reoxygenation of the alveolar epithelial cells (AEC) results in the recruitment of previously endocytosed Na+ pumps back into the cell basolateral membranes (BLM). We will study whether activation of the protein kinase A pathway by terbutaline and forskolin reverses the effects of hypoxia on AEC Na+ pumps and whether this increases lung liquid clearance. We will also determine whether in AEC exposed to prolonged hypoxia (>12 hours) the Na,K-ATPase proteins undergo ubiquitination and degradation via the proteasomal or lysosomal pathways. As such, we will study the effects of hypoxia on the alveolar epithelium by focusing on the mechanisms of Na,K-ATPase regulation in four interrelated aims: in Specific Aim # 1 we propose to determine the role of mitochondrial ROS generated during hypoxia on Na,K-ATPase function and protein endocytosis; in Specific Aim # 2 we will study whether the alveolar epithelial Na,K-ATPase is phosphorylated during hypoxia by PKC triggering the endocytosis of the Na+ pump; in Specific Aim # 3 we will determine whether hypoxia-mediated inhibition of the Na,K-ATPase function and endocytosis of the Na+ pump are reversible upon reoxygenation and whether activation of the PKA pathway by terbutaline and forskolin prevents or reverses the effects of hypoxia; in Specific Aim # 4 we propose to determine whether prolonged exposure of AEC to hypoxia leads to ubiquitination and degradation of the Na,K-ATPase proteins via the proteosomal/lysosomal pathways. Experiments have been conducted for each of the specific aims and the preliminary results support the feasibility of this proposal. Completion of the proposed studies will provide novel information on the effects of hypoxia on the alveolar epithelium, specifically as it pertains to mechanisms of inhibition and degradation of the Na+ pump as well as pathways of reversal of Na,K-ATPase inhibition, which may be of importance for the design of novel approaches to increase edema clearance in patients with pulmonary edema.

急性肺损伤患者会出现低氧血症和气体交换障碍，导致显着的发病率和死亡率。据报道，缺氧可能通过抑制肺泡上皮细胞中的 Na+ 通道和 Na,K-ATP 酶来损害肺清除水肿的能力。本应用的重点是确定 1.5% 或 3%（约 10 或 20 mm Hg）的严重缺氧持续 1 至 4 小时是否会通过激活的特定途径导致 Na+ 泵内吞到细胞内区室中来抑制 Na,K-ATP 酶通过活性氧 (ROS) 和蛋白激酶 C (PKC) 信号分子。我们将确定暴露于缺氧后，肺泡上皮细胞 (AEC) 的复氧是否会导致先前内吞的 Na+ 泵重新招募回细胞基底外侧膜 (BLM)。我们将研究特布他林和毛喉素激活蛋白激酶 A 通路是否可以逆转缺氧对 AEC Na+ 泵的影响，以及这是否会增加肺液体清除率。我们还将确定 AEC 中是否暴露于长时间缺氧（>12 小时）中 Na,K-ATP 酶 蛋白质通过蛋白酶体或溶酶体途径进行泛素化和降解。因此，我们将通过在四个相互关联的目标中关注 Na,K-ATP 酶调节机制来研究缺氧对肺泡上皮的影响：在具体目标#1 中，我们建议确定缺氧期间产生的线粒体 ROS 对 Na 的作用。 ,K-ATP酶功能和蛋白质内吞作用；在具体目标#2中，我们将研究肺泡上皮Na,K-ATP酶在缺氧期间是否被PKC磷酸化，从而触发Na+泵的内吞作用；在具体目标#3中，我们将确定缺氧介导的Na,K-ATP酶功能和Na+泵内吞作用的抑制在复氧后是否可逆，以及特布他林和毛喉素激活PKA途径是否可以预防或逆转缺氧的影响；在具体目标#4中，我们建议确定AEC长时间暴露于缺氧是否会通过蛋白体/溶酶体途径导致Na,K-ATP酶蛋白泛素化和降解。针对每个具体目标都进行了实验，初步结果支持了该提案的可行性。拟议研究的完成将为缺氧对肺泡上皮的影响提供新的信息，特别是因为它涉及Na +泵的抑制和降解机制以及Na，K-ATP酶抑制的逆转途径，这可能是对于设计增加肺水肿患者水肿清除的新方法具有重要意义。