Redox regulation of alveolar fluid balance

肺泡液平衡的氧化还原调节

• 批准号: 7450433
• 负责人: My N. Helms
• 金额: $ 8.91万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7450433
• 关键词: AGTR2 gene; Alveolar; Alveolar Cell; Alveolus; Amiloride; Apical; Binding; Biological Assay; Biological Models; Breathing; Cell Death; Cells; Complex; Condition; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Data; Environment; Epithelial Cells; Epithelium; Equilibrium; Fatigue; Fluid Balance; Gases; Genes; Goals; Homeostasis; Ion Channel; Ion Transport; Lead; Length; Life; Liquid substance; Lung; Measurement; Measures; Mentors; Methods; Modeling; Molecular; Mus; Nitric Oxide; Oxidation-Reduction; Oxygen; Oxygen measurement, partial pressure, arterial; Peroxonitrite; Phase; Physiological; Play; Preparation; Production; Property; Protocols documentation; Rattus; Reactive Nitrogen Species; Reactive Oxygen Species; Regulation; Research; Research Design; Research Personnel; Respiratory physiology; Role; Signal Transduction; Signal Transduction Pathway; Signaling Molecule; Slice; Small Interfering RNA; Sodium Chloride; Superoxide Dismutase; Superoxides; Surface; Techniques; Testing; Time; Tissues; Transgenic Mice; Water; Work; alveolar epithelium; career; cell type; clinically relevant; epithelial Na+ channel; epithelial amiloride-sensitive sodium channel; human NOS2A protein; in vivo; in vivo Model; inhibitor/antagonist; insight; novel strategies; patch clamp; pneumocyte; programs

DESCRIPTION (provided by applicant): The alveolar epithelium in normal lungs is comprised of two morphologically distinct types of cells (type 1 and type 2) that are responsible for maintaining lung fluid balance. Tight regulation of alveolar fluid clearance is essential for maintaining a dry breathing space, and hence, proper gas exchange. It has been established that net ion transport through amiloride-sensitive epithelial sodium channels (ENaC), located on the apical surface of alveolar epithelial cells, play a critical role in fluid clearance in normal lung. However, the specific mechanisms regulating ENaC function are not completely understood. Within the alveoli, complex regulatory mechanisms must also be in place to balance the redox state of type 1 and type 2 cells, since inspired oxygen is converted to superoxide anions (O2-). Excessive O2- production, caused by high oxygen tensions, can lead to tissue damage and cell death, whereas insufficient oxygenation can result in anything from fatigue to life threatening conditions. In vivo, superoxides react quickly and irreversibly with nitric oxide (NO) to form peroxynitrite. We hypothesize that endogenous 02- binding to NO limits nitric oxide inhibition of ENaC function, thereby enhancing alveolar fluid clearance. Indeed, we have preliminary data suggesting that nitric oxide-unresponsive AT1 cells may have elevated levels of O2-, and that increasing O2- enhances Na transport. To investigate the role of redox signaling in alveolar fluid clearance, and more specifically, ENaC function, I will utilize single channel patch clamp analysis to examine ion transport in lung slice preparations, bio-molecular techniques to examine redox signaling in pneumocytes, and perform whole lung studies in vivo. My first aim, performed during the mentored phase, directly examines the role of O2- in alveolar fluid clearance. Successful completion of this aim will naturally transition into aims 2 and 3, which utilizes several protocols that will be established in the mentored phase, as well as incorporate new approaches to studying type 1 and type 2 cells. The second aim will determine the role of NO in lung function, and lastly, the third aim examines the putative reciprocal relationship between O2- and NO regulation of lung fluid balance. The studies proposed have real clinical relevance, and the potential for a very productive independent research career.

描述（由申请人提供）： 正常肺中的肺泡上皮由两种形态上不同类型的细胞（1型和2型）组成，这些细胞负责维持肺液平衡。严格调节肺泡液清除对于维持干呼吸空间以及适当的气体交换至关重要。已经确定，位于肺泡上皮细胞的顶部表面上，通过艾米洛德敏感性上皮钠通道（ENAC）的净离子转运在正常肺中的液体清除率中起着关键作用。但是，调节ENAC功能的特定机制尚未完全理解。在肺泡中，还必须建立复杂的调节机制，以平衡1型和2型细胞的氧化还原状态，因为启发的氧气被转化为超氧化物阴离子（O2-）。高氧张力引起的过多的O2产生会导致组织损伤和细胞死亡，而氧合不足可能导致从疲劳到危及生命的条件。在体内，超氧化物与一氧化氮（NO）形成过氧亚硝酸盐的一氧化氮（NO）反应。我们假设内源性02-结合无极限的ENAC功能抑制一氧化氮，从而增强了肺泡流体清除率。实际上，我们有初步数据表明一氧化氮无反压AT1细胞可能具有O2-的水平升高，并且增加O2-可以增强Na转运。为了研究氧化还原信号在肺泡流体清除率中的作用，更具体地说，ENAC功能，我将利用单通道贴片夹分析来检查肺切片制剂中的离子传输，生物分子技术以检查肺细胞中的氧化还原信号传导，并在体内进行全肺研究。我在指导阶段执行的第一个目标直接检查了O2-在肺泡流体清除率中的作用。该目标的成功完成将自然过渡到目标2和3，该目标利用了将在指导阶段建立的几种方案，并结合了研究1型和2型细胞的新方法。第二个目的将决定NO在肺功能中的作用，最后，第三个目标研究了O2-和NO肺液平衡调节之间的假定相互关系。拟议的研究具有真正的临床意义，并且具有非常有生产力的独立研究生涯的潜力。