Ethanol-mediated cilia motility dysfunction

描述（由申请人提供）：已知大量酒精摄入会损害肺保护自身免受肺炎和支气管炎等感染的能力。肺抵抗这种感染的第一道防线是捕获吸入的颗粒并使用粘膜纤毛装置将它们推出肺。在一个健康的个体中，这个装置产生粘液，粘液捕获吸入的颗粒，然后被纤毛（肺衬里的指状突起）推出肺外。纤毛受损会导致肺炎和肺功能丧失。我们已经确定，酒精通过改变称为激酶的关键蛋白质，严重损害了这种微妙的粘膜纤毛装置。这些激酶中的两种，PKA和PKG，是被充分描述的调节包括肺纤毛的细胞功能的分子。我们在酒精喂养的小鼠中的实验表明，酒精深刻地损害了这些激酶的功能，导致粘膜纤毛装置以需要产生一氧化氮的方式失去正常的“战斗或逃跑”反应。虽然我们已经确定酒精会导致这个问题，但我们不知道如果去除酒精，它会持续多久。我们也不知道酒精是如何改变纤毛中这些重要的激酶依赖性功能的。这使我们假设：慢性酒精暴露导致时间依赖性和可逆性损害的粘膜纤毛功能改变特定的调节蛋白在气道上皮纤毛。在这个提议中，我们提出了旨在回答两个问题的实验：1。酒精引起的黏膜纤毛功能损害是可预防的还是可逆的？和2.哪些纤毛蛋白会受到酒精的影响？我们建议通过四个具体目标提出的实验来回答这些问题：1。描述慢性酒精损害粘膜纤毛功能的时间过程和可逆性; 2.明确氧化应激在酒精介导的一氧化氮依赖性纤毛运动调节中的信号通路和作用。确定存在于被酒精暴露改变的纤毛上的激酶磷酸化靶点;和4.使用分离的纤毛模型鉴定酒精介导的纤毛损伤的分子决定因素。虽然我们已经了解了酒精对粘膜纤毛功能的影响和机制，但我们提出的研究将扩展我们的知识，了解如何预防或治疗这种情况，以及酒精如何改变对纤毛功能和肺部健康至关重要的分子。

DESCRIPTION (provided by applicant): Heavy alcohol intake is known to damage the ability of the lung to protect itself from infections such as pneumonia and bronchitis. The lung's first line of defense against such infection is to trap inhaled particles and propel them out of the lung using the mucociliary apparatus. In a healthy individual, this apparatus produces mucus, which traps inhaled particles, and is then propelled out of the lung by cilia, the fingerlike projections of the lung lining. Damage to cilia results in pneumonia and loss of lung function. We have established that alcohol profoundly injures this delicate mucociliary apparatus by altering critical proteins called kinases. Two of these kinases, PKA and PKG, are well-described molecules that regulate cell functions including lung cilia. Our experiments in alcohol-fed mice have demonstrated that alcohol profoundly impairs the function of these kinases, resulting in a loss of the normal "fight or flight" responsiveness of the mucociliary apparatus in a manner that requires the production of nitric oxide. While we have established that alcohol causes this problem, we do not know how long it persists if alcohol is removed. We also do not know exactly how alcohol alters these important kinase-dependent functions in cilia. This leads us to hypothesize that: Chronic alcohol exposure causes time-dependent and reversible impairment of mucociliary function by altering specific regulatory proteins in airway epithelial cilia. In this proposal, we propose experiments designed to answer two questions: 1. Is alcohol-driven impairment of mucociliary function preventable or reversible? and 2. Which cilia proteins are affected by alcohol? We propose to answer these questions through experiments proposed in four specific aims: 1. Characterize the time course and reversibility of chronic alcohol impairment of mucociliary function; 2. Define the signaling pathways and role that oxidant stress plays in alcohol-mediated nitric oxide-dependent regulation of ciliary motility, 3. Determine the kinase phosphorylation targets present on cilia that are altered by alcohol exposure; and 4. Identify the molecular determinants of alcohol-mediated cilia injury using isolated cilia models. Although we have learned much about the impact and mechanisms of alcohol injury to mucociliary function, the studies we propose will extend our knowledge into how this may be prevented or treated and exactly how alcohol alters molecules critical for cilia function and lung health.