Alcohol increases tight junction permeability & disrupts translocation of ZO-1

描述(申请人提供)：酒精是世界上最常见的滥用药物。在美国，大约一半的成年人经常饮酒。众所周知，酒精对肺部有害。例如，大量饮酒会增加患肺炎、支气管炎和急性呼吸窘迫综合征(ARDS)等肺部疾病的风险。传导呼吸道的第一道防线是呼吸道上皮细胞，它含有紧密连接并表达Zonula occluden-1和occludin。Claudin-1是一种跨膜蛋白，在调节紧密连接通透性中起重要作用。ZO-1是一种外周相关的膜蛋白，连接紧密连接蛋白与肌动蛋白细胞骨架和其他紧密连接蛋白(claudin和occludin)，被认为是稳定紧密连接的重要组成部分。紧密连接位于呼吸道上皮细胞的顶端-侧缘，在那里它们调节细胞的极性，并选择性地调节水、离子、中性分子和炎症细胞通过细胞旁途径的通道。多项研究表明，酒精破坏紧密连接，导致屏障“渗漏”。在肺中，这种情况发生在实质间隔内，导致肺水肿和肺泡水肿。紧密连接在调节传导呼吸道的屏障功能方面也起着关键作用。呼吸道“漏气”是支气管炎和哮喘等呼吸道疾病的基本特征。渗漏增加会导致呼吸道浮肿，这是这些疾病的典型表现。然而，人们对酒精在传导呼吸道中的影响知之甚少。我们最近观察到，酒精损害ZO-1和claudin-1到细胞膜的易位，并增加紧密连接的通透性，导致更多的呼吸道上皮细胞渗漏。我们的数据将这种“泄漏”与酒精诱导的呼吸道蛋白激酶C(PKC)激活的变化联系在一起，导致蛋白质转移到细胞膜的受损。这些发现使我们假设：酒精促进了与ZO-1和Claudin-1在呼吸道上皮细胞中依赖的PKC途径的非局部化有关的呼吸道渗漏。我们建议通过3个具体目标来验证这一假说：1)确定酒精对传导呼吸道上皮紧密连接的功能后果；2)确定酒精损害ZO-1和易位的机制；3)在酒精喂养的小鼠体内模型中建立酒精触发的ZO-1和重排的影响。这些研究的结果将提高我们对酒精对呼吸道上皮屏障功能的影响的理解，这对于理解和治疗酒精滥用者中常见的呼吸道疾病至关重要。Claudin-1(claudin-1) 与公共健康相关：这些创新研究将展示酒精如何调节呼吸道上皮细胞紧密连接的渗透性，并提供有关酒精如何改变呼吸道生理的有价值的信息。)

DESCRIPTION (provided by applicant): Alcohol is the most commonly abused drug in the world. In the United States about half the adult population regularly consumes alcohol. It has been well established that alcohol has harmful effects on the lung. For example, heavy alcohol intake increases the risk for developing pulmonary diseases such as pneumonia, bronchitis and acute respiratory distress syndrome (ARDS). The first line of defense of the conducting airways is the airway epithelium, which contains tight junctions and expresses Zonula occluden-1 (ZO-1), and occludin. Claudin-1 is a transmembrane protein, which is important in regulating tight junction permeability claudin-1, . ZO-1 is a peripherally associated membrane protein, which links tight junction proteins to the actin cytoskeleton, other tight junction proteins (claudin and occludin) and is believed to be an important component in stabilizing tight junctions. Tight junctions are characteristically located at the apical-lateral borders in airway epithelium where they regulate cell polarity and act to selectively regulate the passage of water, ions, neutral molecules and inflammatory cells through the paracellular pathway. A number of studies have shown that alcohol disrupts tight junctions resulting in barrier "leak". In the lung this occurs in the parenchyma compartment resulting in pulmonary edema and alveolar edema. Tight junctions also play a key role in regulating the barrier function in the conducting airways. Airway "leak" is a cardinal feature of airway diseases such as bronchitis and asthma. Increased leak contributes to airway edema, which is a classic finding of these diseases. However, little is known about the effects of alcohol in the conducting airways. We have recently observed that alcohol impairs ZO-1 and claudin-1 translocation to the cell membrane and increases tight junction permeability resulting in "leakier" airway epithelial cells. Our data link this "leakiness" to alcohol-induced changes in airway protein kinase C (PKC) activation resulting in impaired protein translocation to the cell membrane. These findings led us to hypothesize that: Alcohol promotes airway leak linked to the delocalization of ZO-1 and claudin-1 through a PKC-dependent pathway in the airway epithelium. We propose to test this hypothesis with 3 specific aims: 1) Determine the functional consequences of alcohol on tight junctions in the epithelium of the conducting airways; 2) Define the mechanism through which alcohol impairs ZO-1 and translocation; 3) Establish the impact of alcohol-triggered ZO-1 and rearrangement in an in vivo model of alcohol-fed mice. Results for these studies will improve our understanding of the impact alcohol has on airway epithelial barrier functions, which are critical for the understanding and treatment of airway diseases common among alcohol abusers. claudin-1 claudin-1 ) PUBLIC HEALTH RELEVANCE: These innovative studies will demonstrate how alcohol modulates tight junction permeability in airway epithelium and provide valuable information on how alcohol alters airway physiology. )