Mucociliary Function in Chronic Bronchitis

描述（由申请方提供）：在慢性支气管炎中，粘膜纤毛清除功能障碍，尤其是在疾病加重期间。慢性支气管炎急性发作通常与活性氧（ROS）产生增加和气道酸化有关。本项目将研究ROS和细胞内酸化如何调节和/或失调粘液纤毛功能。至少有两种内源性和旁分泌介质控制粘膜纤毛转运系统的关键成分：细胞外ATP和透明质酸片段。我们提出，pannexins，蛋白质相关，但不同的连接蛋白，形成通道的细胞外（称为pannexons），并负责，至少部分，顶端ATP在气道上皮细胞的释放。我们假设在疾病恶化的早期，ROS的产生不仅增加细胞内钙浓度（[Ca 2 +]i）以刺激纤毛搏动频率（CBF）和通过pannexons的顶端ATP释放，而且还降解顶端透明质酸，这反过来刺激CBF并增加气道表面液体（ASL）体积。随后的细胞内酸化，可能是由尼古丁介导的气道中表达的NADPH氧化酶活性上调（Duox）介导的，通过抑制纤毛活性和泛连接蛋白导致粘膜纤毛功能障碍。将以三个具体目标来检验这一假设。具体目标1将检验以下假设：外源性ROS激活信号级联，包括初始[Ca 2 +]i增加以刺激CBF，尽管Duox产生H+导致轻度和暂时的细胞内酸化;此外，直接顶端透明质酸降解激活RHAMM和罗恩以刺激CBF并可能增加ASL体积。具体目标2将检验以下假设：泛连接蛋白负责将ATP释放到气道上皮细胞的顶面，其中ATP在调节包括CBF和ASL体积的粘膜纤毛功能中起重要作用。具体目标3将检验以下假设：持续性细胞内酸化通过直接作用于纤毛搏动以及通过防止ATP通过pannexons释放来抑制粘膜纤毛清除。所提出的实验的结果，使用新的和国家的最先进的方法，将提供新的和重要的机制的见解慢性支气管炎的粘液纤毛清除的调节。

DESCRIPTION (provided by applicant): In chronic bronchitis, mucociliary clearance is dysfunctional, especially during disease exacerbation. Chronic bronchitis exacerbations are commonly associated with increased production of reactive oxygen species (ROS) and airway acidification. This project will examine how ROS and intracellular acidification may regulate and/or dysregulate mucociliary function. At least two endogenous and paracrine mediators control key components of the mucociliary transport system: extracellular ATP and hyaluronan fragments. We propose that pannexins, proteins related to but different from connexins, form channels to the outside of cells (called pannexons) and are responsible, at least in part, for the release of apical ATP in the airway epithelium. We hypothesize that early in disease exacerbation, the production of ROS not only increases intracellular calcium concentrations ([Ca2+]i) to stimulate ciliary beat frequency (CBF) and apical ATP release through pannexons but also degrades apical hyaluronan which in turn stimulates CBF and increases airway surface liquid (ASL) volume. Subsequent intracellular acidification, possibly mediated by cytokine-mediated upregulation of an NADPH oxidase activity expressed in the airway (Duox), leads to mucociliary dysfunction by inhibiting ciliary activity and pannexins. This hypothesis will be tested with three specific aims. Specific Aim 1 will test the hypothesis that exogenous ROS activate a signaling cascade that includes an initial [Ca2+]i increase to stimulate CBF, despite a mild and temporary intracellular acidification due to H+ production by Duox; in addition, direct apical hyaluronan degradation activates RHAMM and RON to stimulate CBF and possibly increase ASL volume. Specific Aim 2 will test the hypothesis that pannexins are responsible for releasing ATP to the apical surface of airway epithelial cells where ATP plays an important role in regulating mucociliary functions including CBF and ASL volume. Specific Aim 3 will test the hypothesis that persistent intracellular acidification inhibits mucociliary clearance by a direct action on ciliary beating as well as by preventing ATP release through pannexons. The results of the proposed experiments, using novel and state-of-the-art methods, will provide new and important mechanistic insights into the regulation of mucociliary clearance in chronic bronchitis.