The injurious effects of hypercapnia on the alveolar epithelium

描述（由申请方提供）：COPD患者和“允许性高碳酸血症”机械通气患者的pCO 2可能显著升高。我们推测高碳酸血症通过特异性下调肺泡上皮Na，K-ATP酶和降低肺泡液体清除率导致肺泡上皮功能障碍。本申请的重点是确定短期（30-60分钟）高碳酸血症是否通过抑制Na，K-ATP酶并促进其经由涉及JNK和ERK激酶和蛋白激酶C（PKC）信号传导分子的特定途径从质膜内吞到细胞内区室而可逆地损害肺泡液体重吸收。我们还建议确定长期（5和7天）高碳酸血症是否会使肺泡上皮对通气诱导的肺损伤敏感，并通过泛素/蛋白体途径不仅引起内吞作用，还引起Na，K-ATP酶降解。感知高碳酸血症的细胞信号在很大程度上是未知的，因此我们将利用C。Elegans已经完全映射，并在C.研究可能参与高pCO 2传感和反应的潜在基因。因此，我们将研究高碳酸血症对肺泡上皮细胞和C。elegans通过四个相互关联的目标：在具体目标1中，我们提出确定高碳酸血症是否降低正常和受损肺中的肺泡液体重吸收，以及这些作用是否是由于高pCO 2或相关的酸中毒，在具体目标2中，我们将确定高碳酸血症促进肺泡上皮细胞中的肺泡上皮Na，K-ATP酶内吞作用的信号传导途径，在具体目标3中，我们将确定高碳酸血症是否通过磷酸化和泛素化抑制Na，K-ATP酶活性，导致Na，K-ATP酶蛋白的内吞作用和降解，在具体目标4中，我们将确定CO2水平升高对C的影响。elegans的发展，运动性和生育能力，并建立了一个系列的基因突变，调节传感和细胞反应的CO2水平升高。针对每个具体目标进行了实验，初步结果支持了该建议的可行性。拟定研究的完成将提供关于高碳酸血症对肺泡上皮影响的新信息，特别是因为它涉及肺泡上皮屏障功能障碍的机制，这可能对高碳酸血症患者的治疗具有重要意义。

DESCRIPTION (provided by applicant): Patients with COPD and mechanically ventilated patients with "permissive hypercapnia" can have significant elevation of pCO2. We hypothesize that hypercapnia causes dysfunction of the alveolar epithelium by specifically downregulating the alveolar epithelial Na,K-ATPase and decreasing alveolar fluid clearance. The focus of this application is to determine whether short term (30-60 min) hypercapnia impairs reversibly alveolar fluid reabsorption by inhibiting the Na,K-ATPase and promoting its endocytosis from the plasma membrane into intracellular compartments via specific pathways involving JNK and ERK kinases and protein kinase C (PKC) signaling molecules. We also propose to determine whether long term (5 and 7 days) hypercapnia sensitizes the alveolar epithelium to ventilation induced lung injury and causes not only endocytosis but degradation of the Na,K-ATPase via the ubiquitin/proteosome pathway. The cellular signals that sense hypercapnia are largely unknown, therefore we will take advantage that the genome of C. Elegans has been completely mapped and conduct experiments in C. elegans to study potential genes that may participate in the sensing and response to high pCO2. As such, we will study the effects of hypercapnia on the alveolar epithelium and C. elegans via four interrelated aims: in Specific Aim 1 we propose to determine whether hypercapnia decreases alveolar fluid reabsorption in normal and injured lungs and whether these effects are due to high pCO2 or the associated acidosis, in Specific Aim 2 we will determine the signaling pathways by which hypercapnia promotes alveolar epithelial Na,K-ATPase endocytosis in alveolar epithelial cells , in Specific Aim 3 we will determine whether hypercapnia inhibits Na,K-ATPase activity via phosphorylation and ubiquitination leading to the endocytosis and degradation of Na,K-ATPase proteins and in Specific Aim 4 we will determine the effect of elevated levels of CO2 on C. elegans development, motility and fertility and establish a collection of lines mutated in genes that regulate the sensing and cellular response to elevated levels of CO2. 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 hypercapnia on the alveolar epithelium, specifically as it pertains to mechanisms of alveolar epithelial barrier dysfunction which may be of importance for the treatment of hypercapnic patients.