The injurious effects of hypercapnia on the alveolar epithelium

描述（由申请方提供）：在采用“允许性高碳酸血症”策略通气的呼吸衰竭患者中观察到高pCO 2水平。我们和其他研究人员最近报道，pCO 2水平升高可能对肺泡上皮细胞产生有害影响。具体而言，我们已经报道了高碳酸血症通过抑制肺泡上皮Na，K-ATP酶和损害肺泡液体清除而导致肺泡上皮的短期和长期功能障碍。因此，本申请的重点是确定在高碳酸血症条件下导致肺泡Na，K-ATP酶下调和肺泡液体清除受损的特定机制/信号通路。我们建议通过四个相互关联的目的来研究高碳酸血症对肺泡上皮的影响：在与特定目的1相关的实验中，我们将确定可溶性腺苷酸环化酶是否参与高碳酸血症诱导的肺泡上皮中Na，K-ATP酶下调;在与具体目标#2有关的实验中，我们将确定高碳酸血症诱导的JNK活化是否导致LIM结构域蛋白7（LMO 7）磷酸化和Na，K-ATP酶内吞作用;在与特定目标3相关的研究中，我们将确定Na，K-ATP酶泛素化是否导致其下调，并提出鉴定特异性E2和E3泛素连接酶;并且在与具体目标#4有关的研究中，我们将确定高碳酸血症是否进一步损害轻度呼吸机诱导的肺损伤（VILI）模型中的损伤，以及sAC，JNK，LMO 7和泛素化途径将逆转高碳酸血症对啮齿动物肺泡液体清除的有害影响。在第一轮拨款中，我们完成了大部分拟议研究，并为每个当前的具体目标进行了初步实验，这些目标支持这项拨款建议的可行性。该实验将为高碳酸血症对肺泡上皮细胞的影响提供新的信息，并揭示导致肺泡上皮功能障碍的机制，这对理解和治疗急性肺损伤患者具有重要意义。

DESCRIPTION (provided by applicant): High pCO2 levels are observed in patients with respiratory failure ventilated with the "permissive hypercapnia" strategy. We and other investigators have recently reported that elevated pCO2 levels can have deleterious effects on the alveolar epithelium. Specifically, we have reported that hypercapnia leads to short and long term dysfunction of the alveolar epithelium by inhibiting the alveolar epithelial Na,K-ATPase and impairing alveolar fluid clearance. Therefore, the focus of this application is to determine the specific mechanisms/signaling pathways that lead to alveolar Na,K-ATPase downregulation and impairment in alveolar fluid clearance during hypercapnic conditions. We propose to study the effects of hypercapnia on the alveolar epithelium via four interrelated aims: in experiments pertaining to specific Aim # 1, we will determine whether the soluble adenylyl cyclase participates in the hypercapnia-induced Na,K- ATPase downregulation in the alveolar epithelium; in experiments pertaining to specific Aim #2, we will determine whether hypercapnia-induced JNK activation leads to LIM-domain only protein 7 (LMO7) phosphorylation and Na,K-ATPase endocytosis; in studies pertaining to specific Aim # 3, we will determine whether Na,K-ATPase ubiquitination leads to its downregulation and propose to identify the specific E2 and E3 ubiquitin ligase; and in studies pertaining to specific Aim # 4, we will determine whether hypercapnia further impairs injury in a model of mild ventilator-induced lung injury (VILI) and whether the downregulation of of sAC, JNK, LMO7 and ubiquitination pathway will reverse the deleterious effects of hypercapnia on alveolar fluid clearance in rodents. In the first cycle of this grant, we have completed most of the proposed research and conducted preliminary experiments for each of the current specific aims which support the feasibility of this grant proposal. The proposed experiments will generate novel information on the effects of hypercapnia on the alveolar epithelium and shed light on the mechanisms leading to alveolar epithelial dysfuntion which is of significance to the understanding and treatment of patients with acute lung injury.