Investigation of how activation of AMP activated protein kinase (AMPK) inhibits Na+ transport across H441 lung epithelial cells.

肺的气道衬有一层薄薄的液体，这对防止吸入物的感染和损害很重要。这一流体层的体积是由钠和氯化物通过选择性通道和泵在气道表面排列的细胞之间的运输控制的。钠从气道转运增加与体液层脱水有关，是囊性纤维化疾病的一个促成因素。相反，钠转运的减少与气道中的液体积聚有关，如新生儿呼吸窘迫综合征和高原肺水肿。我们最近发现，一种感知低细胞能量水平的分子（腺苷单磷酸活化蛋白激酶（AMPK））的激活会减少钠在肺上皮细胞中的转运。因此AMPK可能在这些疾病中具有重要的调节作用。AMPK如何减少钠转运尚不清楚，但它似乎减少了通过酰胺敏感钠通道（ENaC）进入细胞的钠量，以及通过Na+K+ atp酶泵从细胞中挤出的钠量。本项目的目的是研究AMPK的激活是否通过调节细胞膜中ENaC和Na+K+ atp酶的丰度来降低钠转运和ENaC和Na+K+ atp酶的活性。我们将在体外对人类H441肺上皮细胞使用功能和分子方法相结合的方法来做到这一点。确定该分子调节钠转运的细胞机制是阐明其如何调节体内Na+转运的关键第一步。这将有助于我们制定更合适的策略，用于未来的研究和开发肺部疾病的治疗方法，这些疾病表现为体液紊乱。

The airways of the lung are lined with a thin layer of fluid that is important in preventing infection and damage from inhaled agents. The volume of this fluid layer is controlled by the transport of sodium and chloride across the cells that line the surface of the airways through selective channels and pumps. Increased transport of sodium out of the airway is linked with dehydration of the fluid layer and is a contributary factor to the disease, Cystic Fibrosis. Conversely, a decrease in sodium transport is associated with fluid accumulation in the airways as in respiratory distress syndrome of the newborn and high altitude pulmonary oedema. We have recently discovered that activation of a molecule that senses low cellular energy levels (adenosine monophosphate-activated protein kinase (AMPK), decreases sodium transport across the lung epithelial cell. AMPK may therefore have a significant regulatory role in these diseases. How AMPK decreases sodium transport is unknown but it appears to reduce the amount of sodium entering the cell via the amiloride-sensitive sodium channel (ENaC) and the amount of sodium that is extruded from the cell via the Na+K+ATPase pump. It is the purpose of this project to investigate if activation of AMPK decreases sodium transport and the activity of ENaC and Na+K+ATPase by regulating the abundance of these proteins in the cell membrane. We will do this by using a combination of functional and molecular approaches in human H441 lung epithelial cells in vitro. Identifying the cellular mechanism by which this molecule regulates sodium transport is a critical first step in elucidating how it could regulate Na+ transport in vivo. This will help us develop more appropriate strategies for future research and the development of treatments for lung diseases which exhibit dissordered fluid volume.