Cellular Mechanisms of Action of Mineralocorticoid Hormones

描述（由申请人提供）：维持细胞外液容量稳态对血流动力学稳定至关重要，肾脏钠处理异常与心血管疾病和高血压有关。钠排泄的最终调控发生在远端肾元，通过阿米洛胺敏感上皮Na+通道（ENaC）的传导运输。ENaC在上皮细胞顶膜的表达和活性是限制Na+在肾集管、气道上皮和结肠重吸收的速率步骤。醛固酮是ENaC在应答性上皮中表达和活性的主要调节因子。本研究的长期目标是确定矿化皮质激素调节反应性上皮中钠载体运输的机制，并研究醛固酮调节ENaC的机制。当前资助期的工作已经确定了一种特定的甲基转移酶，它可以激活ENaC，敲低该蛋白可以阻断早期醛固酮反应。目前正在进行研究，以确定这种蛋白质是否需要通过基因组或非基因组途径进行早期醛固酮作用，并确定醛固酮如何调节其活性。我们已经证明了ENaC在脂筏中运输到上皮细胞的顶膜，并建立了ENaC在培养上皮细胞中的运输模型。缺乏醛固酮的细胞丧失Na+再吸收活性，同时丧失ENaC的细胞再循环区。随着时间的推移，醛固酮恢复了循环室和Na+再吸收表型。ENaC循环途径的恢复与内体室中多种蛋白质的腺苷酮上调有关。我们将研究醛固酮通过内体和/或再循环途径调节ENaC的顶端表达的机制，并确定哪些特定的醛固酮调节蛋白控制ENaC在内体和外囊室中的活性和表达，以支持ENaC在Na+再吸收状态下的顶端易位。公共卫生相关性：醛固酮是哺乳动物肾脏中调节钠重吸收的主要激素。钠处理异常可导致高血压和心力衰竭等疾病，并与囊性纤维化肺病的进展有关。这些研究旨在确定醛固酮调节肾细胞钠重吸收的细胞机制。

DESCRIPTION (provided by applicant): Maintenance of extrecellular fluid volume homeostasis is essential for hemodynamic stability, and abnormalities of renal sodium handling have been linked to cardiovascular disease and hypertension. Ultimate regulation of sodium excretion occurs in the distal nephron via conductive transport through the amiloride sensitive epithelial Na+ channel (ENaC). ENaC expression and activity in the apical membrane of epithelial cells is the rate limiting step in Na+ reabsorption not only in the kidney collecting duct, but in airway epithelia and colon as well. Aldosterone is the major regulator of ENaC expression and activity in responsive epithelia. The long term objectives of this research are to define the mechanisms through which mineralocorticoid hormones regulate vectorial sodium transport in responsive epithelia and studies are designed to examine the mechanisms of aldosterone regulation of ENaC. Work in the current grant period has identified a specific methyltransferase which activates ENaC and knockdown of this protein blocks the early aldosterone response. Studies are now proposed to determine if this protein is required for early aldosterone action via genomic or non-genomic pathways and determine how aldosterone regulates its activity. We have demonstrated that ENaC trafficks to the apical membrane of epithelial cells in lipid rafts and developed a model of ENaC trafficking in cultured epithelial cells. Cells deprived of aldosterone develop a loss of Na+ reabsorptive activity along with a loss of the cellular recycling compartment for ENaC. The recycling compartment and Na+ reabsorptive phenotype are restored over time by aldosterone. Restoration of the ENaC recycling pathway is associated with adlosterone up-regulation of multiple proteins in the endosomal compartment. We will examine the mechanisms by which aldosterone regulates the apical expression of ENaC from endosomal and/or recycling pathways and determine what specific aldosterone-regulated proteins control the activity and expression of ENaC in endosomes and exocyst compartments to support apical translocation of ENaC in Na+ reabsorptive states. PUBLIC HEALTH RELEVANCE: Aldosterone is the major hormone which regulates sodium reabsorption in the kidney in mammals. Abnormalities of sodium handling can lead to diseases such as hypertension and heart failure and have been implicated in the progression of cystic fibrosis lung disease. These studies are designed to determine the cellular mechanisms by which aldosterone regulates sodium reabsorption in kidney cells.