ENaC regulation by biliary factors

胆道因素对 ENaC 的调节

基本信息

批准号：
10027936
负责人：
OSSAMA B KASHLAN
金额：
$ 49.42万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-21 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10027936
关键词：
Acute Affect Alcoholic Hepatitis Aldosterone Apical Ascites Bile Acids Biliary Bilirubin Binding Binding Sites Blood Blood Volume Body Fluids Body Water Cell model Cells Charge Chemistry Chronic Colon Data Distal Diuretics Drug usage Duct (organ) structure Ductal Epithelial Cell Ductal Epithelium Edema Electrolytes Epithelial Epithelial Cells Epithelium Excretory function Exhibits Extracellular Fluid Goals Hormonal Hyperbilirubinemia Hypertension Hypokalemia In Vitro Ion Transport Ions Kidney Lead Learning Liddle syndrome Liquid substance Liver Liver Failure Liver diseases Measurement Measures Mediating Membrane Mineralocorticoids Modeling Molecular Mus Mutation Nephrons Patients Peripheral Plasma Portal Hypertension Potassium Potassium Channel Regulation Renal tubule structure Renin-Angiotensin-Aldosterone System Reporting Role Sampling Serum Side Site Site-Directed Mutagenesis Sodium Spironolactone Taurocholic Acid Testing Tubular formation Urine Vasodilation Weight Xenopus oocyte biophysical properties chemical property chronic liver disease driving force epithelial Na+ channel experience experimental study extracellular gain of function in vivo mutant novel therapeutics prevent response urinary

项目摘要

Patients with advanced liver disease often experience fluid retention and electrolyte disturbances due in part to activation of the renin-angiotensin-aldosterone system. Aldosterone activates the epithelial Na+ channel (ENaC) in the distal nephron, enhancing urinary Na+ retention and K+ excretion. But many liver disease patients experience fluid retention without apparent activation of the renin-angiotensin-aldosterone system. Liver disease patients also frequently exhibit elevated plasma bile acids and hyperbilirubinemia. Preliminary data show that specific bile acids and conjugated bilirubin activate ENaC in vitro and that taurocholic acid activates ENaC in isolated collecting ducts. The central hypothesis of our proposal is that urinary bile acids and possibly conjugated bilirubin directly activate ENaC, promoting Na+ and fluid retention and K+ excretion. The goal of this proposal is to determine whether this regulation occurs in vivo and to determine the molecular mechanism of regulation. Preliminary data suggest that urine from patients with alcoholic hepatitis contains concentrations of activating bile acids and conjugated bilirubin sufficient to activate the channel, and indeed activate the channel. We will determine whether urine from alcoholic hepatitis patients activates ENaC in experimental cells. We further hypothesize that elevated biliary factors activate ENaC in the distal nephron, promoting Na+ retention and K+ excretion. We will test this hypothesis in isolated collecting ducts. We will also test this hypothesis in vivo by chronically administering biliary factors and quantifying changes in weight, blood K+, urinary Na+ and K+, plasma aldosterone, and total body water. Preliminary data suggest that taurocholic acid promotes Na+ retention, K+ loss, and volume expansion. We will determine the contribution of ENaC to observed changes by using drugs to block ENaC, and by using genetic changes to increase the channel's sensitivity to biliary factors or to decrease ENaC's contribution to renal Na+ and K+ handling. Experiments will also investigate the molecular mechanism of ENaC regulation by bile acids. We hypothesize that activating bile acids interact directly with the channel to enhance channel activity. We will use derivatized bile acids to detect direct binding, and key chemical and biophysical properties of the both the channel and bile acids to dissect the interaction. Key experiments will be performed in multiple cell models, including Xenopus oocytes, cultured epithelial cells, and mouse collecting ducts. Direct ENaC activation by biliary factors elevated in liver disease has not been previously investigated, and may contribute to the volume overload, edema, and electrolyte imbalances associated with liver disease.

晚期肝病患者经常出现液体潴留和电解质紊乱部分原因是肾素-血管紧张素-醛固酮系统的激活。醛固酮激活远端肾单位的上皮 Na+ 通道 (ENaC)，增强尿 Na+ 潴留和 K+ 排泄。但许多肝病患者会出现液体潴留，但没有明显的激活肾素-血管紧张素-醛固酮系统。肝病患者也经常出现血浆胆汁酸升高和高胆红素血症。初步数据表明，特定胆汁酸结合胆红素在体外激活 ENaC，牛磺胆酸在体外激活 ENaC 孤立的集合管。我们提案的中心假设是尿胆汁酸和可能结合胆红素直接激活 ENaC，促进 Na+ 和液体潴留以及 K+ 排泄。该提案的目标是确定这种调节是否发生在体内并确定调控的分子机制。初步数据表明，尿液来自酒精性肝炎患者体内含有一定浓度的活化胆汁酸和结合胆汁酸胆红素足以激活该通道，并且确实激活了该通道。我们将确定酒精性肝炎患者的尿液是否会激活实验细胞中的 ENaC。我们进一步假设升高的胆道因子激活远端肾单位的 ENaC，促进 Na+ 滞留和 K+ 排泄。我们将在孤立的集合管中检验这一假设。我们还将通过长期施用胆道因子并量化胆道因子的变化，在体内检验这一假设体重、血 K+、尿 Na+ 和 K+、血浆醛固酮和体内水分总量。初步数据表明牛磺胆酸促进 Na+ 保留、K+ 损失和体积膨胀。我们将通过使用药物阻断 ENaC 来确定 ENaC 对观察到的变化的贡献，并通过利用基因改变来增加通道对胆道因素的敏感性或减少 ENaC 对肾脏 Na+ 和 K+ 处理的贡献。实验还将研究分子胆汁酸调节 ENaC 的机制。我们假设激活胆汁酸相互作用直接与渠道合作，提升渠道活跃度。我们将使用衍生胆汁酸来检测通道和胆汁酸的直接结合以及关键化学和生物物理特性剖析交互。关键实验将在多种细胞模型中进行，包括非洲爪蟾卵母细胞、培养的上皮细胞和小鼠集合管。直接 ENaC 激活肝脏疾病中胆道因素升高之前尚未被研究过，并且可能有助于与肝病相关的容量超负荷、水肿和电解质失衡。