Bicarbonate Transport by the Maturing Renal Tubule

描述（由申请人提供）：代谢性酸中毒是一种常见但可能危及生命的疾病，当过多的酸在血液中积累时发生。肾脏是主要负责控制血液中酸含量的器官。皮质集管（CCD）是肾小管的关键部分，它适应酸的水平并将其排出体外。长期目标是确定CCD的嵌入细胞如何通过改变其酸和碳酸氢盐的运输来适应酸碱干扰。正常分泌碳酸氢盐的兔ccd在pH 6.8下孵育3小时后分泌酸。这种新型蛋白在适应b嵌入细胞（ic）周围的细胞外基质（ECM）中表达。目的1探讨环孢素A （CsA）如何通过阻止b - ic的适应导致远端肾小管酸中毒。CsA通过抑制亲环蛋白的脯氨酸顺式/反式异构酶活性来抑制母鸡素聚合。使用这些功能的特定抑制剂在培养的ic和ccd中进行研究，在pH 6.8 x 3小时孵育以模拟酸中毒。亲环蛋白的“敲低”将决定哪种亲环蛋白在培养的ic中对母鸡素的聚合起重要作用，以及这种亲环蛋白的缺陷是否会导致酸中毒。目的2研究半乳糖凝集素-3在母鸡素聚合中的作用。半乳糖凝集素-3可以帮助体外聚合母鸡素。这个目的将确定哪些细胞表达半乳糖凝集素-3，以及在酸中毒过程中是否有刺激。目的是确定半乳糖凝集素-3是否在适应CCD的ECM中与母鸡素结合，以及半乳糖凝集素-3的显性阴性是否会阻止CCD中b - ic的母鸡素聚合和适应。本实验旨在探讨表达的亲环蛋白和半乳糖凝集素-3是否能协同体外聚合母鸡素。目的3确定哪些整合素在ic中表达，哪些与母鸡素相互作用。Aim 4研究了母鸡素聚合在CCD ic适应代谢性碱中毒中的作用，假设碱中毒通过降解母鸡素和允许b - ic分泌碳酸氢盐来纠正异常，从而减少了母鸡素聚合。这些研究应该表明母鸡素和相关蛋白如何介导肾脏对酸碱干扰的适应。

DESCRIPTION (provided by applicant): Metabolic acidosis is a common but potentially life-threatening disorder that occurs when too much acid accumulates in the blood. The kidney is the organ that is primarily concerned with controlling the level of acid in the blood. The cortical collecting duct (CCD) is a key segment of kidney tubule that adapts to the level of acid and pumps it out of the body. The long term goal is to determine how intercalated cells of the CCD adapt to acid-base disturbances by changing their transport of acid and bicarbonate. Rabbit CCDs, which normally secrete bicarbonate, exposed to 3h incubation at pH 6.8 secrete acid. The novel protein hensin is expressed in the extracellular matrix (ECM) surrounding adapting B-intercalated cells (ICs). Aim 1 examines how Cyclosporin A (CsA) causes a distal renal tubular acidosis by preventing adaptation of B-ICs. CsA inhibits hensin polymerization via an inhibition of the proline cis/trans isomerase activity of cyclophilin. Studies using specific inhibitors of these functions are performed in cultured ICs and CCDs incubated at pH 6.8 x 3 h to simulate acidosis. A "knockdown" of cyclophilin will determine which cyclophilin is important in polymerizing hensin in cultured ICs and whether a defect in this cyclophilin causes acidosis. Aim 2 examines the role of galectin-3 in hensin polymerization. Galectin-3 can help polymerize hensin in vitro. This aim will determine which cells express galectin-3 and whether there is stimulation during acidosis. The aim will determine if galectin-3 associates with hensin in the ECM in the adapting CCD and whether a dominant negative galectin-3 prevents hensin polymerization and adaptation of B-ICs in CCD. This aim will establish whether expressed cyclophilin and galectin-3 can cooperate in the in vitro polymerization of hensin. Aim 3 determines which integrins are expressed in ICs and which interact with hensin. Aim 4 examines the role of hensin polymerization in the adaptation of CCD ICs to metabolic alkalosis, with the hypothesis that alkalosis reduces hensin polymerization by degrading hensin and allowing B-ICs to secrete bicarbonate to correct the abnormality. These studies should show how hensin and associated proteins mediate the kidney's adaptation to acid-base disturbances.