Regulation of Renal Calcium Transport in Health and Disease

健康和疾病中肾脏钙转运的调节

• 批准号: 9562002
• 负责人: Chou-Long Huang
• 金额: $ 36.45万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-03 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9562002
• 关键词: Acids; Adenine; Affect; Binding; Blood; Blood Circulation; Calcified; Calciphylaxis; Calcium; Cardiovascular system; Cell membrane; Cell surface; Cessation of life; Chronic; Chronic Kidney Failure; Cleaved cell; Defect; Diet; Disaccharides; Disease; Disinhibition; Distal convoluted renal tubule structure; Down-Regulation; Endocrine; Endosomes; Epidemic; Excision; Excretory function; Extracellular Domain; Extracellular Fluid; Fibroblast Growth Factor Receptors; Functional disorder; Future; Galactose; Galectin 1; Gene Expression; General Population; Health; Homeostasis; Hormones; Hydrochlorothiazide; Hyperparathyroidism; Hypocalcemia result; In Vitro; Integral Membrane Protein; Kidney; Kidney Diseases; Lead; Length; Ligands; Link; Maintenance; Mannose; Mediating; Membrane Glycoproteins; Metabolic Diseases; Metabolism; Minerals; Mus; Mutation; Nephrectomy; Neuraminidase; PTH gene; Parathyroid gland; Pathogenesis; Pathway interactions; Patients; Phenotype; Physiological; Play; Polysaccharides; Pruritus; Public Health; Regulation; Renal tubule structure; Retrieval; Role; Secondary Hyperparathyroidism; Secondary to; Serum; Sialic Acids; Staining method; Stains; Surface; Testing; Tissues; Transgenic Mice; Transgenic Organisms; Up-Regulation; Urine; Vitamin D; adverse outcome; anti aging; apical membrane; base; calbindin-D28K; calcification; calcium excretion; combat; extracellular; feeding; hypercalciuria; in vitro activity; in vivo; inorganic phosphate; lactosamine; mortality; mouse model; mutant; novel; paracrine; patch clamp; premature; receptor; renal calcium; response; urinary

Project Summary TRPV5-mediated renal Ca2+ reabsorption is critical for maintaining Ca2+ homeostasis. Klotho is a type-1 transmembrane protein predominantly produced in the kidney. Klotho exists either in the membranous klotho form or the soluble ectodomain that is shedded into urine or systemic circulation to function as a paracrine or endocrine factor. Previously, we have found that soluble klotho increases surface abundance of TRPV5 by removing terminal sialic acids from the N-linked glycan chains of the channel. Removal of sialic acids exposes underlying disaccharide N-acetyl-lactosamine, a ligand for galectin-1 that is ubiquitously present on the external surface of cells. Binding to galectin-1 at the extracellular surface leads to accumulation of functional TRPV5 on the plasma membrane. To support this novel hypothesis for mechanism of action for klotho, we propose the following studies to extend our in vitro findings into in vivo and to investigate pathophysiological relevance of our findings. Aim-1 will test the hypothesis that soluble klotho regulates renal Ca2+ reabsorption via TRPV5 in vivo and that it does so through the putative sialidase activity of klotho. To examine the role of soluble klotho in renal Ca2+ reabsorption in vivo, we will generate transgenic mice that express soluble klotho in the background of klotho-deficient mice (rescued from death by dietary phosphate and vitamin D restriction) and examine urinary calcium excretion, TRPV5 expression, several gene expression parameters related to TRPV5-mediated Ca2+ reabsorption, and patch-clamp recording of TRPV5 channel activity in the native renal tubules. We will also define domains of soluble klotho involved in klotho's putative sialidase activity in vitro and test the effect of mutant soluble Klotho carrying sialidase activity-inactivating mutations in vivo. Chronic kidney disease (CKD) is a klotho-deficient state. Aim 2 will test the hypothesis that defect in TRPV5-mediated renal Ca2+ reabsorption from soluble klotho deficiency contributes to secondary hyperparathyroidism in chronic kidney disease. Elevation of parathyroid hormone (PTH) occurs in patients with CKD as a response to combat hyperphosphatemia and hypocalcemia. Hyperparathyroidism yet causes multiple adverse consequences in CKD. We will test the hypothesis that decrease in soluble klotho and downregulation of TRPV5 plays a role in hyperparathyroidism of CKD by using two mouse models of CKD. The effect of transgenic delivery of wild-type or inactive mutant soluble klotho on serum PTH and Ca2+ levels, urinary Ca2+ excretion, and other mineral metabolites in CKD will be examined. These studies on the mechanism by which klotho stimulates renal calcium reabsorption and its role in mineral metabolism of CKD will be important for our understanding the pathophysiology of mineral disorder of CKD and may help management or deign future therapy of mineral disorder in CKD patients.

项目摘要 TRPV 5介导的肾Ca 2+重吸收对于维持Ca 2+稳态至关重要。Klotho是第一型 主要在肾脏中产生的跨膜蛋白。Klotho要么存在于膜Klotho中， 形式或可溶性胞外域，其脱落到尿液或体循环中以起到旁分泌的作用，或 内分泌因子以前，我们已经发现可溶性klotho通过增加TRPV 5的表面丰度， 从所述通道的N-连接聚糖链去除末端唾液酸。唾液酸的去除暴露了 潜在的二糖N-乙酰基-乳糖胺，半乳糖凝集素-1的配体，其普遍存在于 细胞的外表面。在细胞外表面与半乳糖凝集素-1结合导致功能性半乳糖凝集素的积累。 TRPV 5在细胞膜上的表达。为了支持klotho作用机制的新假设，我们 我建议进行以下研究，将我们的体外研究结果扩展到体内，并研究病理生理学 我们发现的相关性。目的-1将检验可溶性klotho调节肾Ca 2+重吸收的假设 通过TRPV 5在体内，它通过klotho的推定唾液酸酶活性这样做。审查的作用 可溶性klotho在体内肾Ca 2+重吸收中的作用，我们将产生表达可溶性klotho的转基因小鼠， klotho缺陷小鼠（通过饮食磷酸盐和维生素D限制免于死亡）的背景 并检测尿钙排泄、TRPV 5表达、与尿钙排泄相关的几个基因表达参数， TRPV 5介导的Ca ~（2+）重吸收和TRPV 5通道活性的膜片钳记录 小管我们还将定义可溶性klotho的结构域，该结构域参与klotho的体外推定唾液酸酶活性， 测试携带唾液酸酶活性失活突变的突变体可溶性Klotho在体内的作用。慢性肾脏 CKD是一种klotho缺乏状态。目的2将检验TRPV 5介导的肾细胞凋亡缺陷的假设。 可溶性klotho缺乏引起的钙重吸收导致慢性甲状旁腺功能亢进 肾病CKD患者甲状旁腺激素（PTH）升高是对战斗的反应。 高磷血症和低钙血症。甲状旁腺功能亢进仍会导致多种不良后果， CKD。我们将检验可溶性klotho的减少和TRPV 5的下调在细胞凋亡中起作用的假设。 通过使用两种CKD小鼠模型观察CKD的甲状旁腺功能亢进。转基因递送野生型的效果 或无活性突变体可溶性klotho对血清PTH和Ca 2+水平、尿Ca 2+排泄和其他矿物质的影响， 将检查CKD中的代谢物。这些关于Klotho刺激肾脏的机制的研究 钙重吸收及其在CKD矿物质代谢中的作用将对我们理解 CKD矿物质紊乱的病理生理学，并可能有助于管理或设计未来的治疗， CKD患者的疾病。