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.

项目概要 TRPV5 介导的肾 Ca2+ 重吸收对于维持 Ca2+ 稳态至关重要。 Klotho 是 1 型 跨膜蛋白主要在肾脏中产生。 Klotho 存在于膜 klotho 中 形成或可溶性胞外域，脱落到尿液或体循环中，起到旁分泌或 内分泌因素。之前，我们发现可溶性 klotho 通过增加 TRPV5 的表面丰度 从通道的 N 连接聚糖链中去除末端唾液酸。去除唾液酸暴露 潜在的二糖 N-乙酰基-乳糖胺，半乳糖凝集素-1 的配体，普遍存在于 细胞的外表面。与细胞外表面的 galectin-1 结合导致功能性物质的积累 TRPV5 位于质膜上。为了支持 klotho 作用机制的这一新假设，我们 提出以下研究，将我们的体外研究结果扩展到体内并研究病理生理学 我们的研究结果的相关性。 Aim-1 将检验可溶性 klotho 调节肾 Ca2+ 重吸收的假设 体内通过TRPV5，并且通过klotho假定的唾液酸酶活性来实现这一点。要检查的作用 可溶性klotho在体内肾Ca2+重吸收中的作用，我们将产生在体内表达可溶性klotho的转基因小鼠 klotho 缺陷小鼠的背景（通过膳食磷酸盐和维生素 D 限制而免于死亡） 并检查尿钙排泄、TRPV5表达、与相关的几个基因表达参数 TRPV5 介导的 Ca2+ 重吸收，以及天然肾中 TRPV5 通道活性的膜片钳记录 小管。我们还将定义参与 klotho 体外假定唾液酸酶活性的可溶性 klotho 的域，以及 测试携带唾液酸酶活性失活突变的突变体可溶性 Klotho 的体内效果。慢性肾病 疾病（CKD）是一种 klotho 缺陷状态。目标 2 将检验 TRPV5 介导的肾脏缺陷的假设 可溶性 klotho 缺乏导致的 Ca2+ 重吸收导致慢性继发性甲状旁腺功能亢进 肾脏疾病。 CKD 患者的甲状旁腺激素 (PTH) 升高是对战斗的反应 高磷血症和低钙血症。甲状旁腺功能亢进症会导致多种不良后果 慢性肾病。我们将检验以下假设：可溶性 klotho 的减少和 TRPV5 的下调在 通过使用两种 CKD 小鼠模型来研究 CKD 甲状旁腺功能亢进症。野生型转基因递送的效果 或失活突变体可溶性 klotho 对血清 PTH 和 Ca2+ 水平、尿 Ca2+ 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刺激肾脏机制的研究 钙重吸收及其在 CKD 矿物质代谢中的作用对于我们了解 CKD 矿物质紊乱的病理生理学，可能有助于管理或设计未来的矿物质治疗 CKD 患者的疾病。