Regulation of Renal Calcium Transport in Health and Disease

健康和疾病中肾脏钙转运的调节

基本信息

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
项目类别：
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

项目摘要

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 介导的肾脏 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。形成或可溶性胞外域，脱落到尿液或体循环中，起到旁分泌或此前，我们发现可溶性 klotho 可增加 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会暴露。潜在的二糖 N-乙酰基-乳糖胺，半乳糖凝集素-1 的配体，普遍存在于细胞外表面与细胞外表面的 galectin-1 结合导致功能性积累。为了支持 klotho 作用机制的这一新假设，我们研究了质膜上的 TRPV5。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 Aim-1 将检验可溶性 klotho 调节肾脏 Ca2+ 重吸收的假设。体内通过 TRPV5 进行检测，并通过 klotho 的假定唾液酸酶活性来检测其作用。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我们还将定义参与 klotho 假定唾液酸酶活性的可溶性 klotho 的域。测试携带唾液酸酶活性失活突变的可溶性 Klotho 突变体在慢性肾脏中的作用。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慢性肾病患者的甲状旁腺激素 (PTH) 升高是对抗肾脏疾病的一种反应。高磷血症和低钙血症会导致多种不良后果。我们将检验可溶性 klotho 的减少和 TRPV5 的下调在 CKD 中发挥作用的假设。使用两种 CKD 小鼠模型观察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，这些研究涉及 klotho 刺激肾脏的机制。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疗法 CKD 患者的疾病。