Defective flow-dependent tubule transport in the pathogenesis of kidney disease

肾脏疾病发病机制中的血流依赖性肾小管运输缺陷

基本信息

批准号：
10063866
负责人：
Tong Wang Wang
金额：
$ 37.71万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-02-01 至 2022-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10063866
关键词：
Acids Aneurysm Animal Disease Models Apical Architecture Blood Vessels Calcium Cations Cells Cyst Cystic kidney DNA Sequence Alteration Data Defect Development Distal Dopamine Dopamine Antagonists Dopamine Receptor Electrolyte Balance Endocytosis Endoplasmic Reticulum Equilibrium Excretory function Family Feedback Fluid Balance Glomerular Filtration Rate Growth Hydrostatic Pressure Impairment Kidney Kidney Diseases Knockout Mice Liquid substance Measures Mediating Methodology Methods Modeling Mus Mutant Strains Mice Mutation Na(+)-K(+)-Exchanging ATPase Natriuresis Nephrons Null Lymphocytes PKD2 protein Pathogenesis Patients Permeability Phenotype Physiological Play Polycystic Kidney Diseases Potassium Prevention Proton-Translocating ATPases Regulation Renal clearance function Renal tubule structure Role Ryanodine Receptor Calcium Release Channel Signal Transduction Sodium Sodium Chloride Surface TRP channel Testing Tubular formation Variant Work absorption conditional knockout glomerular filtration immunocytochemistry improved in vivo inhibitor/antagonist kidney dysfunction kidney preservation mathematical model mutant mouse model novel novel therapeutics preservation pressure prevent protein intake receptor renal damage shear stress trafficking

项目摘要

Summary The scientific premise of the proposed work is that defective flow-dependent proximal tubule transport may be instrumental in the development and progression of renal dysfunction, a notable example being polycystic kidney disease (PKD). Flow-dependent transport underlies glomerulotubular balance (GTB), whose physiologic importance has been appreciated in normal fluid and electrolyte balance. Whether impaired GTB predisposes to renal damage before GFR is reduced (such as renal cyst formation), has not been studied. Our hypothesis is that episodic increases in kidney tubule hydrostatic pressure promote the creation and growth of renal cysts. This is analogous to the role of arterial pressure in creating vascular aneurysms. There are two control mechanisms, which serve to mitigate the swings in tubule pressure that accompany increases in glomerular filtration (GFR), namely tubuloglomerular feedback and glomerulotubular balance. A mathematical model estimates that with a 50% increase in GFR, impaired GTB increases distal nephron flow, and provokes sharp increases pressure by about 40% in both proximal and distal tubule. This role of GTB, to mitigate GFR- dependent changes of tubule pressure, has never been examined experimentally. During the past 15 years, we have demonstrated that IP3 receptor-mediated intracellular Ca2+ signaling plays a critical role in GTB. Polycystin-2 (PC2) is a nonselective calcium permeable cation channel belonging to the Transient Receptor Potential (TRP) channel family and functions as a Ca2+ channel in the endoplasmic reticulum (ER). Mutations of PC2 abolished the IP3-induced calcium release from the ER. Our preliminary data show a) defective GTB in Pkd2 mutant mice before any renal cysts are formed; and b) Dopamine receptor (DA1) antagonist improves tubule sensitivity to flow and also reduces renal cyst formation in Pkd2 KO mouse, suggesting a new therapeutic method to treat renal disease. In the proposed work, mathematical modeling, renal clearance, microperfusion, immunocytochemistry, and measuring renal tubular pressure in vivo will be used. Hypotheses will be studied in novel nephron-specific conditional knockout PKD animal models, and in PKD null cells to test whether: 1) GTB stabilizes kidney tubule hydrostatic pressure during variations of GFR. This limits tubule distention during GFR elevation, and thus acts to slow cyst formation in Pkd2 KO mice. 2) Inhibition of Na/H- exchanger 3 (NHE3) endocytosis by a DA1 antagonist, increases proximal tubule transporter flow sensitivity, and prevents cyst formation in Pkd2 KO mice. 3) Flow-stimulated NHE3 and/or Na+/K-ATPase trafficking is abrogated in Pkd2 null cells and is similar to blocking the IP3 receptor in WT cells. Inhibition of NHE3 endocytosis by DA1 inhibitor and/or by increasing Ca2+ release from the ER restores the flow-sensing in Pkd2 null cells.

概括这项工作的科学前提是，有缺陷的流量依赖性近端小管运输可能是有助于肾功能障碍的发生和进展，一个著名的例子是多囊肾肾脏疾病（PKD）。血流依赖性转运是肾小球小管平衡 (GTB) 的基础，其生理学人们已经认识到正常液体和电解质平衡的重要性。 GTB 受损是否易诱发肾小球滤过率（GFR）降低之前的肾损伤（如肾囊肿形成），尚未被研究。我们的假设肾小管静水压的间歇性升高促进了肾囊肿的产生和生长。这类似于动脉压在形成血管动脉瘤中的作用。有两个控制机制，用于减轻伴随肾小球增加的肾小管压力波动滤过（GFR），即肾小球反馈和肾小球平衡。数学模型据估计，随着 GFR 增加 50%，受损的 GTB 会增加远端肾单位流量，并引发急剧的近端和远端肾小管的压力增加约 40%。 GTB 的作用是减轻 GFR- 肾小管压力的相关变化从未被实验检验过。在过去的15年里，我们已经证明 IP3 受体介导的细胞内 Ca2+ 信号传导在 GTB 中发挥着关键作用。 Polycystin-2 (PC2) 是一种非选择性钙渗透性阳离子通道，属于瞬时受体电位 (TRP) 通道家族，在内质网 (ER) 中充当 Ca2+ 通道。突变 PC2 消除了 IP3 诱导的 ER 钙释放。我们的初步数据显示 a) 有缺陷的 GTB 在任何肾囊肿形成之前的 Pkd2 突变小鼠； b) 多巴胺受体 (DA1) 拮抗剂改善 Pkd2 KO 小鼠肾小管对血流的敏感性，还减少了肾囊肿的形成，这表明一种新的方法治疗肾病的治疗方法。在拟议的工作中，数学模型、肾脏清除率、将使用微灌注、免疫细胞化学和测量体内肾小管压力。假设将在新型肾单位特异性条件性敲除 PKD 动物模型中进行研究，并在 PKD 无效细胞中进行测试是否：1) GTB 在 GFR 变化期间稳定肾小管静水压。这限制了肾小管 GFR 升高期间膨胀，从而减缓 Pkd2 KO 小鼠囊肿的形成。 2) Na/H-的抑制 DA1 拮抗剂的交换器 3 (NHE3) 内吞作用，增加近端小管转运蛋白的流动敏感性，并防止 Pkd2 KO 小鼠中囊肿的形成。 3) 流量刺激的 NHE3 和/或 Na+/K-ATPase 运输是在 Pkd2 无效细胞中被消除，类似于在 WT 细胞中阻断 IP3 受体。 NHE3 的抑制 DA1 抑制剂的内吞作用和/或增加 ER 中 Ca2+ 的释放可恢复 Pkd2 中的流量感应无效细胞。