Lipid Rafts: Mechanosensors of the distal nephron

脂筏：远端肾单位的机械传感器

• 批准号: 9135629
• 负责人: RAJEEV ROHATGI
• 金额: --
• 依托单位: NORTHPORT VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-10-01 至 2020-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9135629
• 关键词: Acute; Adult; Affect; Apical; Avidity; BIK gene; Blood; Blood Pressure; Blood Vessels; Calcium; Cations; Cell membrane; Cells; Cholesterol; Chronic; Congestive Heart Failure; Consumption; Coronary Arteriosclerosis; Cyclodextrins; Development; Diabetes Mellitus; Diabetic Nephropathy; Diet; Dinoprostone; Disease; Distal; Duct (organ) structure; Ductal Epithelial Cell; Epidemic; Epithelial; Epithelium; Erythrocytes; Essential Hypertension; Etiology; Excretory function; Functional disorder; Homeostasis; Human; Hyperaldosteronism; Hyperinsulinism; Hypertension; In Vitro; Ingestion; Ion Transport; Kidney; Kidney Diseases; Kidney Failure; Link; Liquid substance; Mediating; Membrane Microdomains; Metabolism; Mitogen-Activated Protein Kinases; Morbidity - disease rate; Mus; Natriuresis; Nephrons; Nitric Oxide; Organ; PKD2 protein; PTGS2 gene; Phenotype; Phosphorylation; Play; Population; Prostaglandins; Rattus; Receptor Signaling; Regulation; Research; Research Personnel; Rest; Risk; Rodent; Role; Saline; Sensory; Signal Pathway; Signal Transduction; Sodium; Stroke; Testing; Tissues; Tubular formation; Vascular Diseases; Vasodilation; Viscosity; absorption; antidiuretic; autocrine; base; blood pressure reduction; caveolin 1; chelation; diabetic; environmental change; flotillin; fluid flow; hypercholesterolemia; improved; in vivo; insulin secretion; mechanical force; mitogen-activated protein kinase p38; mortality; novel; paracrine; public health relevance; receptor; renal epithelium; response; salt sensitive; sensor; shear stress

 DESCRIPTION (provided by applicant): Mechanical forces and, in particular, fluid shear stress (FSS), play critical roles in endothelia t regulate vascular reactivity and in renal epithelia to affect transepithelial cation transport that collectively, determine blood pressue (BP). Mechano-regulation of tubular cation transport is an important regulator of renal sodium (Na) homeostasis and infers the existence of mechano-sensors in renal epithelia. Cholesterol (chol)-rich lipid rafts (LRs), signaling platforms located n the plasma membrane (PM), are mechano-sensors that induce paracrine-mediated vasodilation (ie nitric oxide synthesis) in endothelia; however, little is known about their sensory function in renal epithelia. Chol maintains the integrity of LRs, and effects on LR chol content impacts FSS-mediated signaling. The FSS-sensitive signaling pathways which regulate mitogen activated protein kinase (MAPK) and intracellular Ca2+ concentration ([Ca2+]i) in cells (1) are localized to caveolin-1 (cav-1) and flotillin-2 (flot-2), respectively, expressing LRs in collecting duct (CD) cells, and (2) control changes in COX-2 expression and PGE2 release, respectively. Additionally, chol integration into E-prostanoid (EP) receptor expressing LRs can alter signaling that regulates transepithelial Na absorption. In PRELIMINARY STUDIES an anti-diuretic, hypertensive phenotype was uncovered in chol fed mice, volume expanded by parenteral saline, that was associated with suppressed flow-mediated COX-2 activity and PGE2 release in microdissected cortical CDs (CCDs). Thus, the final integrated epithelial response to tubular flow rate is determined by chol calibrated, FSS-mediated PGE2 release and EP receptor stimulation which regulate Na transport. We hypothesize that hypercholesterolemia alters the chol content in LRs of the renal CD to regulate flow/FSS-induced COX-2 expression and PGE2 release which, via autocrine/paracrine EP- receptor based signaling, influences Na transport in the distal nephron. This hypothesis will be tested in the following specific aims (SAs): SA1. Test whether chol-rich LRs are mechanosensory platforms for FSS-mediated stimulation of polycystin-2 (PC-2) and MAPK activation. SA1.a Evaluate the localization of PC-2, a mechano-sensitive channel responsible for apical entry of Ca2+ into CD cells, and MAPK (p38, ERK) within specific populations of LRs in CD cells and microdissected CCDs. SA1.b Determine whether FSS leads to translocation of PC-2 and MAPK out of their respective chol-rich LRs to allow for maximal PC-2 activation and MAPK phosphorylation. SA1.c Test whether LR chol content affects resting localization and FSS-induced translocation of PC-2 and MAPK out of LRs. SA2. Test whether short-term chol ingestion leads to incorporation of chol into the CCD that alters flow-mediated COX stimulation, PGE2 release and Na excretion. SA2.a Evaluate whether dietary chol consumption incorporates into the renal CCD SA2.b Determine whether chol integration into renal CCDs modifies net PGE2 generating capacity, flow- mediated PGE2 release, Na excretion and, in turn, BP. SA3. Test whether short-term chol ingestion inhibits flow-mediated PGE2 release and alters EP receptor dependent signaling, to enhance transepithelial Na transport in microperfused CCDs SA3.a Evaluate whether chol incorporation in CCDs suppresses flow-mediated PGE2 release to augment Na absorption. SA3.b Test whether chol incorporation in CCDs modifies EP receptor signaling to enhance Na absorption.

 描述（由申请人提供）： 机械力，特别是流体剪切应力（FSS），在内皮细胞调节血管反应性和在肾上皮细胞影响跨上皮阳离子转运中起关键作用，所述跨上皮阳离子转运共同决定血压（BP）。肾小管阳离子转运的机械调节是肾钠（Na）稳态的重要调节器，并推断肾上皮细胞中存在机械传感器。富含胆固醇的脂筏（LRs）是位于质膜（PM）上的信号平台，是诱导内皮细胞旁分泌介导的血管舒张（即一氧化氮合成）的机械感受器;然而，关于它们在血管内皮细胞中的感觉功能知之甚少。 肾上皮Chol维持LR的完整性，并且对LR chol含量的影响影响FSS介导的信号传导。调节细胞中丝裂原活化蛋白激酶（MAPK）和细胞内Ca 2+浓度（[Ca 2 +]i）的FSS敏感性信号传导途径（1）分别定位于小窝蛋白-1（cav-1）和漂浮蛋白-2（flot-2），在集合管（CD）细胞中表达LR，和（2）分别控制考克斯-2表达和PGE 2释放的变化。此外，胆固醇整合到表达LR的E-前列腺素（EP）受体中可以改变调节跨上皮Na吸收的信号传导。在初步研究中，在胆固醇喂养的小鼠中发现了抗利尿、高血压表型，通过胃肠外盐水扩大体积，这与微切割皮质CD（CCD）中抑制的流量介导的考克斯-2活性和PGE 2释放相关。因此，最终的综合上皮细胞对肾小管流速的反应是由胆固醇校准，FSS介导的PGE 2释放和EP受体刺激，调节钠转运。我们假设高胆固醇血症改变了肾脏CD LR中的胆固醇含量，以调节流量/FSS诱导的考克斯-2表达和PGE 2释放，其通过基于自分泌/旁分泌EP受体的信号传导影响远端肾单位中的Na转运。这一假设将在以下特定目标（SA）中进行检验：SA 1。测试富含胆碱的LR是否是FSS介导的多囊蛋白-2（PC-2）刺激和MAPK激活的机械感觉平台。SA1.a评价PC-2（一种负责Ca 2+顶端进入CD细胞的机械敏感性通道）和MAPK（p38，ERK）在CD细胞和显微切割CCD中LR特定群体内的定位。SA1.b确定FSS是否导致PC-2和MAPK从其各自的富含chol的LR易位，以允许最大的PC-2活化和MAPK磷酸化。SA1.c测试LR胆固醇含量是否影响PC-2和MAPK的静息定位和FSS诱导的移位出LR。SA 2.检测短期摄入chol是否导致chol掺入CCD，从而改变血流介导的考克斯刺激、PGE 2释放和Na排泄。SA 2.a评价饮食中的胆固醇消耗是否并入肾CCD SA 2.b确定胆固醇并入肾CCD是否改变净PGE 2产生能力、流动介导的PGE 2释放、Na排泄，进而改变BP。SA 3.测试短期chol摄入是否抑制血流介导的PGE 2释放并改变EP受体依赖性信号传导，以增强微灌注CCD中的跨上皮Na转运SA 3.a评价CCD中的chol掺入是否抑制血流介导的PGE 2释放以增加Na吸收。SA3.b测试CCD中的chol掺入是否改变EP受体信号传导以增强Na吸收。