PKD proteins in endothelial cells

内皮细胞中的 PKD 蛋白

• 批准号: 10339327
• 负责人: Jonathan H Jaggar
• 金额: $ 60.23万
• 依托单位: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10339327
• 关键词: Acetylcholine; Arteries; Attenuated; Biotinylation; Blood Pressure; Blood Vessels; Blood flow; Calcium; Cardiovascular Diseases; Cell Line; Cell Wall; Cell physiology; Coupling; Data; Electrophysiology (science); Endothelial Cells; Endothelium; Fluorescence Resonance Energy Transfer; Genetic Models; Hypertension; Image; Immunofluorescence Immunologic; Impairment; Knock-out; Knockout Mice; Ligands; Light; Lighting; Mediating; Membrane Potentials; Methods; Microscope; Modification; Mus; Myography; Nitric Oxide Synthase; Organ; PKD2 protein; Pathologic; Pathway interactions; Pharmacology; Physiological; Potassium Channel; Process; Proteins; RNA Interference; Receptor, Angiotensin, Type 1; Regulation; Resistance; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Stimulus; Superoxide Dismutase; Systemic blood pressure; TRP channel; Telemetry; Tertiary Protein Structure; Testing; Transmembrane Domain; Vasodilation; Vasodilator Agents; Western Blotting; Wnt proteins; autocrine; cell type; mimetics; novel; patch clamp; polycystic kidney disease 1 protein; protein kinase D; protein phosphatase inhibitor-2; response

Project Summary Endothelial cells line the wall of all blood vessels and regulate a wide variety of functions, including contractility which controls systemic blood pressure. Dysfunctional endothelial cells are a hallmark of several cardiovascular diseases, but pathological mechanisms involved are poorly understood. Endothelial cells express both PKD1 (polycystin-1), an eleven transmembrane domain protein, and PKD2 (polycystin-2), a transient receptor potential (TRP) channel. Regulatory mechanisms, physiological functions and pathological involvement during hypertension of PKD1 protein and PKD2 channels in endothelial cells are unclear. Using a wide variety of approaches and inducible, endothelial cell-specific knockout mice, we provide evidence that physical coupling of PKD1 to PKD2 in endothelial cells stimulates vasodilation. Preliminary data also suggest that PKD1/PKD2 channel signaling is dysfunctional during hypertension, which attenuates this vasodilatory signaling mechanism. In this proposal, we will investigate three specific aims. Aim 1 will test the hypothesis that endothelial-dependent physiological stimuli activate PKD1/PKD2 coupling in endothelial cells, leading to vasodilation. Aim 2 will investigate the mechanisms by which endothelial-dependent stimuli activate PKD1/PKD2 channels in endothelial cells to produce vasodilation. Aim 3 will study the hypothesis that hypertension is associated with pathological alterations in PKD1/PKD2 channel signaling in endothelial cells that inhibits vasodilation mediated by these proteins. Methods used will include RT-PCR, Western blotting, biotinylation, FRET, RNAi, co-IP, immunofluorescence, patch-clamp electrophysiology, membrane potential recording, intracellular Ca2+ imaging, arterial myography and blood pressure telemetry. This project will provide significant novel information concerning vasoregulation by endothelial cell PKD1 and PKD2 proteins.

项目摘要 内皮细胞排列在所有血管的壁上，调节多种功能，包括收缩功能。 它控制全身血压。功能障碍的内皮细胞是几个 心血管疾病，但涉及的病理机制知之甚少。内皮细胞 同时表达11个跨膜区蛋白PKD1(Polycystin-1)和PKD2(Polycystin-2 瞬时受体电位(Trp)通道。调控机制、生理功能与病理 高血压时内皮细胞内的PKD1蛋白和PKD2通道的参与尚不清楚。使用 各种各样的方法和可诱导的内皮细胞特异性基因敲除小鼠，我们提供了证据 内皮细胞中PKD1和PKD2的物理偶联可刺激血管扩张。初步数据还表明， PKD1/PKD2通道信号在高血压期间功能失调，从而减弱了这种血管扩张作用 信令机制。在这项提案中，我们将调查三个具体目标。目标1将检验这一假设 内皮依赖的生理刺激激活内皮细胞中的PKD1/PKD2偶联，导致 血管扩张。目标2将研究内皮依赖刺激激活的机制 内皮细胞中的PKD1/PKD2通道产生血管扩张作用。目标3将研究假设 高血压与内皮细胞PKD1/PKD2通道信号的病理改变有关 这抑制了这些蛋白质介导的血管扩张。使用的方法包括RT-PCR，Western blotting， 生物素化，FRET，RNAi，共IP，免疫荧光，膜片钳电生理，膜电位 记录、细胞内钙成像、动脉肌图和血压遥测。这个项目将 提供关于内皮细胞PKD1和PKD2蛋白调节血管的重要新信息。