Regulation of Renal TNF Production and Function

肾 TNF 产生和功能的调节

基本信息

批准号：
7558316
负责人：
NICHOLAS R FERRERI
金额：
$ 39.75万
依托单位：
NEW YORK MEDICAL COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-02-01 至 2012-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7558316
关键词：
Affect Agonist Angiotensin II Antihypertensive Agents Apical Arachidonic Acids Biological Biological Assay Blood Pressure Calcineurin Calcium-Sensing Receptors Cardiovascular Physiology Cell Nucleus Cells Chronic Consumption Cyclosporine DNA Sequence Analysis Data Development Diabetes Mellitus Dinoprostone Dominant-Negative Mutation Exhibits Fibrosis Genes Homeostasis Hypercalcemia Hypertension Ion Transport Kidney Kidney Neoplasms Laboratories Laser Scanning Cytometry Limb structure Link Lipopolysaccharides Mediating Mediator of activation protein Mixed Function Oxygenases Modeling Molecular Mus Na(+)-K(+)-Exchanging ATPase Natriuresis Nephrons Ouabain Potassium Channel Production Prostaglandins Protein Isoforms Rattus Receptor Activation Regulation Regulatory Pathway Renal Hypertension Renal function Reporter Reporting Reverse Transcriptase Polymerase Chain Reaction Sodium Chloride TNFRSF1A gene Testing Thick Time Transfection Tumor Necrosis Factor-alpha VIVIT peptide Water base basolateral membrane blood pressure regulation cell type cyclooxygenase 2 cytokine design extracellular in vivo kidney cortex kidney medulla novel nuclear factors of activated T-cells pressure receptor research study response urinary

项目摘要

DESCRIPTION (provided by applicant): The medullary thick ascending limb (mTAL) is critical to the regulation of salt and water homeostasis and subject to regulation via different monooxygenases that metabolize arachidonic acid. Tumor necrosis factor-alpha (TNF) production is increased in mTAL cells challenged with Angiotensin II (Ang II) and after activation of the calcium-sensing receptor (CaR), and these agonists inhibit ion transport in the mTAL in a TNF- and cyclooxygenase-2 (COX-2)-dependent manner. Thus, the contribution of TNF to prostanoid-dependent mechanisms affecting the regulation of apical K+ channels and Na+-K+-2Cl- cotransporter activity will be determined in mTAL tubules and cells, respectively. Cell will be isolated from wild type littermate control (TNF+/+) and TNF deficient mice. These mice also will be used for in vivo experiments designed to determine the contribution of TNF to the regulation of blood pressure, determined using radiotelemetry, in an Ang II-dependent model of hypertension. The ability of Ang II to regulate COX-2 expression and activity in a TNF-dependent manner also will be determined in vivo. The first demonstration that CaR activation increases nuclear factor of activated T cell (NFAT) activity in any cell type was recently provided by our laboratory. Accordingly, NFAT isoforms present in the mTAL have been identified and will be linked to the production of TNF and PGE2 in mTAL cells after CaR activation. Several cellular and molecular approaches, including electromobility shift assays, laser scanning cytometry, transfection with dominant negative NFAT constructs, and use of NFAT activity reporter constructs, will be used to identify the isoforms that contribute to CaR-mediated TNF production. The biological profile of TNF on cardiovascular function indicates that this cytokine may exhibit either pro- or anti-hypertensive activities in a context-dependent manner. For instance, in mTAL cells, TNF and its relationship to expression of COX-2 and PGE2 may act as part of an antipressor mechanism. Thus, activation of this mechanism in response to extracellular Ca2+ may be initiated by activation of CaR on the basolateral membrane of the TAL, which increases COX-2- derived PGE2 synthesis via a TNF-dependent mechanism. Our findings indicate a mechanism responsible for blood pressure reduction in response to CaR activation by virtue of the inhibitory effect of PGE2 on salt and water transport in the TAL.

描述（由申请人提供）：髓质粗升肢（mTAL）对于盐和水稳态的调节至关重要，并受到代谢花生四烯酸的不同单加氧酶的调节。在用血管紧张素 II (Ang II) 攻击并激活钙敏感受体 (CaR) 后，mTAL 细胞中肿瘤坏死因子-α (TNF) 的产生增加，并且这些激动剂通过 TNF-α 和环氧合酶抑制 mTAL 中的离子转运-2 (COX-2)依赖方式。因此，TNF 对影响顶端 K+ 通道和 Na+-K+-2Cl- 协同转运蛋白活性调节的前列腺素依赖性机制的贡献将分别在 mTAL 小管和细胞中确定。细胞将从野生型同窝对照（TNF+/+）和TNF缺陷小鼠中分离。这些小鼠还将用于体内实验，旨在确定 TNF 对血压调节的贡献，在血管紧张素 II 依赖性高血压模型中使用无线电遥测技术确定。 Ang II 以 TNF 依赖性方式调节 COX-2 表达和活性的能力也将在体内测定。我们的实验室最近首次证明 CaR 激活会增加任何细胞类型中活化 T 细胞核因子 (NFAT) 的活性。因此，mTAL 中存在的 NFAT 亚型已被鉴定，并将与 CaR 激活后 mTAL 细胞中 TNF 和 PGE2 的产生相关。几种细胞和分子方法，包括电动迁移率测定、激光扫描细胞术、显性负性 NFAT 构建体转染以及 NFAT 活性报告构建体的使用，将用于鉴定有助于 CaR 介导的 TNF 产生的亚型。 TNF 对心血管功能的生物学特征表明，这种细胞因子可能以背景依赖性方式表现出促高血压或抗高血压活性。例如，在 mTAL 细胞中，TNF 及其与 COX-2 和 PGE2 表达的关系可能是抗抑郁机制的一部分。因此，该机制响应细胞外 Ca2+ 的激活可能是通过激活 TAL 基底外侧膜上的 CaR 来启动的，这通过 TNF 依赖性机制增加了 COX-2 衍生的 PGE2 合成。我们的研究结果表明，由于 PGE2 对 TAL 中盐和水转运的抑制作用，CaR 激活导致血压降低。