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）：髓质厚的上升肢（MTAL）对于调节盐和水稳态至关重要，并且会通过不同的单加氧酶进行调节，从而代谢蛛网膜酸。在受血管紧张素II（ANG II）挑战的MTAL细胞中以及钙感应受体（CAR）激活后，肿瘤坏死因子-Alpha（TNF）的产生增加，并且这些激动剂抑制MTAL在TNF-和环氧酶-2（COX-2）中的MTAL中的离子转运。因此，将分别在MTAL小管和细胞中确定TNF对影响顶端K+通道和Na+-K+-2CL-共转运蛋白活性的前类前素依赖机制的贡献。细胞将从野生型窝窝对照（TNF+/+）和TNF缺陷小鼠中分离出来。这些小鼠还将用于体内实验，旨在确定TNF在ANG II依赖性高血压模型中使用Radiotelemetry确定的TNF对血压调节的贡献。 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确定。我们的实验室最近提供了第一个证明汽车激活增加任何细胞类型中活化T细胞（NFAT）活性的核因子。因此，已经鉴定出存在于MTAL中的NFAT同工型，并将与CAR激活后MTAL细胞中TNF和PGE2的产生有关。几种细胞和分子方法，包括电动性转移测定，激光扫描细胞仪，具有显性负NFAT构建体的转染以及使用NFAT活性报告基因构建体的使用，可用于识别有助于CAR介导的TNF产生的同种型。 TNF对心血管功能的生物学特征表明，该细胞因子可以以上下文依赖性方式表现出促或抗高血压活性。例如，在MTAL细胞中，TNF及其与COX-2和PGE2表达的关系可能充当抗抑制器机制的一部分。因此，可以通过在TAL的基底外侧膜上激活CAR来激活这种机制，从而使COX-2衍生的PGE2通过TNF依赖性机制增加。我们的发现表明，由于PGE2对TAL中盐和水的抑制作用，导致血压降低的机制响应汽车激活。