Thick ascending limb-derived TNF, salt sensitivity, and blood pressure regulation

厚升肢源性 TNF、盐敏感性和血压调节

• 批准号: 9306934
• 负责人: NICHOLAS R FERRERI
• 金额: $ 41万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9306934
• 关键词: Affect; Affinity; Antihypertensive Agents; Apical; Attenuated; Binding Proteins; Blood Pressure; Blood Vessels; Cardiovascular Diseases; Cell Nucleus; Cells; Characteristics; Cre-LoxP; Cytoplasm; Cytosol; Development; Distal; Equilibrium; Excretory function; Exhibits; Feedback; Genetic; Genetic Transcription; Glycoproteins; Goals; Homeostasis; Hypertension; Individual; Inflammation; Inflammatory; Intake; Kidney; Knockout Mice; Limb structure; Maintenance; Messenger RNA; Modeling; Mus; Myocardial Infarction; Nephrons; Nuclear Translocation; Pathway interactions; Perfusion; Phosphorylation; Physiological; Production; Protein Isoforms; Regulation; Regulatory Pathway; Renal function; Risk Factors; Role; Signal Pathway; Site; Sodium; Sodium Chloride; Stimulus; Stroke; Structure of ascending limb of Henle' s loop; Subfamily lentivirinae; System; TNF gene; Testing; Thick; Transgenic Mice; Tubular formation; Tumor-Derived; autocrine; base; blood pressure regulation; cell type; cytokine; hypertension treatment; in vivo; insight; mRNA Stability; normotensive; novel; nuclear factors of activated T-cells; promoter; response; salt intake; salt sensitive hypertension; targeted treatment; transcription factor; urinary

The central hypothesis of this project is that tumor necrosis factor-alpha (TNF) is produced by the thick ascending limb of Henle's loop (TAL) and is part of an adaptive pathway that attenuates increases in blood pressure in response to high NaCl intake. The TAL-derived TNF inhibits Na+-K+-2Cl- cotransporter (NKCC2) expression and activity and is produced in response to high NaCl intake via a transcriptional mechanism involving NFAT5, also known as tonicity-responsive enchancer binding protein (TonEBP). Thus, TNF production by the TAL in response to high salt intake serves as a negative feedback mechanism to attenuate NKCC2 expression and activity, thereby contributing to the maintenance of blood pressure by limiting an increase in NaCl reabsorption. The maintenance of sodium homeostasis is essential for the regulation of blood pressure and insights regarding novel mechanisms that regulate renal sodium transporters could serve as a framework for development of new antihypertensive therapies. Three main objectives will pursued during this study: 1) Determine the effects of TAL-derived TNF on the blood pressure and renal functional responses to HS intake under normotensive/non-inflammatory conditions, 2) Define the mechanisms by which TNF inhibits NKCC2 expression and activity in the mTAL, and 3) Determine the role of NFAT5 as a regulator of the TAL- derived TNF production and responses to HS intake. TAL cell-specific silencing of TNF will be accomplished using purified lentivirus constructs under the control of a Tamm-Horsfall glycoprotein (THP)-specific promoter construct and by a transgenic mouse strategy to genetically delete TNF in the TAL using the Cre/loxP approach. These models will be used to evaluate the effects of TNF derived from the TAL on blood pressure, renal function and NKCC2 expression and activity under normal and high NaCl conditions. The approaches to study NKCC2 regulation by TNF include in vivo studies in parallel with freshly isolated mTAL tubules, which will be used for cellular determinations of NKCC2 phosphorylation, activity, and analysis of the signaling pathways that are inhibited by TNF. Primary cultures of polarized mouse mTAL cells also will be used to study the mechanisms by which TNF inhibits NKCC2 phosphorylation. TAL-specific lentivirus silencing of NFAT isoforms in vivo will be used to determine the mechanism by which TNF is produced in this segment of the nephron and the expression and translocation from NFAT isoforms from cytoplasm to nucleus will be determined in mTAL tubules and cells. This project will advance our understanding of a novel intrarenal regulatory pathway involving TNF, which contributes to blood pressure homeostasis in response to high salt intake.

该项目的中心假设是肿瘤坏死因子-α（TNF）是由厚膜产生的 亨利氏环（TAL）的上升支，是一个适应性途径的一部分， 压力响应高NaCl摄入量。TAL衍生的TNF抑制Na+-K+-2Cl-协同转运蛋白（NKCC 2） 表达和活性，并通过转录机制响应高NaCl摄入而产生 涉及NFAT 5，也称为张力反应性增强剂结合蛋白（TonEBP）。因此，TNF TAL响应于高盐摄入的产生充当负反馈机制，以减弱 NKCC 2的表达和活性，从而通过限制高血压的发生而有助于维持血压。 增加NaCl重吸收。维持钠稳态对于调节血液是必不可少的 压力和关于调节肾钠转运蛋白的新机制的见解可以作为一个新的研究方向。 开发新的抗高血压疗法的框架。在此期间将实现三个主要目标。 研究：1）确定TAL衍生的TNF对血压和肾功能反应的影响， 在血压正常/非炎症条件下摄入HS，2）定义TNF抑制 mTAL中NKCC 2的表达和活性，以及3）确定NFAT 5作为TAL调节因子的作用- 衍生的TNF产生和对HS摄入的反应。将实现TNF的TAL细胞特异性沉默 使用在Tamm-Horsfall糖蛋白（THP）特异性启动子控制下的纯化慢病毒构建体 构建并通过转基因小鼠策略使用Cre/loxP在TAL中遗传缺失TNF approach.这些模型将用于评价TAL衍生的TNF对血压的影响， 正常和高NaCl条件下的肾功能和NKCC 2表达和活性。的途径 TNF对NKCC 2调节的研究包括与新鲜分离的mTAL小管平行的体内研究， 用于NKCC 2磷酸化、活性的细胞测定和信号通路的分析 被肿瘤坏死因子抑制。极化的小鼠mTAL细胞的原代培养物也将用于研究极化的小鼠mTAL细胞。 TNF抑制NKCC 2磷酸化的机制。NFAT的TAL特异性慢病毒沉默 将使用体内同种型来确定TNF在该节段中产生的机制。 肾单位和NFAT亚型的表达和从细胞质到细胞核的易位将是可能的。 在mTAL小管和细胞中测定。该项目将推进我们对一种新的肾内 涉及TNF的调节途径，其有助于响应高盐的血压稳态 摄入