Mitochondria-mediated effects and therapeutic potential of Atrial Natriuretic Peptide in salt-sensitive hypertension

项目总结 目前尚无针对血压盐敏感型患者的特效药。 不幸的是，盐敏感性的分子机制仍然知之甚少。其中一个 盐敏感型(SS)高血压的主要发病机制涉及 肾脏排盐的能力。心钠素(ANP)是由NPPA编码的一种激素。 促进盐排泄和降低血压，有临床数据表明心钠素水平天生较低 SS高血压的发病情况。在其他影响中，已知ANP(通过cGMP相关机制)是 有利于线粒体生物能量学和生物发生。然而，关于这方面的知识存在差距。 心钠素对肾脏线粒体的影响，特别是对SS高血压的影响。我们的初步研究表明 在高盐攻击期间，NPPA-/-(ANP基因敲除)Dahl SS大鼠表现出更严重的盐敏感性， 减少钠排泄，加重肾脏损伤，这与线粒体损伤和 功能障碍。我们还发现，在NPPA-/-大鼠中，存在着肾脏钠转运体的失调。 与野生型对照相比，采集过程中上皮钠通道(ENaC)的活性升高 风管。慢性注射ANP对盐诱导的SS大鼠血压升高的影响 并减轻器官损伤。 我们假设在SS高血压患者中，ANP缺乏/对ANP敏感性降低是导致肾脏的原因 线粒体功能障碍和相关的钠转运失衡。为了解决这一中心假设 项目中，我们制定了三个具体目标：目标1.确定增加ANP水平是否有益于 SS高血压患者肾盐处理与心功能的关系目的2.确定低水平的肾脏cGMP是否 由于缺乏ANP，导致肾脏线粒体钙和活性氧自由基(ROS)增加。 目的3.验证功能失调导致钙平衡紊乱和ROS过度产生的假说 线粒体影响SS高血压患者的肾脏钠转运。 我们产生了大量的证据来支持这些目标，创造了一个严格而全面的实验 设计并建立了新的尖端技术来解决这一假设。我们招募了斯特朗 协作专业知识，并将实施整体动物研究和活体技术(血液)的组合 通过药物输注、代谢研究和GFR测量进行压力监测)、电生理学(单一 新分离的肾单位和线粒体的通道和全细胞膜片钳)，高级 显微镜、线粒体荧光测定法和呼吸测定法，以及常规分子生物学方法。这个 拟议研究的成功完成将揭开盐敏感性的新的致病机制。

PROJECT SUMMARY There is no specific treatment available for the subpopulation of patients with salt sensitivity of blood pressure (BP); unfortunately, the molecular mechanisms underlying salt-sensitivity remain poorly understood. One of the major proposed mechanisms for the development of salt-sensitive (SS) hypertension involves a defect in the ability of the kidneys to excrete salt. Atrial Natriuretic Peptide (ANP) encoded by Nppa, is a hormone known to promote salt excretion and BP reduction, and there are clinical data implicating inherently low levels of ANP in the development of SS hypertension. Among other effects, ANP (via cGMP-related mechanisms) is known to be beneficial for mitochondrial bioenergetics and biogenesis. However, there is a gap in knowledge regarding the effects of ANP on mitochondria in the kidney, especially in SS hypertension. Our pilot studies demonstrated that during a high salt challenge Nppa-/- (ANP knockout) Dahl SS rats exhibit exacerbated salt-sensitivity, reduced sodium excretion, and aggravated kidney injury, which is associated with mitochondrial damage and dysfunction. We also showed that there is dysregulation of renal sodium transporters, in the Nppa-/- rats compared to wild-type controls, and the activity of the Epithelial Na+ Channel (ENaC) is elevated in the collecting ducts. Chronic ANP infusion in wild-type SS rats resulted in a dramatic attenuation of salt-induced BP increase and alleviated organ damage. We hypothesize that in SS hypertension ANP deficiency/reduced sensitivity to ANP is causative to renal mitochondrial dysfunction and associated sodium transport imbalance. To address the central hypothesis of this project, we developed three specific aims: Aim 1. Establish whether increased ANP levels are beneficial for renal salt handling and cardiac function in SS hypertension. Aim 2. Determine whether low renal cGMP level resulting from lack of ANP causes an increase in renal mitochondrial Ca2+ and reactive oxygen species (ROS). Aim 3. Test the hypothesis that disrupted Ca2+ balance and excessive ROS production by dysfunctional mitochondria affect renal sodium handling in SS hypertension. We generated abundant evidence to support these aims, created a rigorous and comprehensive experimental design and established novel cutting-edge techniques to address the hypothesis. We recruited strong collaborative expertise, and will implement a combination of whole-animal studies and in vivo techniques (blood pressure monitoring with drug infusion, metabolic studies and GFR measurements), electrophysiology (single channel and whole-cell patch-clamp of the freshly isolated nephrons and isolated mitochondria), advanced microscopy, mitochondrial spectrofluorimetry and respirometry, and routine molecular biology approaches. The successful completion of the proposed studies will unravel the novel causative mechanisms of salt-sensitivity.