Significance of Epac signaling in renal Na+ handling and hypertension

项目摘要 肾脏在水盐平衡的调节中起着核心作用，过度的肾脏Na+保护被紧密地结合在一起。 与高血压有关。现有的利尿剂抑制单个肾小管节段中的Na+重吸收， 这通常涉及其他部分的补偿反应，以限制其效率。因此，针对Na+ 同时在多个部分运输系统可能是一种更有效的方式， 高血压我们的研究小组产生了大量的多组分证据， 由cAMP（Epac）激活的亚型1和2是近端小管中肾脏Na+处理的关键调节剂 (PT)和收集管（CD）。Epac 1和Epac 2缺失损害小鼠肾脏Na+保护， 导致饮食Na+限制期间血压降低。这与活动减少有关， PT中钠氢交换器-3（NHE-3）和CD中上皮Na+通道（ENaC）的表达。 此外，RNAseq基因本体富集分析揭示了线粒体功能的改善 以及Epac缺失后PT和CD中活性氧（ROS）产生的减少。有趣的是， 高血压时肾脏Epac表达急剧增加，这是Epac有害分支的争论 过度激活驱动Na+潴留和血压升高的发展。为此我们 开发了新的药理学工具来选择性地抑制Epac亚型，并观察到 这些小分子同时表现出低毒性和有效的生物利用度特征。总的来说，我们假设 Epac 1和Epac 2亚型是PT和PT中钠重吸收的生理相关调节剂， CD对低血容量的反应。相反，Epac级联的过度激活显著有助于 肾钠潴留和高血压的发展，部分是通过增加ROS的产生和氧化 应力我们认为，药物抑制Epac信号可能是一种新的，有效的，安全的策略 对抗高血压和改善肾功能我们计划通过三个具体目标来测试这一点： SA 1.确定与Epac诱导的小鼠肾小管转运变化相关的盐敏感性 缺乏Epac同种型以确定Epac信号传导的肾和肾外组分的作用。SA 2. 研究Epac依赖性调节Na+转运的分子靶点和信号机制 在PT和CD中。SA 3.为了验证Epac 1和Epac 2是有效的治疗靶点的假设， 使用优化的Epac特异性抑制剂治疗高血压。 总之，我们通过合并肾脏领域高度互补和协同的专业知识， 生理学/上皮转运和Epac信号传导/药物发现，来自Dr. Pochynyuk和郑博士在休斯顿UTHSC综合生物学和药理学系。 我们期望发现以前未被认识到的肾脏中Epac信号传导的生理相关手段。 建立Epac阻滞剂治疗高血压的药理学潜力。

PROJECT SUMMARY Kidneys play a central role in regulation of water-salt balance with excessive renal Na+ conservation being tightly associated with hypertension. The existing diuretics inhibit Na+ reabsorption in individual renal tubule segments, which often involves a compensatory response in other segments to limit their efficiency. Thus, targeting Na+ transporting systems in multiple segments simultaneously might represent a more effective way to fight hypertension. Our groups generated abundant multicomponent evidence that exchange protein directly activated by cAMP (Epac) isoforms 1 and 2 are critical regulators of renal Na+ handling in the proximal tubule (PT) and the collecting duct (CD). Epac1 and Epac2 deletion compromises renal Na+ conservation in mice, leading to reduced blood pressure during dietary Na+ restriction. This is associated with decreased activity and expression of the sodium hydrogen exchanger-3 (NHE-3) in PT and epithelial Na+ channel (ENaC) in CD. Furthermore, RNAseq Gene Ontology enrichment analysis revealed an improvement of mitochondria function and reduction in the reactive oxygen species (ROS) production in PT and CD upon Epac deletion. Interestingly, renal Epac expression is drastically increased during hypertension arguing for deleterious ramifications of Epac over-activation in driving Na+ retention and the development of elevated blood pressure. To this end, we have developed novel pharmacological tools to selectively inhibit Epac isoform(s), and observed natriuretic actions of these small molecules while exhibiting low toxicity and potent bioavailability profiles. Overall, we hypothesize that Epac1 and Epac2 isoforms are physiologically relevant regulators of sodium reabsorption in both PT and CD in response to hypovolemia. On the contrary, over-activation of Epac cascade contributes significantly to the development of renal sodium retention and hypertension, in part by increasing ROS production and oxidative stress. We propose that pharmacological inhibition of Epac signaling could be a novel, potent, and safe strategy to counteract hypertension and improve renal function. We plan to test this with 3 specific aims: SA1. Determine salt-sensitivity associated with Epac-induced changes on tubular transport in mice lacking Epac isoforms to establish roles of renal and extra-renal components of Epac signaling. SA2. Examine the molecular targets and signaling mechanisms of Epac-dependent regulation of Na+ transport in the PT and CD. SA3. To test the hypothesis that Epac1 and 2 are effective therapeutic targets of hypertension using optimized Epac specific inhibitors. In summary, we develop this proposal by merging highly complementary and synergistic expertise in renal physiology/epithelial transport and Epac signaling/drug discovery from neighboring laboratories of Dr. Pochynyuk and Dr. Cheng within the Department of Integrative Biology and Pharmacology, UTHSC at Houston. We anticipate to uncover previously unrecognized physiologically relevant means of Epac signaling in the kidney and to establish the pharmacological potential of Epac blockade in treatment of hypertension.