Mechanisms and relevance of ENaC regulation by EGF and Rac1

EGF 和 Rac1 调节 ENaC 的机制和相关性

• 批准号: 8245462
• 负责人: ALEXANDER STARUSCHENKO
• 金额: $ 38.25万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-12-01 至 2016-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8245462
• 关键词: Address; Aldosterone; Binding; Biochemical; Blood Pressure; Blood Volume; Cardiovascular Diseases; Cardiovascular system; Cell membrane; Cells; Chronic; Coupling; Dahl Hypertensive Rats; Data; Development; Diet; Disease; Distal; Electrolytes; Ensure; Environment; Epidermal Growth Factor; Family; Family member; Fluid Balance; Functional disorder; Growth Factor; Homeostasis; Hypertension; In Vitro; Ion Channel; Kidney; Kidney Diseases; Laboratories; Lead; Ligands; Liquid substance; Lung; Mediating; Microscopy; Molecular; Monomeric GTP-Binding Proteins; Nephrons; Pathology; Pathway interactions; Physiological; Physiology; Play; Proteins; Protocols documentation; Publishing; Rat Strains; Rattus; Recycling; Regulation; Role; Signal Pathway; Signal Transduction; Sodium; Sodium Chloride; Specificity; Up-Regulation; WAVE protein; Wisconsin; Work; base; blood pressure regulation; design; epithelial Na+ channel; genetic regulatory protein; hemodynamics; in vivo; inhibitor/antagonist; insight; instrument; medical schools; member; overexpression; receptor; research study; response; rhoB p20 GDI; salt sensitive; small hairpin RNA; trafficking

DESCRIPTION (provided by applicant): Long term control of blood pressure involves Na+ homeostasis through the precise regulation of the Epithelial Na+ Channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN). ENaC dysfunction is causative for disturbances in total body Na+ levels associated with abnormal regulation of blood volume, blood pressure, and lung fluid balance. We provide preliminary evidence that epidermal growth factor (EGF) can serve as an ENaC ligand and hypothesize that members of the EGF-family and Rac1 modulate ENaC-mediated Na+ transport in the ASDN and participate in the development of salt-sensitive hypertension. EGF and its related EGF-family members bind to ErbB receptors and act as signaling factors responsible for renal development, physiology and pathophysiology. Under physiological conditions, ErbB receptors play an important role in the regulation of renal hemodynamics and electrolyte handling by the kidney, while in different pathophysiological states ErbB activation may mediate either beneficial or detrimental effects on the kidney. Stimulation of ErbB receptors activates an intracellular cascade involving small GTPases, particularly Rac1. Small G proteins and their regulatory proteins contribute to the pathology of renal and cardiovascular diseases. Our preliminary results, including electrophysiological experiments using isolated, split open ASDN, demonstrate that ENaC is regulated by EGF and Rac1, possibly through a convergent mechanism. Dahl salt-sensitive (SS) rats used in these studies develop severe hypertension on high-salt diet. We provide preliminary data indicating that ENaC contributes to the development of hypertension in the SS rat strain. Furthermore, our preliminary data reveal that EGF concentration is reduced in the SS rats, which we propose would enhance ENaC activity, sodium retention and hypertension. Building upon this preliminary data and our previously published findings, the specific objective of this proposal is to determine whether EGF acting through Rac1 is important for physiologic control of renal sodium handling through regulation of ENaC and define the precise mechanisms of EGF- and Rac1-mediated changes in ENaC activity. A combination of electrophysiological, immunohistochemical, biochemical, microscopy and chronic studies in vivo and in vitro will be used in this proposal to provide mechanistic insights on how ENaC is regulated by member of the EGF-family and Rac1 and how changes in this pathway contributes to salt induced hypertension in SS rats. These studies will address three Specific Aims: 1) To identify and quantify the role of EGF and related growth factors in the physiological regulation of ENaC activity in the ASDN and establish the role of this pathway in the development of salt-sensitive hypertension; 2) To establish the physiological role of RhoGDI and Rac1 in regulation of ENaC and determine the role of Rac1 in mediating EGF effects on ENaC; and 3) To define the cellular and molecular mechanism by which Rac1 modulates ENaC activity: do WAVEs convey Rac1 regulation to ENaC? PUBLIC HEALTH RELEVANCE: The control of blood pressure occurs via Na+ homeostasis in the kidney and involves the precise regulation of the Epithelial Na+ Channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN). The current proposal will study how members of the epidermal growth factors family and small GTPase Rac1 modulate ENaC- mediated Na+ transport in the ASDN and participate in the development of salt-sensitive hypertension. This work has the potential to provide new insights into the control and function of this important ion channel and uncover mechanisms involved in diseases associated with fluid imbalance and hypertension.

描述（由申请人提供）：血压的长期控制涉及通过精确调节醛固酮敏感的远端肾单位 (ASDN) 中的上皮 Na+ 通道 (ENaC) 来实现 Na+ 稳态。 ENaC 功能障碍会导致全身 Na+ 水平紊乱，并与血容量、血压和肺液平衡调节异常相关。我们提供了表皮生长因子（EGF）可以作为 ENaC 配体的初步证据，并假设 EGF 家族成员和 Rac1 调节 ASDN 中 ENaC 介导的 Na+ 转运，并参与盐敏感性高血压的发展。 EGF 及其相关 EGF 家族成员与 ErbB 受体结合，并充当负责肾脏发育、生理学和病理生理学的信号因子。在生理条件下，ErbB 受体在肾脏血流动力学和电解质处理的调节中发挥重要作用，而在不同的病理生理状态下，ErbB 激活可能介导对肾脏的有益或有害影响。 ErbB 受体的刺激会激活涉及小 GTP 酶（尤其是 Rac1）的细胞内级联反应。小 G 蛋白及其调节蛋白有助于肾脏和心血管疾病的病理学。我们的初步结果，包括使用分离的、分裂开放的 ASDN 进行的电生理学实验，表明 ENaC 可能通过收敛机制受到 EGF 和 Rac1 的调节。这些研究中使用的 Dahl 盐敏感 (SS) 大鼠在高盐饮食下会出现严重的高血压。我们提供的初步数据表明 ENaC 有助于 SS 大鼠品系中高血压的发生。此外，我们的初步数据显示 SS 大鼠中 EGF 浓度降低，我们认为这会增强 ENaC 活性、钠潴留和高血压。基于这些初步数据和我们之前发表的研究结果，本提案的具体目标是确定 EGF 通过 Rac1 发挥作用是否对于通过调节 ENaC 对肾脏钠处理的生理控制很重要，并确定 EGF 和 Rac1 介导的 ENaC 活性变化的精确机制。该提案将结合电生理学、免疫组织化学、生化、显微镜和体内外长期研究，以提供关于 EGF 家族成员和 Rac1 如何调节 ENaC 以及该途径的变化如何导致 SS 大鼠盐诱导高血压的机制见解。这些研究将解决三个具体目标：1）确定和量化 EGF 和相关生长因子在 ASDN 中 ENaC 活性生理调节中的作用，并确定该途径在盐敏感性高血压发展中的作用； 2) 建立RhoGDI和Rac1在ENaC调节中的生理作用，并确定Rac1在介导EGF对ENaC影响中的作用； 3) 定义 Rac1 调节 ENaC 活性的细胞和分子机制：WAVE 是否将 Rac1 调节传递给 ENaC？ 公共卫生相关性：血压的控制是通过肾脏中的 Na+ 稳态进行的，涉及醛固酮敏感的远端肾单位 (ASDN) 中上皮 Na+ 通道 (ENaC) 的精确调节。目前的提案将研究表皮生长因子家族的成员和小 GTPase Rac1 如何调节 ASDN 中 ENaC 介导的 Na+ 转运并参与盐敏感性高血压的发展。这项工作有可能为这一重要离子通道的控制和功能提供新的见解，并揭示与体液失衡和高血压相关疾病的机制。