Role of Cyp2j-epoxygenases, sEH and PPARs in adenosine-induced vascular response

Cyp2j-环氧合酶、sEH 和 PPAR 在腺苷诱导的血管反应中的作用

• 批准号: 8666036
• 负责人: Mohammed A Nayeem
• 金额: $ 36.97万
• 依托单位: WEST VIRGINIA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8666036
• 关键词: ADORA2A gene; Acetylcholine; Adenosine; Adenosine A1 Receptor; Adenylate Cyclase; Affect; Agonist; Antiinflammatory Effect; Aorta; Bathing; Blood Pressure; Blood Vessels; Cardiovascular Diseases; Cell Line; Cell physiology; Clinical; Coronary artery; Country; Coupling; Cyclic AMP-Dependent Protein Kinases; Data; Development; Down-Regulation; Endothelial Cells; Endothelium; Epoxide hydrolase; Exercise; Exhibits; Experimental Designs; GTP-Binding Proteins; Genes; Genetic Polymorphism; Goals; Health; Human; Hypertension; Image; Individual; Kidney; Link; Measures; Mediating; Mitogen-Activated Protein Kinase Inhibitor; Mus; Myocardial perfusion; Organ; Outcome; PKA inhibitor; PPAR gamma; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacology; Population; Prevention; Protein Kinase C Alpha; Proteins; Publishing; Purinergic P1 Receptors; Receptor Gene; Relaxation; Reporting; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Transduction; Smooth Muscle Myocytes; Technology; Testing; Therapeutic Uses; Transgenic Mice; Vascular Endothelium; Vasodilation; Western Blotting; Wild Type Mouse; base; blood pressure regulation; drinking water; endothelial dysfunction; genetic variant; inhibitor/antagonist; innovation; knockout gene; mRNA Expression; novel; overexpression; renal artery; response; tool; vasoconstriction

DESCRIPTION (provided by applicant): Endothelial dysfunction is associated with corresponding changes in vascular tone and increases in contractility, a condition that may cause hypertension in susceptible individuals which may have allelic variants in cyp2j-epoxygenases, soluble epoxide hydrolase (sEH), A2A, and A1 adenosine receptors (A2A AR & A1 AR) genes. These allelic variants may have similarities to our transgenic mice which may regulate vascular tone and blood pressure (BP). Our preliminary data have suggested a possible link between adenosine-induced relaxation and opening of KATP channels through A2A AR-cyp2j-PKA-PPARγ pathway. Also, it is possible that a link may exist between adenosine-induced contraction and closing of KATP channels through A1 AR-sEH-cyp4a- PPARα pathway. A combination of pharmacological tools and transgenic mice would allow us to identify the possible targets as a long term goal to treat population which may have allelic variants leading to hypertension. Therefore, there is a critical need to explore the possible mechanism involving cyp2j2-epoxygenases, sEH/cyp4a, A1 AR/A2A AR, PPARα/γ, PKA/PKCα/ϵ and KATP channels in adenosine-induced vascular responses. Our central hypothesis is that adenosine induces vascular relaxation and decreases in BP through cyp2j-epoxygenases via A2A AR-cyp2j-PKA-PPARγ signaling leading to opening of KATP channels. On the other hand, adenosine induces vascular contraction and increases in BP through sEH via A1 AR-sEH-cyp4a-PPARα pathway leading to closing of KATP channels. To test this hypothesis, we propose to explore in depth mechanism(s) using A2A AR-/-, A1 AR-/-, cyp2j5-/-, sEH-/-, Tie2-cyp2j2Tr (endothelial-cyp2j2 overexpressed), Tie2-sEHTr (endothelial-sEH overexpressed), wild-type mice, immortal renal endothelial cell line from H-2Kb- tsA58 mouse, mouse aortic endothelial cells (MAEC) and mouse aortic smooth muscle cells (MASMC). Further, we will also explore the possible treatment with sEH inhibitors (AUDA/t-AUCB) in drinking water (or gavage) for A2A AR-/-, cyp2j5-/- and Tie2-sEHTr mice which may have high BP. We will measure BP, and we will use aortas/renal arteries (organ bath/DMT-wire myograph) with treatments (adenosine-receptors agonists & antagonists), cyp-epoxygenases, sEH, adenylyl cyclase, PKCα/ϵ, MAPK and PKA inhibitors, PPARα/γ, EETs and KATP channel (activators & inhibitors). EETs & DHETs will be analyzed (UPLC-MS/MS). Western blot & RT-PCR will be used for proteins & mRNA expression. We propose 3 specific aims to determine: (1) whether the cyp2j-epxygenases or sEH affects BP, adenosine-induced vascular response and EETs/DHETs; (2) whether the presence or absence of A2A AR affects adenosine-induced vascular response through PPARs via cyp2j-epoxygenases, sEH (3) whether the presence /overexpression of cyp2j-epoxygenases or sEH regulate KATP channels through A2A AR-cyp2j-PKA-PPARγ/A1 AR-sEH-cyp4a-PPARα pathway in adenosine-induced vascular response. Such results can have a positive impact, as the identified components are expected to provide new targets to curb clinical problems linked with dysfunctional endothelium leading to hypertension.

描述（由申请人提供）：内皮功能障碍与血管张力的相应变化和收缩力的增加相关，这是一种可能导致易感个体高血压的病症，易感个体可能具有cyp 2 j-环氧合酶、可溶性环氧化物水解酶（sEH）、A2 A和A1腺苷受体（A2 A AR & A1 AR）基因的等位基因变体。这些等位基因变体可能与我们的转基因小鼠具有相似性，这些转基因小鼠可能调节血管张力和血压（BP）。我们的初步研究结果提示腺苷对KATP通道的舒张作用可能通过A2 A AR-cyp 2 j-PKA-PPARγ途径而与KATP通道的开放有关。腺苷引起的KATP通道收缩可能通过A1 AR-sEH-cyp 4a- PPARα途径与KATP通道关闭有关。药理学工具和转基因小鼠的组合将使我们能够确定可能的靶点，作为治疗可能具有导致高血压的等位基因变体的人群的长期目标。因此，研究cyp 2 j2-环氧合酶、sEH/cyp 4a、A1 AR/A2 A AR、PPARα/γ、PKA/PKCα/PKC β和KATP通道在腺苷诱导血管反应中的作用机制具有重要意义。我们的中心假设是腺苷通过cyp 2 j-环氧合酶诱导血管舒张并降低血压，通过A2 A AR-cyp 2 j-PKA-PPARγ信号传导导致KATP通道开放。另一方面，腺苷通过A1 AR-sEH-cyp 4a-PPAR α途径引起血管收缩和血压升高，导致KATP通道关闭。为了验证这一假设，我们提出使用A2 A AR-/-、A1 AR-/-、cyp 2 j5-/-、sEH-/-、Tie 2-cyp 2 j2 Tr（内皮细胞-cyp 2 j2过表达），Tie 2-sEHTr（内皮-sEH过表达），野生型小鼠，来自H-2Kb-tsA 58小鼠的永生肾内皮细胞系，小鼠主动脉内皮细胞（MAEC）和小鼠主动脉平滑肌细胞（MASMC）。此外，我们还将探索在饮用水（或灌胃）中使用sEH抑制剂（AUDA/t-AUCB）对可能具有高BP的A2 A AR-/-、cyp 2 j5-/-和Tie 2-sEHTr小鼠的可能治疗。我们将测量血压，我们将使用治疗（腺苷受体激动剂和拮抗剂）、cyp-epoxygenases、sEH、腺苷酸环化酶、PKCα/PKC β、MAPK和PKA抑制剂、PPARα/γ、EkB和KATP通道（激活剂和抑制剂）的动脉/肾动脉（器官浴/DMT-钢丝肌电描记器）。将分析E2和DH 2（UPLC-MS/MS）。蛋白质印迹和RT-PCR将用于蛋白质和mRNA表达。我们提出了三个具体的目标来确定：（1）cyp 2 j-表氧化酶或sEH是否影响血压，腺苷诱导的血管反应和EclO/DHclO;（2）A2 A AR的存在或不存在是否通过PPARs经由cyp 2 j-环氧合酶影响腺苷诱导的血管反应，sEH（3）cyp 2 j-环氧酶或sEH的存在/过表达是否通过腺苷中的A2 A AR-cyp 2 j-PKA-PPARγ/A1 AR-sEH-cyp 4a-PPAR α途径调节KATP通道-诱导血管反应。这样的结果可能会产生积极的影响，因为所鉴定的成分有望提供新的靶点，以遏制与导致高血压的内皮功能障碍相关的临床问题。