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

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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.