Effects of epoxyeicosatrienoic acids on levels of eNOS phosphorylation and relevant signaling transduction pathways involved.
Endothelial nitric oxide synthase (eNOS) is a key enzyme responsible for the regulation of vascular homeostasis. Many humor factors and mechanical forces can affect eNOS activity via phosphorylation modification but the mechanisms involved vary with stimuli applied. We have demonstrated that cytochrome P450 (CYP) epoxygenase-dependent metabolites of arachidonic acid, epoxyeicosatrienoic acids (EETs), can robustly up-regulate eNOS expression and its activity, however the relevant signaling pathways responsible for activity regulation are not well known. In this study, we explored the role of PI3 kinase (PI3K)/protein kinase B (Akt) signaling pathway in eNOS expression and its phosphorylation in response to EETs via direct addition of EETs into cultured bovine aorta endothelial cells (BAECs) and recombinant adeno-associated virus-mediated transfection of CYP epoxygenase genes CYPF87V and CYP2C11 to produce endogenous EETs followed by co-treatment with PI3K or Akt inhibitor. Results show that both exogenous and endogenous EETs could remarkably enhance eNOS expression and its phosphorylation at Ser1179 and Thr497 residues; PI3K inhibitor LY294002 could inhibit EETs-induced increase in eNOS-Ser(P)1179 but had no effect on the change of eNOS-Thr(P)497, while Akt inhibitor could attenuate the increase in phosphor-eNOS at both residues; both of the two inhibitors could block EETs-enhanced eNOS expression. These results lead to conclusions: (i) EETs-mediated regulation of eNOS activity may be related with the changes of phosphorylation level at eNOS-Ser1179 via P13K/Akt and eNOS-Thr497 via Akt; (ii) PI3K/Akt signaling pathway is involved in the up-regulation of eNOS expression by EETs.
Chen
Ruijuan
R
The Institute of Hypertension and Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science & Technology, Wuhan 430030, China.
Jiang
Jiangang
J
Xiao
Xiao
X
Wang
Daowen
D
D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000502:physiology; D002417:Cattle; D002478:Cells, Cultured; D000072467:Cytochrome P450 Family 2; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D005231:Fatty Acids, Unsaturated / Q000502:physiology; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism* / Q000502:physiology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D039181:Steroid 16-alpha-Hydroxylase / Q000502:physiology; D015854:Up-Regulation / Q000187:drug effects
D015126:8,11,14-Eicosatrienoic Acid; D001189:Aryl Hydrocarbon Hydroxylases; D000072467:Cytochrome P450 Family 2; D005231:Fatty Acids, Unsaturated; D052250:Nitric Oxide Synthase Type III; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D039181:Steroid 16-alpha-Hydroxylase; C528813:15-hydroxy-11,12-epoxyeicosatrienoic acid; C047008:17-octadecynoic acid; C050715:8,9-epoxyeicosatrienoic acid; C046782:14,15-epoxy-5,8,11-eicosatrienoic acid; C450279:CYP2C11 protein, rat