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Characterization of GPR40 as a Low Affinity Epoxyeicosatrienoic Acid (EET) Receptor in Vascular Cells
Author(s) -
Park SangKyu,
Herrnreiter Anja,
Pfister Sandra L.,
Falck John R.,
Campbell William B.
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.827.6
Subject(s) - free fatty acid receptor 1 , vascular smooth muscle , chemistry , calcium in biology , microbiology and biotechnology , mapk/erk pathway , agonist , umbilical vein , receptor , medicine , biochemistry , phosphorylation , endocrinology , biology , in vitro , smooth muscle
Endothelium‐derived EETs have a number of vascular actions including dilating arteries, promoting angiogenesis and reducing inflammation. These actions may be mediated by high and low affinity G protein‐coupled receptors. GPR40 is activated by long chain free fatty acids and EETs. We investigated the role of GPR40 and its signaling mechanisms in some vascular actions of EETs. 14,15‐EET, 11,12‐EET and the GPR40 agonist GW9508 increased intracellular calcium concentration in HEK293 cells overexpressing human GPR40 (EC 50 = 0.66 ± 0.13 μM, 0.98 ± 0.09 μM and 19 ± 0.37 nM, respectively) but were without effect in non‐transfected HEK293 cells. 8,9‐EET, 11,12‐DHET and 14,15‐DHET were less active. GW1100 (10 μM), the GPR40 antagonist, blocked these increases in calcium by EETs and GW9508. Using PCR and immunoblotting, GPR40 expression was detected in human and bovine endothelial cells (ECs), smooth muscle cells and arteries. 11,12‐EET caused concentration‐related relaxations of U46619‐preconstricted bovine coronary arteries; however, these relaxations were not altered by GW1100 (EC 50 = 0.46 ± 0.14 μM vs 0.63 ± 0.37 μM; control vs GW1100). 11,12‐EET increased intracellular calcium in human umbilical vein ECs. In these ECs, 11,12‐EET (1 μM) increased MAP kinase phosphorylation of ERK and increased the expression of connexin 43 (Cx43) and cyclooxygenase‐2 (COX‐2). GW1100 and the MAP kinase inhibitor U0126 (1 μM) both inhibited 11,12‐EET‐mediated ERK phosphorylation and Cx43 and COX‐2 increases. In addition, the NFκB inhibitor andrographolide (40 μM) also inhibited the 11,12‐EET‐mediated COX‐2 increase in ECs. Our results indicate that GPR40 is a low affinity EET receptor that is expressed in vascular cells and arteries. GPR40 does not mediate EET‐induced vasorelaxation;, however, 11,12‐EET through GPR40 increases Cx43 and COX‐2 expression in ECs via ERK phosphorylation. Thus, Cx43 and COX‐2 play important roles in cell to cell gap junctional communication and prostaglandin production to mediate some of vascular actions of EETs. Support or Funding Information This work was funded by grants from the National Institutes of Health (HL‐83297), Regulation of adrenal tone by steroidogenesis cells.