The Biological Actions of 11,12-Epoxyeicosatrienoic Acid in Endothelial Cells Are Specific to theR/S-Enantiomer and Require the G_sProtein

Abstract

Cytochrome P450-derived epoxides of arachidonic acid [i.e., the epoxyeicosatrienoic acids (EETs)] are important lipid signaling molecules involved in the regulation of vascular tone and angiogenesis. Because many actions of 11,12-cis-epoxyeicosatrienoic acid (EET) are dependent on the activation of protein kinase A (PKA), the existence of a cell-surface G s -coupled receptor has been postulated. To assess whether the responses of endothelial cells to 11,12-EET are enantiomer specific and linked to a potential G protein-coupled receptor, we assessed 11,12-EETinduced, PKA-dependent translocation of transient receptor potential (TRP) C6 channels, as well as angiogenesis. In primary cultures of human endothelial cells, (6)-11,12-EET led to the rapid (30 seconds) translocation a TRPC6-V5 fusion protein, an effect reproduced by 11(R),12(S)-EET, but not by 11(S),12(R)-EET or (6)-14,15-EET. Similarly, endothelial cell migration and tube formation were stimulated by (6)-11,12-EET and 11(R),12(S)-EET, whereas 11(S),12(R)-EET and 11,12-dihydroxyeicosatrienoic acid were without effect. The effects of (6)-11,12-EET on TRP channel translocation and angiogenesis were sensitive to EET antagonists, and TRP channel trafficking was also prevented by a PKA inhibitor. The small interfering RNA-mediated downregulation of G s in endothelial cells had no significant effect on responses stimulated by vascular endothelial growth or a PKA activator but abolished responses to (6)-11,12-EET. The downregulation of G q /11 failed to prevent 11,12-EET-induced TRPC6 channel translocation or the formation of capillary-like structures. Taken together, our results suggest that a G s -coupled receptor in the endothelial cell membrane responds to 11(R),12(S)-EET and mediates the PKA-dependent translocation and activation of TRPC6 channels, as well as angiogenesis.

GPR40 is a low-affinity epoxyeicosatrienoic acid receptor in vascular cells

“…EETs have diverse biological actions including vasodilation, inhibition of inflammation, promoting angiogenesis and organ protection (5,6,10,18). These actions and the cellular components to these actions occur over a wide concentration range from 10 -9 -10 -5 M (4).…”

“…Additionally, 11,12-and 14,15-EETs inhibit smooth muscle cell migration and aromatase expression (8,9). On endothelial cells, 11,12-EET promotes cell growth, decrease apoptosis, migration, tube formation and angiogenesis, increases tissue plasminogen activator (tPA) release, increases cyclooxygenase-2 (COX-2) expression and alters connexin-43 (Cx43) expression and gap junctions (10)(11)(12)(13)(14)(15)(16)(17). EET isomers also inhibit platelet adhesion to the endothelium, and 11,12-and 8,9-EETs inhibit inflammation by decreasing leukocyte adhesion to the endothelium (18,19).…”

The Pharmacology of the Cytochrome P450 Epoxygenase/Soluble Epoxide Hydrolase Axis in the Vasculature and Cardiovascular Disease

“…This concept is supported by the fact that there are also differences in responsiveness to different EET stereoisomers and not just the regioisomers. For example, 11(R),12(S)-EET is a more potent activator of renal artery K Ca channels (Zou et al, 1996) and rat airway electrical parameters 1114 Fleming (Pascual et al, 1998), as well as endothelial cell TRP channel translocation and angiogenesis (Ding et al, 2014), than 11(S),12(R)-EET. However, this is not a universal observation because 11(S),12(R)-EET is reportedly more effective than 11(R),12(S)-EET in activating cardiac K ATP channels (Lu et al, 2002).…”

11,12-Epoxyeicosatrienoic acid induces vasodilator response in the rat perfused mesenteric vasculature

“…found that RuR completely overturned smooth muscle hyperpolarization induced by 11, 12‐EET in arteries from WT mice. Ding et al 41. reported that in the endothelial cell membrane, a G(s)‐coupled receptor responds to 11(R),12(S)‐EET and mediates the PKA‐dependent translocation and stimulation of TRPC6 channels.…”