The Biological Actions of 11,12‐Epoxyeicosatrienoic Acid in Endothelial Cells are Specific to the R/S Enantiomer and Require the Gs Protein

Cytochrome P450 (CYP)‐derived epoxides of arachidonic acid, i.e., the epoxyeicosatrienoic acids (EETs), are important lipid signaling molecules. Since many actions of 11,12‐EET are dependent on the activation of protein kinase A (PKA), the existence of a cell surface Gs‐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‐EET‐induced, PKA‐dependent translocation of transient receptor potential (TRP) C6 channels as well as angiogenesis. In primary cultures of human endothelial cells, (±)‐11,12‐EET led to the rapid (30 seconds) translocation a TRPC6V5 fusion protein, an effect reproduced by 11(R),12(S)‐EET, but not 11(S),12(R)‐EET or (±)‐14,15‐EET. Similarly, endothelial cell migration and tube formation were stimulated by (±)‐11,12‐EET and 11(R),12(S)‐EET while 11(S),12(R)‐EET and 11,12‐dihydroxyeicosatrienoic acid were without effect. The effects of (±)‐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 siRNA‐mediated downregulation of Gs in endothelial cells had no significant effect on responses stimulated by VEGF or a PKA activator but abolished responses to (±)‐11,12‐EET. The down regulation of Gq/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 Gs‐coupled receptor in the endothelial cell membrane responds to 11(R),12(S)‐EET, mediates the PKA‐dependent translocation and activation of TRPC6 channels, as well as angiogenesis.