Elongation of arachidonic and eicosapentaenoic acids limits their availability for thrombin-stimulated release from the glycerolipids of vascular endothelial cells

Abstract

This study has examined the thrombin-stimulated release of polyunsaturated fatty acids from endothelial glycerolipids. Human umbilical vein endothelial cells were incubated with 1.25 μM [ 14C]arachidonate or [ 14C]eicosapentaenoate and then exposed to thrombin in buffered saline plus albumin. After an incorporation period of 0.5 h, the thrombin-stimulated release of the two radiolabeled fatty acids was quite similar. By contrast, after 24 h of fatty acid incorporation, the thrombin-stimulated release of radiolabeled fatty acid from cells incubated with [ 14C]eicosapentaenoate was only 25–30% of that from cells with [ 14C]arachidonate. Analysis of cellular glycerolipids indicated that 23 and 72%, respectively, of the incorporated [ 14C]arachidonate and [ 14C]eicosapentaenoate had been elongated to 22-carbon fatty acids in 24 h. Both 20- and 22-carbon 14C-labeled fatty acids were released to albumin in the medium in control incubations. Addition of thrombin stimulated the release of [ 14C]arachidonate and [ 14C]eicosapentaenoate, but not of their respective elongation products. Furthermore, endothelial cells incorporated exogenous [ 14C]docosatetraenoate into cellular glycerolipids but did not release it in response to thrombin. Thus, the thrombin-stimulated release of polyunsaturated fatty acids from vascular endothelial cells is highly selective for arachidonate and eicosapentaenoate. These results suggest that the extensive elongation of eicosapentaenoate by these cells serves to remove n-3 polyunsaturated fatty acids from the pool of cellular acyl groups which are released in response to thrombin and are thus made available for metabolism by cyclooxygenase and lipoxygenase enzymes.