Formation of biologically active autacoids is regulated by calcium influx in endothelial cells.
The blocker of receptor-mediated calcium entry SK&F 96365 was used to evaluate the contribution of calcium influx to the formation of biologically active endothelial prostanoids and endothelium-derived relaxing factor (EDRF). SK&F 96365 inhibited histamine-stimulated calcium entry into human umbilical vein endothelial cells but not its discharge from intracellular stores as determined spectrofluorometrically by changes of intracellular calcium concentration in fura-2-loaded cells. Concordantly, SK&F 96365 inhibited histamine-induced endothelial synthesis of 6-keto-prostaglandin F1 alpha and thromboxane B2 in a dose-dependent manner. To assess the functional significance of endothelial formation of prostacyclin and EDRF to platelets, the cAMP- and cGMP-dependent phosphorylation of two platelet proteins, rap1B and a 50-kD vasodilator-stimulated phosphoprotein (VASP), was analyzed in coincubation experiments of endothelial cells with platelets. Autacoids released by histamine-stimulated endothelial cells caused the phosphorylation of rap1B and VASP in platelets, which was only partly inhibited by either indomethacin or NG-monomethyl-L-arginine but was almost completely suppressed by SK&F 96365. The concomitant endothelial release of thromboxane A2 had no effect on protein kinase C- and calcium-dependent phosphorylation of platelet proteins. The results demonstrate that blockade of receptor-mediated calcium entry by SK&F 96365 markedly reduced the release of biologically active prostacyclin and EDRF from endothelial cells. Thus, calcium influx but not calcium release from intracellular stores plays a critical role in the receptor-stimulated formation and liberation of prostacyclin and EDRF in endothelial cells.
Kruse
H J
HJ
Institut für Prophylaxe und Epidemiologie der Kreislaufkrankheifen, University of Munich, FRG.
Grünberg
B
B
Siess
W
W
Weber
P C
PC
D000818:Animals; D012898:Autacoids / Q000096:biosynthesis*; D001792:Blood Platelets / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015815:Cell Adhesion Molecules; D002478:Cells, Cultured; D003584:Cytological Techniques; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism; D006632:Histamine / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism; D008840:Microfilament Proteins; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011453:Prostaglandins / Q000378:metabolism; D013552:Swine; D020668:rap GTP-Binding Proteins; D000098662:Vasodilator-Stimulated Phosphoprotein
D012898:Autacoids; D002121:Calcium Channel Blockers; D015815:Cell Adhesion Molecules; D008565:Membrane Proteins; D008840:Microfilament Proteins; D010750:Phosphoproteins; D011453:Prostaglandins; D000098662:Vasodilator-Stimulated Phosphoprotein; D006632:Histamine; D019204:GTP-Binding Proteins; D020668:rap GTP-Binding Proteins; D002118:Calcium