Functional implications of cAMP and Ca2+ on prostaglandin I2 and thromboxane A2 synthesis by human endothelial cells.

Abstract

Human umbilical vein endothelial cells were incubated with ATP, ADP, thrombin, or ionophore A23187 for as long as 600 seconds. A statistically significant rise in the prostaglandin I2 (prostacyclin; PGI2) and thromboxane A2 (TxA2) release was observed after 45 seconds. The maximum amount of cytosolic free Ca2+ was reached between 20 and 30 seconds. A statistically significant elevation of intracellular adenosine 3',5'-cyclic monophosphate (cAMP) levels was observed only in response to ATP and ADP after 120 seconds. Furthermore, adenosine, caffeine, phorbol 12-myristate 13-acetate (PMA), and adenosine 5'-(alpha, beta-methylene)triphosphate were tested alone or in combination with ATP or thrombin. PMA inhibited ATP-stimulated eicosanoid biosynthesis but had no effect on thrombin. Of the agents used to increase cytosolic free Ca2+ concentrations, only PMA failed to provoke eicosanoid release. Similarly, only PMA failed to induce a Ca2+ flux in the absence of extracellular Ca2+. The data presented show that PGI2 and TxA2 release from human umbilical cord endothelial cells is closely associated with Ca2+ mobilized from intracellular sources. Extracellular Ca2+, as well as changes in intracellular cAMP levels, has no influence on endothelial eicosanoid synthesis, at least for short-term regulation (less than or equal to 600 seconds).