This study was designed to evaluate the effect of thrombin on prostacyclin (PGI 2) production in cultured human vascular endothelial cells in association with intracellular Ca 2+ and with the gene expression of prostaglandin H 2 synthase (PGHS) and phospholipase A 2 (PLA 2) using competitive polymerase chain reaction. Thrombin enhanced the PGI 2 synthesis dependent with time. Additionally, thrombin increased the intracellular Ca 2+, which stimulates PLA 2, resulting in arachidonic acid cleavage from membrane phospholipids and its subsequent conversion into PGI 2 through the PGHS pathway. The elevation of intracellular Ca 2+ was a result of Ca 2+ influx and Ca 2+ release from its intracellular storage sites. In this study, PGHS-1 mRNA was constitutively expressed, whereas PGHS-2 mRNA was not. With the stimulation of thrombin, cytosolic PLA 2 (cPLA 2) mRNA increased 9-fold at 15 min, PGHS-1 mRNA increased 3.4-fold at 180 min, and PGHS-2 mRNA increased 38-fold at 60 min. These results suggest that the elevation of intracellular Ca 2+ and the expression of cPLA 2, PGHS-1, and PGHS-2 mRNA cause PGI 2 generation.