Evidence for a two-step process in prostaglandin secretion. Intracellular accumulation of prostacyclin precedes its release from human endothelial cells in culture

Cultured endothelial cells (EC) from human umbilical vein were incubated with U-14C]arachidonic acid (AA) followed by a challenge with thrombin (2 units/ml) or calcium ionophore A23187 (5 microM) for 0.5-10 min at 37 degrees C. In both cases, AA was rapidly liberated from phospholipids and converted into prostaglandin I2 (PGI2), as determined by the radioactivity of the stable derivative 6-keto-PGF1 alpha. Maximal liberation of AA and synthesis of PGI2 were achieved within 2 min, but the two compounds first accumulated in EC prior to their release into supernatants. This finding, which was never reported before, raises the question of the mechanism of AA and PG release through the cell membranes and offers a convenient model to investigate this still obscure process.