Human foreskin microvascular endothelial cells synthesize and release tissue-type plasminogen activator (t-PA) in similar amounts as do endothelial cells from umbilical cord artery and vein. Human thrombin increases the production of t-PA by these cells, which could be visualized from 8 h after addition of 0.1-5 units/ml thrombin by fibrin autography after SDS polyacrylamide gel electrophoresis of the endothelial cell conditioned media. Thrombin also increased the secretion of t-PA antigen. Together with t-PA, human microvascular cells release urokinase-type plasminogen activator (u-PA) antigen and endothelial cell-type PA inhibitor, PA inhibitor-1, which were both demonstrated by specific immunoprecipitation from radiolabeled endothelial cell conditioned medium. Thrombin increases the release of u-PA antigen, but no u-PA activity could be demonstrated. Thrombin induced a two-fold stimulation of the synthesis and secretion of PA inhibitor-1 antigen. At 0.1 unit/ml thrombin also an increase in PA inhibitor activity was found. At high concentrations of thrombin a decrease of PA inhibitor activity was found, due to the conversion of the active 46 kD PA inhibitor-1 into a 42 kD product without PA inhibitor activity. Our data indicate that interaction of thrombin with microvascular endothelial cells will shift the balance between t-PA, u-PA and PA inhibitor-1, and thus affects the regulation of fibrinolysis.