Endothelial cells play a central role in fibrinolysis due to their production of both tissue plasminogen activator (t-PA) and plasminogen activator inhibitor type-1 (PAI-1). The purpose of this study was to test the hypothesis that the production of t-PA and PAI-1 from human umbilical vein endothelial cells (HUVEC) and from human adult vein endothelial cells (HAVEC) adopt the same pathways for the regulation of fibrinolysis, and that differences in PAI-1 and t-PA production are only quantitative. HUVEC and HAVEC cultures were exposed to phorbol ester (PMA), endotoxin (LPS) or thrombin, singly or in combination with forskolin or H7. The conditioned medium was collected after 16 h and analyzed for t-PA and PAI-1 antigens. The basal production of both t-PA and PAI-1 was significantly higher from HUVEC than from HAVEC. Exposure to PMA, thrombin or forskolin gave a similar response from the two cell types. In contrast, the results from HUVEC and HAVEC differed significantly, not only in magnitude but also in direction, when the cells were exposed to forskolin coincubated with PMA, LPS or thrombin, and in magnitude alone when exposed to LPS only. The results indicate that there are not only quantitative but also qualitative differences in the production of t-PA and PAI-1 from HUVEC and HAVEC. These differences must be taken into account when data from cells of different origin are compared.