Thrombin induction of plasminogen activator-inhibitor in cultured human endothelial cells.

Abstract

We have examined the effect of thrombin on the activity of plasminogen activator (PA) and plasminogen activator-inhibitor (PA-I) in medium conditioned by primary cultures of human umbilical vein endothelial cells. PA activity was measured by fibrinolytic and esterolytic assays, and total tissue-type PA (tPA) antigen by radioimmunoassay. Net PA-I activity was assayed by titration of human urokinase esterolytic activity. Incubation of confluent endothelial cell cultures with thrombin for 24 h caused a sixfold increase in PA-I activity. The effect of thrombin was half-maximal at -0.4 U/ml (<4 nM), and required concomitant RNA and protein synthesis. The stimulation of PA-I activity required active a-thrombin and was not obtained with 'y-thrombin nor with thrombin catalytically inactivated with hirudin. Because of the excess of PA-I, PA activity was not measurable in either control or thrombin-treated cells. Thrombin did, however, increase medium concentration of tPA antigen by approximately fourfold. The thrombin-induced PA-I inhibited both tPA and urokinase, did not lose activity upon acidification, and was stable to sodium dodecyl sulfate and thiol reduction. We conclude that physiologic concentrations of thrombin increase both PA-I activity and tPA antigen in medium conditioned by human umbilical vein endothelial cells. Because there was always a several-fold increase in the net activity of free PA-I, these observations suggest that the net effect of thrombin is to decrease fibrinolytic activity in human endothelial cells. Thus, thrombin, in addition to its role in coagulation, may protect clots from premature lysis by increasing the amount of a specific fibrinolytic inhibitor.

Urokinase and Tissue-type Plasminogen Activator Are Required for the Mitogenic and Chemotactic Effects of Bovine Fibroblast Growth Factor and Platelet-derived Growth Factor-BB for Vascular Smooth Muscle Cells

“…This observation suggests that SMC may possess a t-PA receptor analogous to the SMC thrombin or factor Xa receptors which exhibits a proteolytic mechanism of receptor activation (34 -36, 57). The fact that PAI-1 has been shown to be expressed by SMC following PDGF, transforming growth factor ␤, bFGF, or thrombin stimulation (58,59), together with the identification of a functionally active PAI-1 accessible to t-PA, identified in the extracellular matrix of cultured endothelial cells and SMC (9,10,48,60) gives an added dimension to our observation showing that PAI-1 inhibited the mitogenic effect of t-PA. Indeed, it suggests that SMCs might thereby regulate their own plasminogen activators in an autocrine fashion.…”

The cytokine‐mediated imbalance between coagulant and anticoagulant mechanisms in sepsis and endotoxaemia

“…Also, the administration of the specific thrombin inhibitor hirudin could not prevent the endotoxin-induced activation and inhibition of fibrinolysis [32]. These observations suggest that endotoxininduced effects on coagulation and fibrinolysis are regulated independently and that earlier hypotheses of thrombin-mediated activation or inhibition of fibrinolysis (partly based on in vitro studies) may not be correct [33][34][35].…”

Characterization of a functional thrombin receptor. Issues and opportunities.

“…Thrombin is chemotactic for monocytes (4), mitogenic for lymphocytes and for mesenchymal cells including vascular smooth muscle cells (5,6), and has a number ofeffects upon the vascular endo-thelium. These include stimulating endothelial production of prostacyclin (7), platelet-activating factor (8), plasminogen activator-inhibitor (9), and the potent smooth muscle cell mitogen platelet-derived growth factor (10). Thrombin also induces neutrophil adherence to the vessel wall by an endothelial-dependent mechanism (1 1), probably by causing surface expression of GMP-140 (12).…”