The inactivation of single-chain urokinase-type plasminogen activator by thrombin on cultured human endothelial cells.
Single-chain urokinase-type plasminogen activator (scu-PA) is cleaved by thrombin, resulting in an inactive molecule called thrombin-cleaved two-chain urokinase-type plasminogen activator (tcu-PA/T). There is no knowledge about cell-mediated inactivation of scu-PA. We have studied whether scu-PA bound to cultured human umbilical vein endothelial cells (HUVEC) could be inactivated by thrombin. High molecular weight scu-PA was bound to HUVEC and incubated with increasing amounts of thrombin for 30 min at 37 degrees C. Cell-bound urokinase-type plasminogen activator (u-PA) was released and levels of scu-PA, tcu-PA/T and active two-chain u-PA were measured using sensitive bioimmunoassays. Cell-bound scu-PA was efficiently inactivated by thrombin. Fifty percent inactivation of scu-PA occurred at about 0.2 nM thrombin. In the presence of monoclonal anti-urokinase receptor IgG, at least 50% of the binding of scu-PA to HUVEC was inhibited. The relative amount of tcu-PA/T that was generated by thrombin was not affected by the monoclonal antibody. These results indicated that scu-PA bound to HUVEC via the urokinase receptor can be inactivated by thrombin. The efficient inactivation of cell-bound scu-PA suggests that a cofactor for thrombin may be involved, like thrombomodulin or glycosaminoglycans. It is concluded that scu-PA bound to the urokinase receptor on a cell surface can be inactivated by thrombin, which may have profound effects on u-PA-mediated local fibrinolysis and extracellular proteolysis during processes in which thrombin is also involved.
Braat
E A
EA
TNO Prevention and Health, Gaubius Laboratory, P.O. Box 2215, 2301 CE, Leiden, The Netherlands.
Collen
A
A
Jie
A F
AF
Grimbergen
J M
JM
Rijken
D C
DC
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D015536:Down-Regulation; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D005342:Fibrinolysis; D006801:Humans; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D013917:Thrombin / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000378:metabolism*
D000911:Antibodies, Monoclonal; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D013917:Thrombin; D014568:Urokinase-Type Plasminogen Activator