Thrombin decreases the urokinase receptor and surface-localized fibrinolysis in cultured endothelial cells

Academic Article

Abstract

  • The endothelial cell (EC) urokinase receptor plays an important role in the localization and receptor-mediated activation of EC-bound plasminogen and hence surface-localized fibrinolysis. Thrombin induced a rapid (<5 minute), time- (0 to 30 minutes) and dose- (0.1 to 8 L/mL) dependent decrease in the specific binding of I-labeled two-chain urokinase-type plasminogen activator (tcu-PA) or diisopropylfluorophosphate-tcu-PA to urokinase-type plasminogen activator receptor (u-PAR) in cultured ECs from various sources (range, 21% to 50%). The thrombin receptor activation peptide but not control peptide showed a similar but reduced decrease in the specific binding of I-labeled tcu-PA to u-PAR. Incubation of thrombin-treated cultures (10 to 12 hours) in complete medium restored I-labeled tcu-PA ligand binding to normal levels. u-PAR mRNA levels rapidly (1 hour) increased and peaked 10 to 12 hours after thrombin treatment as analyzed by reverse transcriptase-polymerase chain reaction. Decreased thrombin-induced I-labeled tcu-PA binding correlated with the time-dependent decrease in surface-localized plasmin generation, as measured by the direct activation of I-labeled Glu-plasminogen and quantification of the 20-kD light chains of I-labeled plasmin. After incubation with thrombin, plasmin generation was decreased 50% to 56% (125 to 152 fmol/3 to 3.5 x 10 cells). Isolation of metabolically labeled S-labeled u-PAR from the media of thrombin and phospholipase C-treated human aortic cultures yielded ~10- and ~12-fold more 55-kD M(r) and ~6-fold more 35-kD M(r) S-labeled u-PAR forms than control cultures, respectively. The u-PAR antigen forms (M(r), 54 kD) and the glycosyl-phosphatidylinositol-anchored protein CD59 (M(r), 20 kD) were also simultaneously identified by immunoprecipitation in the media of thrombin-treated cultures. This suggests that thrombin may release u-PAR and decrease u-PA ligand binding through a common pathway involving phospholipase C. These results establish a novel interrelation between thrombin and EC fibrinolysis and suggest that thrombin may also have an additional regulatory role in the net expression of surface-localized EC fibrinolytic activity. 125 125 125 125 125 125 4 35 35
  • Authors

    Digital Object Identifier (doi)

    Author List

  • Li XN; Varma VK; Parks JM; Benza RL; Koons JC; Grammer JR; Grenett H; Tabengwa EM; Booyse FM
  • Start Page

  • 410
  • End Page

  • 419
  • Volume

  • 15
  • Issue

  • 3