title: Activation of protein C and thrombin activable fibrinolysis inhibitor on cultured human endothelial cells.
abstract: ESSENTIALS: It is unknown if thrombin activatable fibrinolysis inhibitor (TAFI) and protein C compete on cells. TAFI and protein C activation on endothelial cells was simultaneously quantified. TAFI and protein C do not compete for activation on endothelial cells. TAFI and protein C are independently recognized by the thrombin-thrombomodulin complex.
  
  
  BACKGROUND
  When bound to thrombomodulin (TM), thrombin is a potent activator of protein C (PC) and thrombin activable fibrinolysis inhibitor (TAFI). By binding PC and presenting it to the thrombin-TM complex, endothelial cell PC receptor (EPCR) enhances PC activation. It is unknown whether PC and TAFI compete for the thrombin-TM complex on endothelial cells.
  
  
  OBJECTIVE
  To compare PC and TAFI activation on the surface of cultured human endothelial cells in the absence or presence of JRK1535 and/or CTM1009, inhibitory antibodies directed against EPCR and TM, respectively, and to determine whether PC and TAFI compete with each other for activation.
  
  
  METHODS
  PC and TAFI activation on endothelial cells were compared, and the effect of PC on TAFI activation and TAFI on PC activation was determined in the absence or presence of JRK1535 and/or CTM1009.
  
  
  RESULTS
  In the absence of antibodies, activation of PC was four-fold faster than that of TAFI. Blocking EPCR with JRK1535 resulted in a 53-fold decrease in PC activation and no effect on TAFI activation. Blocking TM with CTM1009 inhibited both TAFI and PC activation. Neither TAFI nor PC competed with each other in the absence or presence of JRK1535.
  
  
  CONCLUSIONS
  PC and TAFI are concurrently activated in a TM-dependent manner and do not compete for the thrombin-TM complex, raising the possibility that they interact with distinct activation complexes. EPCR selectively enhances PC activation so that PC and TAFI activation kinetics become comparable on endothelial cells.
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, Hamilton, ON, Canada.|Department of Medicine, McMaster University, Hamilton, ON, Canada.
authors.forename: C
authors.initials: C
authors.lastname: Wu
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, Hamilton, ON, Canada.|Department of Medicine, McMaster University, Hamilton, ON, Canada.
authors.forename: P Y
authors.identifier: http://orcid.org/0000-0002-0504-3064
authors.initials: PY
authors.lastname: Kim
authors.affiliation: Department of Pathology and Molecular Medicine, Queen's University, Kingston, ON, Canada.
authors.forename: L L
authors.initials: LL
authors.lastname: Swystun
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, Hamilton, ON, Canada.|Department of Medicine, McMaster University, Hamilton, ON, Canada.
authors.forename: P C
authors.initials: PC
authors.lastname: Liaw
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, Hamilton, ON, Canada.|Department of Medicine, McMaster University, Hamilton, ON, Canada.|Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, Canada.
authors.forename: J I
authors.initials: JI
authors.lastname: Weitz
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D001667:Binding, Competitive; D025901:Carboxypeptidase B2 / Q000378:metabolism*; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000201:enzymology*; D000074283:Endothelial Protein C Receptor; D004789:Enzyme Activation; D061307:Human Umbilical Vein Endothelial Cells / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D011485:Protein Binding; D011486:Protein C / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism; D018180:Thrombomodulin / Q000378:metabolism
chemical_list: D015703:Antigens, CD; D000074283:Endothelial Protein C Receptor; C478370:PROCR protein, human; D011486:Protein C; D011956:Receptors, Cell Surface; C546931:THBD protein, human; D018180:Thrombomodulin; D025901:Carboxypeptidase B2; D013917:Thrombin
keywords: carboxypeptidase B2; endothelial cells; enzyme activation; protein C; thrombin-activable fibrinolysis inhibitor; thrombomodulin