title: The role of thrombin receptors PAR1 and PAR4 for PAI-1 storage, synthesis and secretion by human platelets.
abstract: BACKGROUND
  Arterial thrombi contain more platelets than venous thrombi and are more resistant to fibrinolysis. This resistance could partly be due to plasminogen activator inhibitor 1 (PAI-1) secreted by platelets. The aim of this study was to elucidate differences between thrombin receptors protease-activated receptor (PAR) 1 and 4 and platelet storage, secretion and synthesis of platelet PAI-1, as compared to other platelet α-granule proteins such as VEGF and endostatin.
  
  
  METHODS
  Human isolated platelets were incubated with thrombin (0.5 U/ml), PAR1-activating peptide (AP) (0.4-30 μM) or PAR4-AP (1.5-300 μM) for up to 24 hours. ELISA, western blot and fluorescence microscopy were used to measure secretion, contents and localization of PAI-1, VEGF and endostatin.
  
  
  RESULTS
  Our results show that PAI-1 and VEGF might be co-localized and that endostatin does not co-localize with either PAI-1 or VEGF. PAI-1 and VEGF show a similar secretion pattern, being more sensitive to low grade PAR1 activation, but secretion was also observed with higher concentrations of PAR4-APs. PAI-1 is secreted in an active form. PAI-1 mRNA was found in platelets, and elevated levels of PAI-1 were detected after 24 hours incubation of platelets.
  
  
  CONCLUSIONS
  PAI-1 and VEGF, but not endostatin, might be stored in the same α-granule in human platelets. PAI-1 and VEGF also show a similar secretion pattern, being more sensitive to PAR1 than to PAR4 activation, but the secretion is not exclusively selective. Our results also show that platelet PAI-1 is increased if incubated for 24 hours, both with addition of PAR1-activating peptide and without activation, which could indicate de novo synthesis.
authors.affiliation: Department of Clinical and Experimental Medicine, Linköping University, Sweden. martina.nylander@liu.se
authors.forename: Martina
authors.initials: M
authors.lastname: Nylander
authors.forename: Abdimajid
authors.initials: A
authors.lastname: Osman
authors.forename: Sofia
authors.initials: S
authors.lastname: Ramström
authors.forename: Emma
authors.initials: E
authors.lastname: Aklint
authors.forename: Anders
authors.initials: A
authors.lastname: Larsson
authors.forename: Tomas L
authors.initials: TL
authors.lastname: Lindahl
mesh_terms: D001792:Blood Platelets / Q000378:metabolism*; D002478:Cells, Cultured; D043169:Endostatins / Q000378:metabolism*; D006801:Humans; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism*; D044463:Receptor, PAR-1 / Q000378:metabolism*; D018179:Receptors, Thrombin / Q000378:metabolism*; D013917:Thrombin / Q000493:pharmacokinetics*
chemical_list: D043169:Endostatins; D017395:Plasminogen Activator Inhibitor 1; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; C051321:SERPINE1 protein, human; D013917:Thrombin; C112908:protease-activated receptor 4