title: Plasminogen associates with phosphatidylserine-exposing platelets and contributes to thrombus lysis under flow.
abstract: The interaction of plasminogen with platelets and their localization during thrombus formation and fibrinolysis under flow are not defined. Using a novel model of whole blood thrombi, formed under flow, we examine dose-dependent fibrinolysis using fluorescence microscopy. Fibrinolysis was dependent upon flow and the balance between fibrin formation and plasminogen activation, with tissue plasminogen activator-mediated lysis being more efficient than urokinase plasminogen activator-mediated lysis. Fluorescently labeled plasminogen radiates from platelet aggregates at the base of thrombi, primarily in association with fibrin. Hirudin attenuates, but does not abolish plasminogen binding, denoting the importance of fibrin. Flow cytometry revealed that stimulation of platelets with thrombin/convulxin significantly increased the plasminogen signal associated with phosphatidylserine (PS)-exposing platelets. Binding was attenuated by tirofiban and Gly-Pro-Arg-Pro amide, confirming a role for fibrin in amplifying plasminogen binding to PS-exposing platelets. Confocal microscopy revealed direct binding of plasminogen and fibrinogen to different platelet subpopulations. Binding of plasminogen and fibrinogen co-localized with PAC-1 in the center of spread platelets. In contrast, PS-exposing platelets were PAC-1 negative, and bound plasminogen and fibrinogen in a protruding "cap." These data show that different subpopulations of platelets harbor plasminogen by diverse mechanisms and provide an essential scaffold for the accumulation of fibrinolytic proteins that mediate fibrinolysis under flow.
authors.affiliation: Institute of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom; and.
authors.forename: Claire S
authors.identifier: http://orcid.org/0000-0001-8127-6102
authors.initials: CS
authors.lastname: Whyte
authors.affiliation: Department of Biochemistry and Anesthesiology, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands.
authors.forename: Frauke
authors.initials: F
authors.lastname: Swieringa
authors.affiliation: Department of Biochemistry and Anesthesiology, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands.
authors.forename: Tom G
authors.initials: TG
authors.lastname: Mastenbroek
authors.affiliation: Institute of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom; and.
authors.forename: Ausra S
authors.initials: AS
authors.lastname: Lionikiene
authors.affiliation: Department of Biochemistry and Anesthesiology, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands.
authors.forename: Marcus D
authors.initials: MD
authors.lastname: Lancé
authors.affiliation: Department of Biochemistry and Anesthesiology, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands.
authors.forename: Paola E J
authors.initials: PE
authors.lastname: van der Meijden
authors.affiliation: Department of Biochemistry and Anesthesiology, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands.
authors.forename: Johan W M
authors.initials: JW
authors.lastname: Heemskerk
authors.affiliation: Institute of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom; and.
authors.forename: Nicola J
authors.identifier: http://orcid.org/0000-0002-7452-0813
authors.initials: NJ
authors.lastname: Mutch
mesh_terms: D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D003435:Crotalid Venoms / Q000494:pharmacology; D005337:Fibrin / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism*; D005342:Fibrinolysis / Q000187:drug effects; D005343:Fibrinolytic Agents / Q000494:pharmacology; D005434:Flow Cytometry; D006490:Hemostatics / Q000494:pharmacology; D006629:Hirudins / Q000494:pharmacology; D037181:Lectins, C-Type; D018613:Microscopy, Confocal; D009842:Oligopeptides / Q000494:pharmacology; D010718:Phosphatidylserines / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D012212:Rheology; D033081:Shear Strength; D013917:Thrombin / Q000494:pharmacology; D013927:Thrombosis / Q000378:metabolism*; D000077466:Tirofiban; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014443:Tyrosine / Q000031:analogs & derivatives / Q000494:pharmacology; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
chemical_list: D003435:Crotalid Venoms; D005343:Fibrinolytic Agents; D006490:Hemostatics; D006629:Hirudins; D037181:Lectins, C-Type; D009842:Oligopeptides; D010718:Phosphatidylserines; C028890:convulxin; D014443:Tyrosine; C034585:glycyl-prolyl-arginyl-proline; D005337:Fibrin; D013917:Thrombin; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator; D000077466:Tirofiban