title: Endogenous platelet fibrinogen surface expression on activated platelets.
abstract: Intracellular platelet fibrinogen surface expression was studied in arabinogalactan-purified, resting, and thrombin-stimulated platelets. Platelet fibrinogen is derived from endocytosis of plasma fibrinogen by megakaryocytes. Like a variety of other adhesive proteins, it is stored in the platelet alpha-granule. Platelet fibrinogen surface expression was studied by using the antigen-binding fragments of a murine monoclonal antibody to platelet fibrinogen, F26, and an immunopurified polyclonal antifibrinogen antibody. Studies correlating platelet fibrinogen surface expression with the presence of the glycoprotein IIb-IIIa (GPIIb-IIIa) complex showed that in the presence of ethylene glycol tetraacetic acid (EGTA) at 37 degrees C, neither the GPIIb-IIIa complex nor platelet fibrinogen was expressed on the surface of thrombin-activated platelets. Similar experiments performed in the presence of EGTA and calcium showed proportional expression of the GPIIb-IIIa complex and platelet fibrinogen. The addition of Arg-Gly-Asp-Ser-containing peptides, the pentadecapeptide of the fibrinogen gamma-chain carboxy terminus, or the monoclonal antibody 10E5, when directed against the GPIIb-IIIa complex before thrombin activation, inhibited 65% to 94% of the platelet fibrinogen expression, as determined with the polyclonal and monoclonal antigen-binding fragments. When these same inhibitory agents were added immediately after or 5 minutes after thrombin, the amount of inhibition decreased significantly. Similar studies with a washed platelet system revealed that when the inhibitors of platelet fibrinogen expression were added before thrombin stimulation, the degree of inhibition observed was only 24% to 38%. This suggests that the major portion of platelet fibrinogen expression involves the release of platelet fibrinogen and its subsequent binding to GPIIb-IIIa. This binding may occur within the open canalicular system or on the platelet surface; in either case, wherever the site of released platelet fibrinogen binding occurs, it can be markedly inhibited by the RGD-containing peptides and the gamma-chain fibrinogen peptides. Approximately 10% to 30% of platelet fibrinogen may be expressed prebound to a platelet receptor, or else it is released and binds to a platelet receptor other than the GPIIb-IIIa complex.
authors.affiliation: Hematology Service, Clinical Center, National Institutes of Health, Bethesda, MD 20892.
authors.forename: H R
authors.initials: HR
authors.lastname: Gralnick
authors.forename: S
authors.initials: S
authors.lastname: Williams
authors.forename: L
authors.initials: L
authors.lastname: McKeown
authors.forename: G
authors.initials: G
authors.lastname: Connaghan
authors.forename: B
authors.initials: B
authors.lastname: Shafer
authors.forename: K
authors.initials: K
authors.lastname: Hansmann
authors.forename: M
authors.initials: M
authors.lastname: Vail
authors.forename: J
authors.initials: J
authors.lastname: Fenton
mesh_terms: D000595:Amino Acid Sequence; D001792:Blood Platelets / Q000378:metabolism*; D002412:Cations / Q000097:blood; D002462:Cell Membrane / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D005340:Fibrinogen / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D015539:Platelet Activation; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D013917:Thrombin / Q000494:pharmacology
chemical_list: D002412:Cations; D007140:Immunoglobulin Fab Fragments; D009842:Oligopeptides; D010980:Platelet Membrane Glycoproteins; D004533:Egtazic Acid; C047981:arginyl-glycyl-aspartic acid; D005340:Fibrinogen; D013917:Thrombin