title: Studies of activated GPIIb/IIIa receptors on the luminal surface of adherent platelets. Paradoxical loss of luminal receptors when platelets adhere to high density fibrinogen.
abstract: The accessibility of activated GPIIb/IIIa receptors on the luminal surface of platelets adherent to damaged blood vessels or atherosclerotic plaques is likely to play a crucial role in subsequent platelet recruitment. To define better the factors involved in this process, we developed a functional assay to assess the presence of activated, luminal GPIIb/IIIa receptors, based on their ability to bind erythrocytes containing a high density of covalently coupled RGD-containing peptides (thromboerythrocytes). Platelets readily adhered to wells coated with purified type I rat skin collagen and the adherent platelets bound a dense lawn of thromboerythrocytes. With fibrinogen-coated wells, platelet adhesion increased as the fibrinogen-coating concentration increased, reaching a plateau at about 11 micrograms/ml. Thromboerythrocyte binding to the platelets adherent to fibrinogen showed a paradoxical response, increasing at fibrinogen coating concentrations up to approximately 4-6 micrograms/ml and then dramatically decreasing at higher fibrinogen-coating concentrations. Scanning electron microscopy demonstrated that the morphology of platelets adherent to collagen was similar to that of platelets adherent to low density fibrinogen, with extensive filopodia formation and ruffling. In contrast, platelets adherent to high density fibrinogen showed a bland, flattened appearance. Immunogold staining of GPIIb/IIIa receptors demonstrated concentration of the receptors on the filopodia, and depletion of receptors on the flattened portion of the platelets. Thus, there is a paradoxical loss of accessible, activated GPIIb/IIIa receptors on the luminal surface of platelets adherent to high density fibrinogen. Two factors may contribute to this result: engagement of GPIIb/IIIa receptors with fibrinogen on the abluminal surface leading to the loss of luminal receptors, and loss of luminal filopodia that interact with thromboerythrocytes. These data provide insight into the differences in thrombogenicity between surfaces, and may provide a mechanism for purposefully passivating platelet-reactive artificial surfaces.
authors.affiliation: Division of Hematology, State University of New York at Stony Brook 11794-8151.
authors.forename: B S
authors.initials: BS
authors.lastname: Coller
authors.forename: J L
authors.initials: JL
authors.lastname: Kutok
authors.forename: L E
authors.initials: LE
authors.lastname: Scudder
authors.forename: D K
authors.initials: DK
authors.lastname: Galanakis
authors.forename: S M
authors.initials: SM
authors.lastname: West
authors.forename: G S
authors.initials: GS
authors.lastname: Rudomen
authors.forename: K T
authors.initials: KT
authors.lastname: Springer
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003432:Cross-Linking Reagents; D004912:Erythrocytes / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D009842:Oligopeptides; D010973:Platelet Adhesiveness / Q000187:drug effects; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D009842:Oligopeptides; D010980:Platelet Membrane Glycoproteins; D005340:Fibrinogen