title: The glycoprotein Ib-IX-V complex contributes to tissue factor-independent thrombin generation by recombinant factor VIIa on the activated platelet surface.
abstract: Several lines of evidence suggest that recombinant factor VIIa (rFVIIa) is able to activate factor X on an activated platelet, in a tissue factor-independent manner. We hypothesized that, besides the anionic surface, a receptor on the activated platelet surface is involved in this process. Here, we showed that, in an ELISA setup, a purified extracellular fragment of GPIbalpha bound to immobilized rFVIIa. Surface plasmon resonance established a affinity constant (K(d)) of approximately 20 nM for this interaction. In addition, CHO cells transfected with the GPIb-IX-V complex could adhere to immobilized rFVIIa, whereas wild-type CHO cells could not. Furthermore, platelets sti-mulated with a combination of collagen and thrombin adhered to immobilized rFVIIa under static conditions. Platelet adhesion was inhibited by treatment with O-sialoglycoprotein endopeptidase, which specifically cleaves GPIbalpha from the platelet surface. In addition, rFVIIa-mediated thrombin generation on the activated platelet surface was inhibited by cleaving GPIbalpha from its surface. In summary, 3 lines of evidence showed that rFVIIa interacts with the GPIb-IX-V complex, and this interaction enhanced tissue factor-independent thrombin generation mediated by rFVIIa on the activated platelet surface. The rFVIIa-GPIbalpha interaction could contribute to cessation of bleeding after administration of rFVIIa to patients with bleeding disorders.
authors.affiliation: Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Cees
authors.initials: C
authors.lastname: Weeterings
authors.forename: Philip G
authors.initials: PG
authors.lastname: de Groot
authors.forename: Jelle
authors.initials: J
authors.lastname: Adelmeijer
authors.forename: Ton
authors.initials: T
authors.lastname: Lisman
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism*; D003094:Collagen / Q000737:chemistry; D006224:Cricetinae; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015942:Factor VIIa / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D015539:Platelet Activation; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D020349:Surface Plasmon Resonance; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D019038:Platelet Glycoprotein GPIb-IX Complex; D011994:Recombinant Proteins; D003094:Collagen; C103587:recombinant FVIIa; D015942:Factor VIIa; D013917:Thrombin