title: Tissue factor-enriched vesicles are taken up by platelets and induce platelet aggregation in the presence of factor VIIa.
abstract: We investigated the interactions of vesicles containing human tissue factor (TF) with platelets and evaluated responses induced by rFVIIa using standard aggregometry, ultrastructural and flow-cytometry techniques. Washed platelets were exposed to a preparation of placental human TF (pTF) or to a relipidated formulation of recombinant human TF (rTF). Under stirring conditions, pTF induced reversible aggregation with platelets returning to their resting state after 5 minutes. This reversible response to pTF was partially inhibited by antibodies against CD62-P, but not by antithrombin agents, and was not observed with rTF. Sequential ultrastructural studies revealed uptake of both TF preparations by platelets involving traffic of vesicles through channels of the open canalicular system (OCS). Immunocytochemical studies on cryosections identified TF in the OCS, and occasionally in the alpha-granules of the platelets. These processes were faster with pTF than with rTF, but both TF preparations accumulated in platelets at the end of incubation periods. Flow cytometry studies revealed the presence of other cellular antigens (CD62-P, CD14 and CD45) associated to the pTF. Addition of rFVIIa to washed platelets exposed to pTF or rTF, caused a thrombin dependent irreversible platelet aggregation. Our studies demonstrate that platelets possess mechanisms to capture and incorporate TF-rich vesicles. These processes are accelerated by the presence of other cellular antigens in the vesicles. Our findings may explain the hemostatic action of rFVIIa in severely hemodiluted patients, but are also relevant for the understanding of potential implications of TF-associated to platelets in the propagation of thrombus.
authors.affiliation: Servicio de Hemoterapia-Hemostasia, Hospital Clinic, Centre de Deagnostic biomedic, IDIBAPS, Universidad de Barcelona, Spain.
authors.forename: Irene
authors.initials: I
authors.lastname: Lopez-Vilchez
authors.forename: Gines
authors.initials: G
authors.lastname: Escolar
authors.forename: Maribel
authors.initials: M
authors.lastname: Diaz-Ricart
authors.forename: Berta
authors.initials: B
authors.lastname: Fuste
authors.forename: Ana M
authors.initials: AM
authors.lastname: Galan
authors.forename: James G
authors.initials: JG
authors.lastname: White
mesh_terms: D017304:Annexin A5 / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism; D001792:Blood Platelets / Q000276:immunology / Q000378:metabolism* / Q000648:ultrastructure; D015942:Factor VIIa / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D017493:Leukocyte Common Antigens / Q000378:metabolism; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008854:Microscopy, Electron; D019007:P-Selectin / Q000378:metabolism; D010316:Particle Size; D010743:Phospholipids / Q000378:metabolism; D010924:Placental Extracts / Q000737:chemistry / Q000378:metabolism; D010974:Platelet Aggregation; D010979:Platelet Function Tests; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D060149:Tetraspanin 30; D013925:Thromboplastin / Q000378:metabolism*; D013997:Time Factors
chemical_list: D017304:Annexin A5; D015703:Antigens, CD; C554586:CD63 protein, human; D018950:Lipopolysaccharide Receptors; D008081:Liposomes; D019007:P-Selectin; D010743:Phospholipids; D010924:Placental Extracts; D010980:Platelet Membrane Glycoproteins; D011994:Recombinant Proteins; D060149:Tetraspanin 30; D013925:Thromboplastin; D017493:Leukocyte Common Antigens; D015942:Factor VIIa