title: A model of platelet aggregation involving multiple interactions of thrombospondin-1, fibrinogen, and GPIIbIIIa receptor.
abstract: Thrombospondin-1 (TSP) may, after secretion from platelet alpha granules, participate in platelet aggregation, but its mode of action is poorly understood. We evaluated the capacity of TSP to form inter-platelet cross-bridges through its interaction with fibrinogen (Fg), using either Fg-coated beads or Fg bound to the activated GPIIbIIIa integrin (GPIIbIIIa*) immobilized on beads or on activated fixed platelets (AFP), i.e. in a system free of platelet signaling and secretion mechanisms. Aggregation at physiological shear rates (100-2000 s(-1)) was studied in a microcouette device and monitored by flow cytometry. Soluble TSP bound to and induced aggregation of Fg-coated beads dose-dependently, which could be blocked by the amino-terminal heparin-binding domain of TSP, TSP18. Soluble TSP did not bind to GPIIbIIIa*-coated beads or AFP, unless they were preincubated with Fg. The interaction of soluble TSP with Fg-GPIIbIIIa*-coated beads or Fg-AFP resulted in the formation of aggregates via Fg-TSP-Fg cross-bridges, as demonstrated in a system where direct cross-bridges mediated by GPIIbIIIa*-Fg on one particle and free GPIIbIIIa* on a second particle were blocked by the RGD mimetic Ro 44-9883. Soluble TSP increased the efficiency of Fg-mediated aggregation of AFP by 30-110% over all shear rates and GPIIbIIIa* occupancies evaluated. Surprisingly, TSP binding to Fg already bound to its GPIIbIIIa* receptor appears to block the ability of this occupied Fg to recognize another GPIIbIIIa* receptor, but this TSP can indeed cross-bridge to another Fg molecule on a second platelet. Finally, TSP-coated beads could directly coaggregate at shear rates from 100 to 2000 s(-1). Our studies provide a model for the contribution of secreted TSP in reinforcing inter-platelet interactions in flowing blood, through direct Fg-TSP-Fg and TSP-TSP cross-bridges.
authors.affiliation: Unité 353 INSERM, Institut d'Hématologie, Université Paris VII, Hôpital St Louis, Cedex 10, Paris, France.
authors.forename: A
authors.initials: A
authors.lastname: Bonnefoy
authors.forename: R
authors.initials: R
authors.lastname: Hantgan
authors.forename: C
authors.initials: C
authors.lastname: Legrand
authors.forename: M M
authors.initials: MM
authors.lastname: Frojmovic
mesh_terms: D000085:Acetates / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism*; D017022:Flow Injection Analysis; D008954:Models, Biological; D010974:Platelet Aggregation / Q000502:physiology*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D011485:Protein Binding; D019700:Thrombospondin 1 / Q000378:metabolism*; D019699:Thrombospondins / Q000378:metabolism; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D000085:Acetates; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D010980:Platelet Membrane Glycoproteins; D019700:Thrombospondin 1; D019699:Thrombospondins; D014443:Tyrosine; D005340:Fibrinogen; C084062:lamifiban