title: Fibrinogen and von Willebrand factor-independent platelet aggregation in vitro and in vivo.
abstract: BACKGROUND
  Fibrinogen (Fg) has been considered essential for platelet aggregation. However, we recently demonstrated formation of occlusive thrombi in Fg-deficient mice and in mice doubly deficient for Fg and von Willebrand factor (Fg/VWF(-/-)).
  
  
  RESULTS
  Here we studied Fg/VWF-independent platelet aggregation in vitro and found no aggregation in citrated platelet-rich plasma of Fg/VWF(-/-) mice. Surprisingly, in Fg/VWF(-/-) plasma without anticoagulant, adenosine diphosphate induced robust aggregation of Fg/VWF(-/-) platelets but not of beta(3)-integrin-deficient (beta(3) (-/-)) platelets. In addition, beta(3) (-/-) platelets did not significantly incorporate into thrombi in Fg/VWF(-/-) mice. This Fg/VWF-independent aggregation was blocked by thrombin inhibitors (heparin, hirudin, PPACK), and thrombin or thrombin receptor activation peptide (AYPGKF-NH(2)) induced aggregation of gel-filtered Fg/VWF(-/-) platelets in 1 mm Ca(2+) PIPES buffer. Notably, aggregation in PIPES buffer was only 50-60% of that observed in Fg/VWF(-/-) plasma. Consistent with the requirement for thrombin in vitro, hirudin completely inhibited thrombus formation in Fg/VWF(-/-) mice. These data define a novel pathway of platelet aggregation independent of both Fg and VWF. Although this pathway was not detected in the presence of anticoagulants, it was observed under physiological conditions in vivo and in the presence of Ca(2+)in vitro.
  
  
  CONCLUSIONS
  beta(3) integrin, thrombin, and Ca(2+) play critical roles in this Fg/VWF-independent aggregation, and both plasma and platelet granule proteins contribute to this process.
authors.affiliation: Department of Laboratory Medicine and Pathobiology, St Michael's Hospital, University of Toronto, Ottawa, ON, Canada.
authors.forename: H
authors.initials: H
authors.lastname: Yang
authors.forename: A
authors.initials: A
authors.lastname: Reheman
authors.forename: P
authors.initials: P
authors.lastname: Chen
authors.forename: G
authors.initials: G
authors.lastname: Zhu
authors.forename: R O
authors.initials: RO
authors.lastname: Hynes
authors.forename: J
authors.initials: J
authors.lastname: Freedman
authors.forename: D D
authors.initials: DD
authors.lastname: Wagner
authors.forename: H
authors.initials: H
authors.lastname: Ni
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D005340:Fibrinogen / Q000737:chemistry* / Q000235:genetics*; D006629:Hirudins / Q000494:pharmacology; D066298:In Vitro Techniques; D039661:Integrin beta3 / Q000502:physiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D010974:Platelet Aggregation; D013917:Thrombin / Q000502:physiology*; D013927:Thrombosis; D014841:von Willebrand Factor / Q000737:chemistry* / Q000235:genetics*
chemical_list: D006629:Hirudins; D039661:Integrin beta3; D014841:von Willebrand Factor; D005340:Fibrinogen; D013917:Thrombin; D002118:Calcium