title: Ultrastructural analysis of thrombin-induced interaction between human platelets and liposomes carrying fibrinogen γ-chain dodecapeptide as a synthetic platelet substitute.
abstract: BACKGROUND
  The dodecapeptide HHLGGAKQAGDV (H12) in the carboxy-terminus of the fibrinogen γ-chain is a specific binding site of the ligand for platelet GPIIb/IIIa complex. We have evaluated liposomes carrying fibrinogen γ-chain dodecapeptide as a synthetic platelet substitute.
  
  
  OBJECTIVE
  We examined the interaction between human platelets and H12-liposomes during thrombin-induced activation using flow cytometry and electron microscopy (EM).
  
  
  RESULTS
  After thrombin-activation, a remarkable time-dependent increase in binding of the H12-liposomes to platelets was found by flow cytometry. A large-sized swollen open canalicular system (OCS) was observed in the spheroidal platelets from 60 sec to 5 min after thrombin-activation, but intact H12-liposomes were not evident by conventional EM. Cryoultramicrotomy and immunogold staining with anti-H12 antibody were successful in identifying the liposomes; they appeared as small particles with a unit membrane around 0.2 to 0.4 μm in diameter, and gold labels representing H12 were distributed homogeneously on the surface. Abundant H12-liposomes were localized not only on the surface membrane but also in the lumen of the large-sized swollen OCS in the platelets at 60 sec after thrombin-activation. The formation of the large-sized swollen OCS was inhibited by pre-incubation with unbound H12, EDTA or anti-GPIIb/IIIa antibody. In thrombin-induced platelet aggregates we observed electron-transparent areas between adherent platelets, in which abundant H12-liposomes were distributed.
  
  
  CONCLUSIONS
  We demonstrate morphologically that H12-liposomes bind to thrombin-activated platelets and accumulate between adherent platelets like fibrinogen, leading to large-scale aggregation.
authors.affiliation: Laboratory of Electron Microscopy, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan. suzuki-hd@igakuken.or.jp
authors.forename: Hidenori
authors.initials: H
authors.lastname: Suzuki
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Okamura
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Ikeda
authors.forename: Shinji
authors.initials: S
authors.lastname: Takeoka
authors.forename: Makoto
authors.initials: M
authors.lastname: Handa
mesh_terms: D001792:Blood Platelets / Q000187:drug effects* / Q000502:physiology; D001802:Blood Substitutes / Q000494:pharmacology*; D005340:Fibrinogen / Q000378:metabolism / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D008081:Liposomes / Q000097:blood / Q000494:pharmacology; D008854:Microscopy, Electron; D015539:Platelet Activation / Q000187:drug effects; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000097:blood / Q000494:pharmacology*; D017713:Platelet Transfusion; D013917:Thrombin / Q000378:metabolism*
chemical_list: D001802:Blood Substitutes; D008081:Liposomes; D010975:Platelet Aggregation Inhibitors; C027539:fibrinopeptides gamma; D005340:Fibrinogen; D013917:Thrombin