title: Functional significance of adenosine 5'-diphosphate receptor (P2Y(12)) in platelet activation initiated by binding of von Willebrand factor to platelet GP Ibalpha induced by conditions of high shear rate.
abstract: BACKGROUND
  The role of the adenosine 5'-diphosphate receptor P2Y(12) in platelet activation initiated by the von Willebrand factor (VWF)-GP Ibalpha interaction under high shear rate was investigated.
  
  
  RESULTS
  Blood samples were obtained from 11 donors. Shear-induced platelet aggregation was detected by optically modified cone-plate viscometer. Shear-induced VWF binding, P-selectin expression, and microparticle release were detected by flow cytometry. Platelet interaction with immobilized VWF was also investigated by parallel-plate flow chamber equipped with epifluorescent videomicroscopy. Effects of a selective P2Y(12) antagonist AR-C69931 MX were tested. AR-C69931 MX inhibited shear-induced platelet aggregation in a dose-dependent manner, achieving the maximum inhibition at 100 nmol/L. The extent of aggregation after exposure to a shear rate of 10 800 s(-1) for 6 minutes in the presence of 100 nmol/L AR-C69931 MX was 32.4+/-8.2% (mean+/-SD), which was significantly lower than the value in the controls of 69.7+/-9.6% (P<0.01). The inhibiting effects of AR-C69931 MX were reversed by exogenous addition of adenosine 5'-diphosphate. Shear-induced VWF binding and P-selectin surface translocation, which occurred in 4696+/-911 and 5964+/-784, respectively, of 10 000 measured platelets, was also inhibited by AR-C69931 MX (100 nmol/L) to 1948+/-528 and 2797+/-718, respectively (P=0.0018 and P=0.0009). Microparticle release was similarly inhibited. In a flow chamber experiment, firm platelet attachment on immobilized VWF was inhibited by AR-C69931 MX, whereas transient interaction was not influenced. All the above reactions were completely inhibited by blocking VWF-GP Ibalpha interaction.
  
  
  CONCLUSIONS
  We have demonstrated that the stimulation of P2Y(12) is involved in platelet activation initiated by the binding of VWF to GP Ibalpha induced by a high shear rate.
authors.affiliation: Division of Cardiology, Department of Medicine, Tokai University School of Medicine, Kanagawa, Japan. shinichi@is.icc.u-tokai.ac.jp
authors.forename: Shinya
authors.initials: S
authors.lastname: Goto
authors.forename: Noriko
authors.initials: N
authors.lastname: Tamura
authors.forename: Koji
authors.initials: K
authors.lastname: Eto
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Ikeda
authors.forename: Shunnosuke
authors.initials: S
authors.lastname: Handa
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000249:Adenosine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D008565:Membrane Proteins; D008856:Microscopy, Fluorescence; D018715:Microscopy, Video; D019007:P-Selectin / Q000096:biosynthesis; D015539:Platelet Activation / Q000187:drug effects / Q000502:physiology*; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism*; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058925:Receptors, Purinergic P2Y12; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D019007:P-Selectin; C548576:P2RY12 protein, human; D010975:Platelet Aggregation Inhibitors; D019038:Platelet Glycoprotein GPIb-IX Complex; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D014841:von Willebrand Factor; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; C117446:cangrelor