Maintenance of GPIIb-IIIa avidity supporting "irreversible" fibrinogen binding is energy-dependent.
The interaction between fibrinogen and GPIIb-IIIa on stimulated platelets is multiphasic, progressing from reversible to irreversible ligand binding, associated with stabilization of platelet aggregates and clot retraction. Because fibrinogen binding to platelets has been linked to "outside-in" signaling events such as postreceptor occupancy protein tyrosine kinase and phosphatidylinositol-3 kinase activation, this study examined intracellular signaling requirements involved in stabilizing 125I-fibrinogen binding to adenosine diphosphate-treated platelets with selective inhibitors of protein tyrosine kinase (herbimycin A) (10 micromol/L) and phosphatidylinositol-3 kinase (Wortmannin) (10 nmol/L) and metabolic inhibitors antimycin A (7.3 micromol/L) and 2 deoxyglucose (6 mmol/L). Preincubation of platelets with herbimycin A or Wortmannin inhibited fibrinogen binding by 80% to 92% and was accompanied by markedly decreased tyrosine phosphorylation of a range of proteins migrating between 60 kDa and 125 kDa. The addition of inhibitors 5 minutes after adenosine diphosphate-induced fibrinogen binding also resulted in decreased tyrosine phosphorylation and dissociation of approximately 50% of bound fibrinogen within 60 minutes but failed to cause dissociation of irreversibly bound fibrinogen. In contrast, platelet exposure to metabolic inhibitors 5 minutes or 60 minutes after fibrinogen binding resulted in complete, spontaneous fibrinogen dissociation. These data suggest that the maintenance of GPIIb-IIIa avidity supporting irreversible fibrinogen binding to intact platelets is not affected by inhibitors of protein tyrosine kinase or phosphatidylinositol-3 kinase but involves other energy-dependent pathways.
Peerschke
E I
EI
Department of Pathology, Weill Medical College of Cornell University, New York, New York, USA.
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000911:Antibodies, Monoclonal; D000963:Antimetabolites / Q000494:pharmacology; D000968:Antimycin A / Q000494:pharmacology; D016227:Benzoquinones; D001792:Blood Platelets / Q000201:enzymology / Q000276:immunology; D003847:Deoxyglucose / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004734:Energy Metabolism / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005079:Excipients / Q000494:pharmacology; D005340:Fibrinogen / Q000378:metabolism*; D006801:Humans; D047029:Lactams, Macrocyclic; D010766:Phosphorylation; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D011107:Polylysine; D011485:Protein Binding / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D015398:Signal Transduction / Q000502:physiology; D014443:Tyrosine / Q000378:metabolism; D000077191:Wortmannin
D000730:Androstadienes; D000900:Anti-Bacterial Agents; D000911:Antibodies, Monoclonal; D000963:Antimetabolites; D016227:Benzoquinones; D004791:Enzyme Inhibitors; D005079:Excipients; D047029:Lactams, Macrocyclic; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D011809:Quinones; D017828:Rifabutin; D011107:Polylysine; D014443:Tyrosine; D000244:Adenosine Diphosphate; D000968:Antimycin A; C020754:herbimycin; D005340:Fibrinogen; D003847:Deoxyglucose; D011505:Protein-Tyrosine Kinases; D000077191:Wortmannin; D004121:Dimethyl Sulfoxide