A subpopulation of platelets responds to thrombin- or SFLLRN-stimulation with binding sites for factor IXa.
Strong agonists cause platelets to expose a procoagulant surface supporting the assembly of two important coagulation enzyme complexes. Equilibrium binding has determined the density of high affinity saturable factor IXa binding sites to be 500-600 sites/platelet. We have now used flow cytometry to visualize the binding of factor IX and IXa to thrombin- or SFLLRN-activated platelets. Concentrations of these agonists that are half-maximal or maximal in kinetic studies resulted in only a small subpopulation (4-20%) of platelets binding factor IX or IXa with the density of binding sites for factor IX being about half of that for factor IXa, consistent with previous equilibrium binding studies. A small subpopulation (5 +/- 1.5%) of platelets stimulated with either agonist also exposed annexin V binding sites, and this subpopulation of platelets also bound factor IXa. Annexin V decreased factor IXa binding in the presence or absence of factor VIIIa, and factor IXa could also decrease annexin V binding on some platelets indicating a common binding site in agreement with previous studies. All platelets binding factor IXa were positive for glycoprotein IX, at the same glycoprotein IX surface density as seen in platelets negative for factor IXa binding. These studies refine the results from equilibrium binding studies and suggest that, on average, only a small subpopulation (approximately 10%) of PAR 1-stimulated platelets expose approximately 6000 factor IXa binding sites/platelet.
London
Fredda S
FS
Sol Sherry Thrombosis Research Center, Department of Biochemistry, Temple University School of Medicine, 3400 N. Broad Street, Philadelphia, PA 19140, USA. flon@temple.edu
Marcinkiewicz
Mariola
M
Walsh
Peter N
PN
D017304:Annexin A5 / Q000494:pharmacology; D001665:Binding Sites; D001792:Blood Platelets / Q000187:drug effects*; D003029:Coagulants / Q000494:pharmacology; D004396:Coloring Agents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005164:Factor IX / Q000378:metabolism; D015949:Factor IXa / Q000737:chemistry*; D005170:Factor X / Q000378:metabolism; D015945:Factor XIa / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D020128:Inhibitory Concentration 50; D007700:Kinetics; D008024:Ligands; D010446:Peptide Fragments / Q000494:pharmacology*; D011485:Protein Binding; D044463:Receptor, PAR-1 / Q000378:metabolism; D018179:Receptors, Thrombin / Q000378:metabolism; D013917:Thrombin / Q000494:pharmacology*
D017304:Annexin A5; D003029:Coagulants; D004396:Coloring Agents; D008024:Ligands; D010446:Peptide Fragments; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; C082835:thrombin receptor peptide (42-47); D005164:Factor IX; D005170:Factor X; D015949:Factor IXa; D015945:Factor XIa; D013917:Thrombin