Binding properties and inhibition of platelet aggregation by a monoclonal antibody to CD31 (PECAM-1).
CD31/platelet endothelial cell adhesion molecule 1 (PECAM-1) is expressed on platelets, endothelial cells, myeloid cells, monocytes, and certain lymphocyte subsets. It has been shown that CD31 is involved in the homophilic and heterophilic cellular adhesion and leukocyte transendothelial migration, but little is known about the role of CD31 in platelet functions. Previously we have shown that monoclonal antibody (MAb) AAP2 produced in our laboratory bound to a 110-kD platelet antigen and gave an enhanced binding to activated platelet membrane. In this study we demonstrated that platelet lysate depleted of the antigen through adsorption by an AAP2-solidified affinity column was bound by MAbs against CD62 and CD42 but not by MAb 5.6E against CD31 or AAP2 on the immunoblot. Rabbit antibodies against CD31 completely inhibited the binding of AAP2 to platelets in the flow cytometry analysis. This indicates that AAP2 is specifically against CD31. 125I-labeled AAP2 bound to resting platelets with 5587 +/- 1765 sites/platelet and a Kd of 1 nmol/L and to thrombin-activated platelets with 17,625 +/- 4865 sites/platelet and a Kd of 0.24 nmol/L. Addition of 10 micrograms/mL AAP2 inhibited the aggregation induced by 4 mumol/L ADP by 78.6%, 6 mumol/L epinephrine by 79.4%, 1 microgram/mL collagen by 78.7%, and 0.25 U/mL thrombin by 29%. The platelet aggregation was completely restored when higher concentration of agonists were used. MAb 5.6E did not have any effect on platelet aggregation. These results suggest that AAP2 binds to a special epitope of CD31 on platelets and that CD31 is involved in platelet aggregation.
Wu
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XW
Hemophilia and Thrombosis Center/Sylvester Cancer Center, University of Miami, Fla., USA.
Lian
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EC
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D000918:Antibody Specificity; D001667:Binding, Competitive; D004837:Epinephrine / Q000494:pharmacology; D000939:Epitopes / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D010974:Platelet Aggregation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000276:immunology*; D011485:Protein Binding; D011817:Rabbits; D013917:Thrombin / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010975:Platelet Aggregation Inhibitors; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D000244:Adenosine Diphosphate; D013917:Thrombin; D004837:Epinephrine