title: Orientation and specificity of fibrin protofibril binding to ADP-stimulated platelets.
abstract: We have investigated the molecular basis of platelet:fibrin binding by studying interactions between platelets and protofibrils, soluble two-stranded polymers of fibrin, which are intermediates on the fibrin assembly pathway. The specificity of these interactions was examined with transmission electron microscopy (TEM), which clearly showed thin fibers with lengths to 150 nm attached to the cell surface of normal, stimulated platelets. Immunogold electron microscopy using rabbit anti-human fibrinogen as the first stage antibody verified the identity of the surface-bound molecules, and the immunogold distribution paralleled that observed with the fibrin/fibrinogen molecules alone. Contacts between the ends of the fibers and the platelets were frequently observed, but lateral contacts were also evident. Given the diameter at the point of fibrin contact (18.2 +/- 1.3 nm), it is possible that several glycoprotein receptors were involved in binding each protofibril. Morphometric analyses demonstrated that normal platelets stimulated by ADP in the absence of exogenous fibrin(ogen) or in the presence of fibrin protofibrils and antibodies directed against the GPIIb/IIIa complex lacked this molecular layer on the surface. Neither protofibrils nor fibrin fibers adhered to the surface of Glanzmann's thrombasthenic platelets, as demonstrated by TEM and microfluorimetry. Synthetic peptides of sequence RGDS and HHLGGAKQAGDV effectively blocked the binding of protofibrils to the surface of normal, stimulated platelets while synthetic GHRP had no effect. These results provide direct evidence for multiple points of attachment between fibrin protofibrils and the glycoprotein IIb/IIIa complexes present in a functional conformation on the surface of normal, stimulated platelets.
authors.affiliation: Department of Pathology, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, NC 27103.
authors.forename: J C
authors.initials: JC
authors.lastname: Lewis
authors.forename: C
authors.initials: C
authors.lastname: Johnson
authors.forename: P
authors.initials: P
authors.lastname: Ramsamooj
authors.forename: R R
authors.initials: RR
authors.lastname: Hantgan
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000906:Antibodies / Q000276:immunology; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005337:Fibrin / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D008854:Microscopy, Electron; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism; D010980:Platelet Membrane Glycoproteins / Q000276:immunology; D011487:Protein Conformation; D013915:Thrombasthenia / Q000097:blood / Q000473:pathology
chemical_list: D000906:Antibodies; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D000244:Adenosine Diphosphate; D005337:Fibrin