Spreading of platelets on fibrin is mediated by the amino terminus of the beta chain including peptide beta 15-42.
We have investigated the adhesion and spreading of platelets on polymerized fibrin of varying structure to identify sites that mediate these interactions. Fibrin was prepared with thrombin to remove both fibrinopeptide A (FPA) and fibrinopeptide B (FPB) and with reptilase or Agkistrodon contortrix thrombin-like enzyme (ACTE) to selectively remove FPA or FPB, respectively. Residual fibrin-bound enzymes were inhibited with D-phenylalanyl-L-prolyl-L-arginyl chloromethyl ketone (PPACK). Platelet adhesion was independent of fibrinopeptide cleavage and was equal on fibrin prepared with each of the three enzymes. In contrast, FPB cleavage increased spreading as quantitated by fluorescence microscopy of platelets stained for glycoprotein IIb-IIIa. The 24% +/- 4% spreading on reptilase-fibrin was significantly less than the 70% +/- 8% on thrombin-fibrin or 65% +/- 9% on ACTE-fibrin (P < .0005 for both). Protease III from Crotalus atrox venom was used to specifically cleave residues B beta 1-42 from fibrinogen to further investigate the role of the beta chain N-terminus in promoting platelet spreading. After clotting with thrombin, this fibrin derivative lacked beta 15-42 and supported significantly less spreading. A monoclonal antibody (MoAb) reactive with beta 15-21 inhibited spreading on thrombin-fibrin as did peptide beta 15-42, while control MoAbs and peptides had no significant effect. These results indicate that adhesion and spreading of platelets on fibrin are mediated by different interactions, and that spreading can be mediated by FPB cleavage and the amino terminus of the beta chain including residues beta 15-42.
Hamaguchi
M
M
Department of Medicine, University of Rochester School of Medicine and Dentistry, NY.
Bunce
L A
LA
Sporn
L A
LA
Francis
C W
CW
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects; D004591:Electrophoresis, Polyacrylamide Gel; D005337:Fibrin / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D005340:Fibrinogen / Q000302:isolation & purification; D005344:Fibrinopeptide A / Q000302:isolation & purification; D005345:Fibrinopeptide B / Q000302:isolation & purification; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis / Q000494:pharmacology; D010973:Platelet Adhesiveness / Q000187:drug effects / Q000502:physiology*; D012701:Serotonin / Q000097:blood
D000590:Amino Acid Chloromethyl Ketones; D000911:Antibodies, Monoclonal; D009842:Oligopeptides; D005344:Fibrinopeptide A; D012701:Serotonin; D005345:Fibrinopeptide B; D005337:Fibrin; D005340:Fibrinogen; C049554:glycyl-arginyl-glycyl-aspartyl-serine; C040308:phenylalanyl-prolyl-arginine-chloromethyl ketone