The cleaved peptide of PAR1 is a more potent stimulant of platelet-endothelial cell adhesion than is thrombin.
OBJECTIVE
Platelet-endothelial cell adhesion is an important pathologic response to vessel injury or inflammation. On binding to its endothelial or platelet G protein-linked seven-transmembrane domain receptor, protease-activated receptor-1 (PAR1), thrombin releases a 41-amino acid peptide (TR(1-41)). We examined the effect of TR(1-41) on platelet activation and on platelet-endothelial cell adhesion.


METHODS
A monolayer of confluent human saphenous vein endothelial cells was incubated with washed human platelets. Platelets were stimulated with either TR(1-41), TR(21-41), scrambled TR(1-41), adenosine diphosphate (ADP)-epinephrine (EPI), thrombin, or thrombin receptor activating peptide (TRAP). Platelet activation was identified with flow cytometry. The magnitude of platelet-endothelial cell adhesion was determined with a laser scanning cytometer that scanned the monolayer of endothelial cells and identified fluorescently bound platelets.


RESULTS
Maximal thrombin stimulation (0.1 U/mL) induced a threefold increase in platelets bound to endothelial cells compared with buffer alone. Stimulation with TR(1-41) (20 mmol/L) tripled the number of platelets bound to endothelial cells compared with thrombin. Scrambled sequence of TR(1-41) (20 mmol/L) and TR(21-41) (20 mmol/L), neither of which induces platelet activation, had minimal effect on platelet adhesion. Both TRAP (20 mmol/L) and ADP-EPI (20 mmol/L) induced less platelet-endothelial cell adhesion than did thrombin. TR(1-41)-induced platelet-endothelial cell adhesion was partially blocked by glycoprotein (GP)IIb-IIIa-specific monoclonal antibody, 10E5 (10 mg/mL).


CONCLUSIONS
TR(1-41), the cleaved peptide of PAR1, is a more potent stimulant of platelet-endothelial cell adhesion than is thrombin, TRAP, or ADP-EPI, and this adhesion is at least in part mediated by the platelet GPIIb-IIIa receptor.
Claytor
R Brannon
RB
Department of Surgery, Division of Vascular Surgery, and Center for Platelet Function Studies, University of Massachusetts Medical School, Worcester, Mass. 01655, USA. brannon@umassmed.edu
Michelson
Alan D
AD
Li
Jian-Ming
JM
Frelinger
A L
AL
Rohrer
Michael J
MJ
Garnette
Charles S C
CS
Barnard
Marc R
MR
Krueger
Lori A
LA
Furman
Mark I
MI
D002448:Cell Adhesion / Q000187:drug effects* / Q000502:physiology; D002455:Cell Division / Q000187:drug effects* / Q000502:physiology; D004727:Endothelium / Q000187:drug effects* / Q000503:physiopathology; D006490:Hemostatics / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D010446:Peptide Fragments / Q000494:pharmacology*; D010973:Platelet Adhesiveness / Q000187:drug effects* / Q000502:physiology; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin / Q000187:drug effects* / Q000502:physiology; D012501:Saphenous Vein / Q000187:drug effects / Q000503:physiopathology; D013917:Thrombin / Q000494:pharmacology*; D020246:Venous Thrombosis / Q000503:physiopathology*
D006490:Hemostatics; D010446:Peptide Fragments; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; C111242:thrombin receptor peptide (1-41); D013917:Thrombin