Two sites (23-30, 76-90) on rat P-selectin mediate thrombin activated platelet-neutrophil interactions.
The lectin-like domain of P-selectin, an adhesive receptor (also known as PADGEM, GMP-140 or CD62) is implicated in platelet or endothelial cell interactions with leukocytes. The aim of this study was to characterize the lectin-like domain of rat P-selectin by the use of synthetic peptides. The lectin and EGF-like domains of rat P-selectin were cloned in our laboratory and shown to present very strong homologies to its human counterpart. Peptides corresponding with the lectin-like domain of P-selectin were tested for their ability to inhibit thrombin-activated platelets rosetting to neutrophils. Peptides 23-30 (A) and 76-90 (C), but not peptide 51-61 (B), inhibited thrombin activated rat platelets interactions with rat neutrophils (A = 33%, C = 46%, P < 0.05). Using a combination of peptides (A+B = 35%, P = 0.008 and A+C = 62%, P < 0.001), we observe different degrees of inhibition of platelets binding to neutrophils. The IC50 of peptides A+C was 0.11 mM. LYP-20, an anti-human P-selectin monoclonal antibody, was also observed to inhibit thrombin-activated rat platelets binding to rat neutrophils in a very significant manner (57% of inhibition, P < 0.001). Moreover, heparin inhibited thrombin-stimulated platelet/neutrophils rosetting (36% of inhibition, P < 0.01). These results show the importance of two sites (23-30 and 76-90) on the lectin-like domain of P-selectin in mediating platelet-neutrophil interactions in rats. Such peptides may be potent in vivo inhibitors of cell-cell interactions involving P-selectin.
Chignier
E
E
INSERM U 331, Faculté de Médecine Alexis Carrel, Institut Pasteur de Lyon, France.
Sparagano
M H
MH
McGregor
L
L
Thillier
A
A
Pellecchia
D
D
McGregor
J L
JL
D000818:Animals; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000502:physiology*; D002450:Cell Communication / Q000187:drug effects*; D003001:Cloning, Molecular; D009504:Neutrophils / Q000502:physiology*; D019007:P-Selectin; D010446:Peptide Fragments / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry* / Q000502:physiology*; D051381:Rats; D012397:Rosette Formation; D013917:Thrombin / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D019007:P-Selectin; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D013917:Thrombin