A member of the selectin family (GMP-140/PADGEM) is expressed on thrombin-stimulated rat platelets in vitro.
1. Granule membrane protein (GMP-140) is an integral alpha-granule membrane glycoprotein, expressed on the surface of human platelets following degranulation, and is part of a new family of adhesion molecules (selectins) related to the endothelial leukocyte adhesion molecule (ELAM-1) and to the lymphocyte homing receptors in man (Leu-8/TQ1) and in mouse (gp90MEL-14). 2. The cross-reactivity with rat platelets of the monoclonal antibodies (MAb), LYP20 and S12, directed against human GMP-140 was examined, with the purpose of assessing the homology of GMP-140 between human and rat platelets and of using positive MAbs to detect platelet activation in vivo in response to vascular disease in rats. 3. By ELISA technique, LYP20 gave a greater OD reading with thrombin-stimulated rat platelets than with resting platelets. 4. 125I-LYP20 bound significantly more to thrombin-stimulated rat platelets (3875 +/- 750 molecules/platelet) than to resting platelets (645 +/- 240 molecules/platelet, P less than 0.01) with 50% maximum binding at 0.13 +/- 0.02 microgram/ml; 125I-S12 did not bind to rat platelets. 5. By fluorescence-activated flow cytometry there were significantly more fluorescent thrombin-stimulated platelets (56 +/- 7% of total), compared with resting platelets (8 +/- 1% of total, P less than 0.001). 6. Western blots of rat platelet lysates showed that LYP20 bound to a single band identified, under non-reducing conditions, as having the same apparent M(r) as GMP-140. 7. LYP20 immunoprecipitated a protein which became radiolabelled on the surface of thrombin-activated rat platelets; S12 did not recognize any protein.(ABSTRACT TRUNCATED AT 250 WORDS)
Winocour
P D
PD
INSERM Unit 331, Faculté de Médecine, Institut Pasteur, Lyon, France.
Chignier
E
E
Parmentier
S
S
McGregor
J L
JL
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D015703:Antigens, CD / Q000276:immunology / Q000378:metabolism*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D008297:Male; D019007:P-Selectin; D015539:Platelet Activation; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats; D011919:Rats, Inbred Strains; D013917:Thrombin / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins; D013917:Thrombin