Structural and functional characterization of the mouse von Willebrand factor receptor GPIb-IX with novel monoclonal antibodies.
Five novel monoclonal antibodies (mAbs; p0p 1-5) were used to characterize the structural and functional properties and the in vivo expression of the murine GPIb-IX complex (von Willebrand factor receptor). The molecular weights of the subunits are similar to the human homologs: GPIbalpha (150 kd), GPIbbeta (25 kd), and GPIX (25 kd). Activation of platelets with thrombin or PMA predominantly induced shedding of glycocalicin (GC; 130 kd) but only low levels of receptor internalization. The GC concentration in normal mouse plasma was found to be at least 10 times higher than that described for human plasma (approximately 25 microg/mL versus 1-2 microg/mL). Two additional cleavage sites for unidentified platelet-derived proteases were found on GPIbalpha, as demonstrated by the generation of 3 N-terminal fragments during in vitro incubation of washed platelets (GC, 60 kd, 45 kd). Occupancy of GPIbalpha with p0p mAbs or F(ab)(2)-fragments resulted in aggregate formation in vitro and rapid irreversible thrombocytopenia in vivo, irrespective of the exact binding epitopes of the individual antibodies. GPIb-IX was not detectable immunohistochemically on endothelial cells in the major organs under normal or inflammatory conditions. The authors conclude that the mouse system might become an interesting model for studies on GPIb-IX function and regulation.
Bergmeier
W
W
Department of Molecular Oncology, General Surgery, University of Witten-Herdecke, Wuppertal, Germany.
Rackebrandt
K
K
Schröder
W
W
Zirngibl
H
H
Nieswandt
B
B
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D004730:Endothelium, Vascular / Q000737:chemistry; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D015539:Platelet Activation / Q000187:drug effects; D010974:Platelet Aggregation / Q000187:drug effects; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000032:analysis; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry* / Q000276:immunology / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000276:immunology / Q000502:physiology; D013047:Specific Pathogen-Free Organisms; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013921:Thrombocytopenia / Q000209:etiology; D014657:Vasculitis / Q000378:metabolism
D000911:Antibodies, Monoclonal; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; C023592:glycocalicin; C028254:von Willebrand factor receptor; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate