Purification of the Porcine Platelet GP IIb-IIIa Complex and the Propolypeptide of von Willebrand Factor

Abstract

SummaryPlatelet membrane glycoproteins (GP) are involved in platelet adhesion and aggregation. The glycoprotein IIb-IIIa complex (GP IIbIIIa) is a Ca2+-dependent heterodimer that binds fibrinogen and other adhesive proteins, thereby mediating platelet aggregation and adhesion. We have purified two major glycoproteins from pig platelets by Concanavalin A-Sepharose, Heparin-Sepharose and Sephacryl S-300 HR chromatography (Fitzgerald et al. Anal Biochem, 1985): i) the GP IIb-IIIa complex, GP IIb Mr = 140,000 and GP IIIa a single chain of Mr = 95,000-100,000; and ii) a predominant glycoprotein of high molecular weight, the propolypeptide of von Willebrand factor (Mr = 80,000-100,000). Western-blot analysis of the purified GP IIb-IIIa showed that only certain monoclonal antibodies against the human receptor specifically recognize the porcine complex. Differences between the porcine and human GP IIb-IIIa glycoproteins could partially explain the decreased inhibitory effects of GP IIb/IIIa-antagonists (against the human receptor) in porcine platelets.

Effect of gemfibrozil on peripheral atherosclerosis and platelet activation in a pig model of hyperlipidemia

“…Before immunisation, rabbits were bled in order to obtain preimmune γ‐globulins and polyclonal antibodies against pig fibrinogen were prepared by conventional methods [ 21]. Purified polyclonal antibodies (γ‐globulins) against pig fibrinogen ( pabBp29 and pabBp30 ) were characterised by ELISA and Western‐blot.…”

Differential Kinetics of Cell Surface Loss of von Willebrand Factor and Its Propolypeptide after Secretion from Weibel-Palade Bodies in Living Human Endothelial Cells

“…Previous biochemical studies have shown that although proregion does associate transiently with VWF inside the secretory pathway (10), it does not associate with extracellular VWF, matrix components, or the endothelial cell surface, indicating that it is unlikely to play an important role in endothelial-platelet adhesion (9). A putative role in regulating collagen-induced platelet-platelet aggregation has been suggested (11,12), and it may also act as a ligand for the VLA-4 (Very Late Antigen-4) integrin present on monocytes (13). VLA-4 plays an important role in the recruitment of monocytes into the blood vessel wall and is implicated in the vascular accumulation of monocytes that marks the early development of atherosclerotic lesions (14).…”

Atherosclerosis and Thrombosis: Lessons from Animal Models

“…Western-blot analysis of purified GP IIb-IIIa showed that only certain monoclonal antibodies with epitopes on fragments of the human receptor specifically recognized the porcine receptor. These differences help explain the decreased inhibitory effects of antihuman anti-IIb-IIIa antagonists when used as pharmacological antithrombotic drugs in pigs (122)(123)(124).…”