Anti-glycoprotein VI monoclonal antibodies directly aggregate platelets independently of FcγRIIa and induce GPVI ectodomain shedding

Mohammad Al-Tamimi, Fi Tjen Mu, Jane F. Arthur, Yang Shen, Masaaki Moroi, Michael C. Berndt, Robert K. Andrews, Elizabeth E. Gardiner

Research output: Contribution to journalArticlepeer-review

37 Citations (Scopus)

Abstract

Adhesion of circulating platelets to the blood vessel wall initiates thrombus formation in haemostasis and thrombotic disease. The platelet collagen receptor, glycoprotein (GP) VI, is critical for thrombus formation at arterial shear rates and is a potential therapeutic target for anti-thrombotic drugs. In this study, we evaluate eight newly-derived, purified murine anti-human GPVI monoclonal antibodies (mAbs) for their effect on GPVI-dependent platelet aggregation and GPVI ectodomain shedding. All mAbs were raised against the ligand-binding GPVI ectodomain encompassing two immunoglobulin domains (residues 21-234, excluding the signal sequence) and recognized full-length GPVI in human platelet lysates by western blotting. The majority of antibodies induced aggregation in both human platelet-rich plasma (PRP) and washed platelets independently of the Fc receptor, FcγRIIa (not inhibited by the blocking anti-FcγRIIa mAb, IV.3), whereas one mAb (11A7) neither induced aggregation nor inhibited aggregation in response to GPVI ligands, collagen, and collagen-related peptide (CRP). In contrast, Fab fragments of mAb 12A5 strongly blocked collagen- and CRP-, but not convulxin-induced aggregation. In addition, it is shown for the first time in vitro that anti-GPVI mAbs can induce metalloproteinase-dependent ectodomain shedding of human GPVI, generating an ∼10-kDa remnant that remained platelet-associated and an ∼55-kDa soluble fragment. In conclusion, this analysis of anti-GPVI mAbs provides useful tools for studying the functional role of platelet GPVI.

Original languageEnglish
Pages (from-to)75-82
Number of pages8
JournalPlatelets
Volume20
Issue number2
DOIs
Publication statusPublished - Mar 2009
Externally publishedYes

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