Flow studies on human GPVI-deficient blood under coagulating and noncoagulating conditions

Abstract

The role of glycoprotein VI (GPVI) in platelets was investigated in 3 families bearing an insertion within the GP6 gene that introduces a premature stop codon prior to the transmembrane domain, leading to expression of a truncated protein in the cytoplasm devoid of the transmembrane region. Western blotting and flow cytometry of GP6hom (homozygous) platelets confirmed loss of the full protein. The level of the Fc receptor γ-chain, which associates with GPVI in the membrane, was partially reduced, but expression of other receptors and signaling proteins was not altered. Spreading of platelets on collagen and von Willebrand factor (which supports partial spreading) was abolished in GP6hom platelets, and spreading on uncoated glass was reduced. Anticoagulated whole blood flowed over immobilized collagen or a mixture of von Willebrand factor, laminin, and rhodocytin (noncollagen surface) generated stable platelet aggregates that express phosphatidylserine (PS). Both responses were blocked on the 2 surfaces in GP6hom individuals, but adhesion was not altered. Thrombin generation was partially reduced in GP6hom blood. The frequency of the GP6het (heterozygous) variant in a representative sample of the Chilean population (1212 donors) is 2.9%, indicating that there are ∼4000 GP6hom individuals in Chile. These results demonstrate that GPVI supports aggregation and PS exposure under flow on collagen and noncollagen surfaces, but not adhesion. The retention of adhesion may contribute to the mild bleeding diathesis of GP6hom patients and account for why so few of the estimated 4000 GP6hom individuals in Chile have been identified.

Assessment of thrombotic risk during long-term treatment of immune thrombocytopenia with fostamatinib

“… 38 Conversely, patients with genetic deficiency in GPVI did not have significant disruption of hemostasis or other platelet activities. 39 , 40 In vivo experiments have shown that SYK deficiency or administration of the active metabolite of fostamatinib (R406) and other SYK inhibitors has a negligible effect on hemostasis. 20 , 41 Importantly, in platelet-mediated hemostasis, platelet activation and aggregation are also stimulated by thrombin, thromboxane A 2 , and ADP, which all signal through G protein-coupled receptors that are not dependent on signaling via SYK.…”

Platelet calcium signaling by G-protein coupled and ITAM-linked receptors regulating anoctamin-6 and procoagulant activity

“…This feature makes GPVI an attractive target for the development of improved anti-platelet drugs to prevent thrombosis. Indeed, GPVI-deficient patients described so far present with no more than a minor bleeding tendency [43,47]. Interestingly, a genetic variant of GP6 was found to associate with an increased risk of venous thrombosis [48].…”

Platelet activation by charged ligands and nanoparticles: platelet glycoprotein receptors as pattern recognition receptors

“…77 In addition, DEP-mediated activation has previously been shown to be abolished in platelets from GPVI-deficient mice. 1 Here, we expanded on this to show negatively charged DEPs (20-70 nm) and negative charged surfaces also activate human platelets through GPVI, using platelets from homozygous patients (Figure 3A), who have a mutation in the GP6 gene preventing expression 78 (see Supplementary Materials for full patient details). These patients have no GPVI expression as shown by flow cytometry and western blotting, whereas levels of other glycoprotein receptors, including CLEC-2, GPIIb/IIIa and FcγRIIA are within the normal range.…”