Is β₂-glycoprotein I prothrombotic?

In this issue of Blood, Kulkarni et al report that mice deficient in β₂-glycoprotein I (β2GPI) are relatively protected from thrombosis.¹ They suggest that this decreased propensity toward thrombosis is mediated by reduced protease-activated receptor 3 (PAR3)-facilitated platelet activation.

β2GPI is an abundant plasma glycoprotein and major autoantigen in antiphospholipid antibody syndrome (APS). APS is an autoimmune disease that presents with thrombosis or obstetric complications in the presence of characteristic autoantibodies.^2,3 These autoantibodies include the lupus anticoagulant, anticardiolipin antibodies, and anti-β2GPI antibodies.^2,3 Anti-β2GPI antibodies are proposed to be pathogenic in APS. In support of this are mouse studies demonstrating a direct prothrombotic effect of exogenous anti-β2GPI antibodies.⁴ The mechanism by which anti-β2GPI antibodies mediate a prothrombotic effect remains uncertain.³ 

A key barrier to understanding the pathophysiologic effect of anti-β2GPI antibodies in APS is a limited understanding of the physiologic role of β2GPI in coagulation. Some previous studies have suggested that β2GPI has an antithrombotic effect, and others have suggested a prothrombotic effect.^5,6 

The present study addresses the question of what role β2GPI plays in in vivo hemostasis and thrombosis. The authors generated β2GPI-deficient mice using CRISPR/Cas9 and carried out a detailed analysis of their thrombotic and bleeding phenotypes. Relative to wild-type mice, the β2GPI-deficient mice were protected from thrombosis and had prolonged tail bleeding times. These phenotypes were rescued by infusion of purified β2GPI. To determine the mechanisms responsible for protection of the β2GPI-deficient mice from thrombosis, the authors first compared their plasma coagulation laboratory parameters to those of wild-type mice; they found no significant differences. They next tested the function of platelets from the β2GPI-deficient mice. They found that the β2GPI-deficient mice had reduced PAR3-facilitated activation of platelets by thrombin. The authors hypothesize that reduced PAR3-facilitated platelet activation is the mechanism by which β2GPI deficiency leads to decreased thrombotic potential.

The findings of this study contrast with results from a number of previous studies that have suggested an antithrombotic effect of β2GPI.^5-7 Most notably, a similar study by Passam et al reported that β2GPI-null mice had a strong prothrombotic phenotype.⁶ There are several potential explanations for the discrepant findings in these 2 studies. A strength of the Kulkarni et al study is use of littermate controls from the same vivarium. They posit that differences in genotype and microbiota between the wild-type and β2GPI-deficient mice could have led to the opposite result in the Passam et al study. The impact of the intestinal microbiome on hemostasis and thrombosis has been increasingly appreciated in recent years.⁸ Thorough analyses of the intestinal microbiome composition of the mice used these studies would help to advance understanding of potential microbiome contributions to the observed phenotypes. Independent replication of these studies in other laboratories, using littermate controls, would be very useful given the conflicting reports from these 2 high-quality studies.

This study advances our understanding of the role of β2GPI in murine physiology and also raises some important questions that require additional investigation. First, what is the role of β2GPI in human hemostasis and thrombosis? Although this report suggests that β2GPI is prothrombotic in mice, differences between murine and human physiology make the results difficult to extrapolate to humans. The article suggests that β2GPI modulates murine platelet function in a manner that depends on PAR3. In mice, PAR3 is an important cofactor for thrombin-induced platelet activation. In humans, PAR3 is expressed only at very low levels in platelets, and is generally thought to have little to no role in platelet function. Further investigations are needed to determine what impact, if any, β2GPI has on the function of human platelets. Also highlighting the potential difficulties in extending the findings from these studies to humans are several human studies pointing to an anticoagulant role for β2GPI.⁷ Larger epidemiologic studies are needed to test for associations between β2GPI levels and bleeding and thrombotic outcomes. Another key question is: how do anti-β2GPI antibodies lead to a prothrombotic phenotype in APS? Supplemental data included in this report suggest that anti-β2GPI immunoglobulin G (IgG) enhances murine platelet responsiveness as compared with control IgG, but this effect is not inhibited by an anti-PAR3 antibody. This suggests that anti-β2GPI-induced platelet hyperresponsiveness is independent of PAR3. Further in vitro and in vivo studies of anti-β2GPI antibodies and their impact on the function of murine and human platelets would be useful to clarify the potential role for anti-β2GPI-induced platelet hyperresponsiveness in APS.

In summary, this report by Kulkarni et al sheds new light on the physiologic role of β2GPI, suggesting a prothrombotic function in contrast to some prior reports. Hopefully this work will stimulate further investigations into the biochemistry and in vivo functions of β2GPI and anti-β2GPI antibodies, which may lead to translationally relevant insights into the biology of APS.

Conflict-of-interest disclosure: M.A.H. declares no competing financial interests.