Abstract
Objective
Results of previous studies suggest that anti–β~2~‐glycoprotein I (anti‐β~2~GPI) antibodies in complex with β~2~GPI activate platelets in a dysregulated manner, potentially contributing to the prothrombotic tendency associated with the antiphospholipid syndrome (APS). We undertook this study to investigate the possible contribution of the GPIb‐IX‐V receptor to platelet activation mediated by the anti‐β~2~GPI antibody–β~2~GPI complex.
Methods
In vitro methods were used in the present study. The interaction between β~2~GPI and the GPIbα subunit of the GPIb‐IX‐V receptor was delineated using direct binding and competitive inhibition assays. The interaction between the anti‐β~2~GPI antibody–β~2~GPI complex and platelets was studied using a novel method in which the Fc portion of the antibody was immobilized using protein A coated onto a microtiter plate. Platelet activation was assessed by two methods; one involved measuring thromboxane B~2~ production and the other involved assessment of the activation of the phosphatidylinositol 3‐kinase/Akt/glycogen synthase kinase 3β intracellular signaling pathway. The contribution of the GPIbα receptor to platelet activation induced by the anti‐β~2~GPI antibody–β~2~GPI complex was assessed by observing the influence of 2 anti‐GPIbα antibodies (AK2 and SZ2) directed against distinct epitopes.
Results
This study showed that β~2~GPI could bind to the GPIbα receptor. The anti‐β~2~GPI antibody–β~2~GPI complex was able to activate platelets, and this effect was inhibited by anti‐GPIbα antibody directed against epitope Leu‐36–Gln‐59, but not by anti‐GPIbα antibody directed against residues Tyr‐276–Glu‐282.
Conclusion
Our findings show that inappropriate platelet activation by the anti‐β~2~GPI antibody–β~2~GPI complex via the GPIbα receptor may contribute to the prothrombotic tendency associated with APS.