Interaction of the antiphospholipid syndrome autoantigen beta-2 glycoprotein I with DNA and neutrophil extracellular traps.

Beta-2 glycoprotein I (ß2GPI) is a phospholipid-binding plasma protein and prominent autoantigen in antiphospholipid syndrome (APS). Here, we tested the hypothesis that ß2GPI might bind to not only phospholipids, but also cell-free DNA and neutrophil extracellular traps (NETs). We report that ß2GPI interacts with cell-free DNA from different species, as well as NETs, in a dose-dependent manner, retarding their migration in an agarose-gel electrophoretic mobility shift assay. We confirm the direct binding interaction of ß2GPI with DNA and NETs by ELISA. We also demonstrate that ß2GPI colocalizes with NET strands by immunofluorescence microscopy. Finally, we provide evidence that ß2GPI-DNA complexes can be detected in the plasma of APS patients, where they positively correlate with an established biomarker of NET remnants. Taken together, our findings indicate that ß2GPI interacts with DNA and NETs, and suggest that this interaction may play a role as a perpetuator and/or instigator of autoimmunity in APS.

Release of neutrophil extracellular traps by neutrophils stimulated with antiphospholipid antibodies: a newly identified mechanism of thrombosis in the antiphospholipid syndrome.

OBJECTIVE: Antiphospholipid antibodies (aPL), especially those targeting ß2 -glycoprotein I (ß2 GPI), are well known to activate endothelial cells, monocytes, and platelets, with prothrombotic implications. In contrast, the interaction of aPL with neutrophils has not been extensively studied. Neutrophil extracellular traps (NETs) have recently been recognized as an important activator of the coagulation cascade, as well as an integral component of arterial and venous thrombi. This study was undertaken to determine whether aPL activate neutrophils to release NETs, thereby predisposing to the arterial and venous thrombosis inherent in the antiphospholipid syndrome (APS). METHODS: Neutrophils, sera, and plasma were prepared from patients with primary APS (n = 52) or from healthy volunteers and characterized. No patient had concomitant systemic lupus erythematosus. RESULTS: Sera and plasma from patients with primary APS had elevated levels of both cell-free DNA and NETs, as compared to healthy volunteers. Freshly isolated neutrophils from patients with APS were predisposed to high levels of spontaneous NET release. Further, APS patient sera, as well as IgG purified from APS patients, stimulated NET release from control neutrophils. Human aPL monoclonal antibodies, especially those targeting ß2 GPI, also enhanced NET release. The induction of APS NETs was abrogated with inhibitors of reactive oxygen species formation and Toll-like receptor 4 signaling. Highlighting the potential clinical relevance of these findings, APS NETs promoted thrombin generation. CONCLUSION: Our findings indicate that NET release warrants further investigation as a novel therapeutic target in APS.