Anti–Neutrophil Extracellular Trap Antibodies in Antiphospholipid Antibody–Positive Patients: Results From the Antiphospholipid Syndrome Alliance for Clinical Trials and InternatiOnal Networking Clinical Database and Repository

Abstract

Objective This study aimed to elucidate the presence, antigen specificities, and potential clinical associations of anti–neutrophil extracellular trap (anti‐NET) antibodies in a multinational cohort of antiphospholipid (aPL) antibody–positive patients who did not have lupus. Methods Anti‐NET IgG/IgM levels were measured in serum samples from 389 aPL‐positive patients; 308 patients met the classification criteria for antiphospholipid syndrome. Multivariate logistic regression with best variable model selection was used to determine clinical associations. For a subset of the patients (n = 214), we profiled autoantibodies using an autoantigen microarray platform. Results We found elevated levels of anti‐NET IgG and/or IgM in 45% of the aPL‐positive patients. High anti‐NET antibody levels are associated with more circulating myeloperoxidase (MPO)–DNA complexes, which are a biomarker of NETs. When considering clinical manifestations, positive anti‐NET IgG was associated with lesions affecting the white matter of the brain, even after adjusting for demographic variables and aPL profiles. Anti‐NET IgM tracked with complement consumption after controlling for aPL profiles; furthermore, patient serum samples containing high levels of anti‐NET IgM efficiently deposited complement C3d on NETs. As determined by autoantigen microarray, positive testing for anti‐NET IgG was significantly associated with several autoantibodies, including those recognizing citrullinated histones, heparan sulfate proteoglycan, laminin, MPO–DNA complexes, and nucleosomes. Anti‐NET IgM positivity was associated with autoantibodies targeting single‐stranded DNA, double‐stranded DNA, and proliferating cell nuclear antigen. Conclusion These data reveal high levels of anti‐NET antibodies in 45% of aPL‐positive patients, where they potentially activate the complement cascade. While anti‐NET IgM may especially recognize DNA in NETs, anti‐NET IgG species appear to be more likely to target NET‐associated protein antigens.

An Emerging Role for Anti-DNA Antibodies in Systemic Lupus Erythematosus

“…In another study, IgG and IgM antibodies to NETs measured by ELISA were significantly elevated in patients with primary APS and in SLE without aPL relative to the healthy controls [58]. In a recent larger cohort of primary APS, 45% of the aPL-positive patients had IgG and/or IgM anti-NET antibodies [59]. Importantly, an analysis of the associations of anti-NET antibodies with clinical manifestations revealed that IgG antibodies were associated with lesions affecting the white matter of the brain, while IgM antibodies were tracked with complement consumption.…”

How We Interpret Thrombosis with Thrombocytopenia Syndrome?

“…On the other hand, the presence of aPLs in vivo promotes coagulation and causes thrombosis. As a mechanism for causing thrombosis, antiphospholipid antibodies inhibit the activation of protein C, inhibit β2-glycoprotein-I (β2-GPI) that suppresses phospholipid-dependent coagulation reactions [94,95], inhibit or impair thrombomodulin [96] and heparan sulfate [97] on vascular endothelial cells, suppress prostacyclin production from vascular endothelial cells that inhibits platelet aggregation [98], and increase the production and release of VWF [99,100] and plasminogen activator inhibitor [101]. Clinical findings are often found in deep-vein thrombosis, thrombocytopenia, recurrent miscarriage/stillbirth (infertility), cerebral infarction, migraine, and livedo [102].…”

Antibodies to Neutrophil Extracellular Traps: Novel Markers for the Antiphospholipid Syndrome

“…In this issue of Arthritis & Rheumatology , a provocative serologic analysis by Zuo et al provides a novel perspective on autoantibody expression in patients with antiphospholipid antibodies (aPL) (1). As the data from a large patient cohort indicate, patients whose serum is persistently positive for APL, including those with the antiphospholipid syndrome (APS), produce IgM and IgG antibodies to neutrophil extracellular traps (NETs), a large DNA‐rich structure released by activated neutrophils (2,3).…”

“…As shown in their study, 45% of 389 patients with aPL, 309 of whom had APS, had anti‐NET antibodies. Of interest, the presence of the IgG anti‐NET antibodies was associated with white matter lesions, while IgM anti‐NET antibodies were associated with complement activation (1). In this and other studies in which anti‐NET antibodies have been quantified, the NETs in the supernatants of activated cells are partially digested with micrococcal nuclease to create an antigen preparation suitable for assay in microtiter plates.…”