Background/Purpose: Neutrophil activation, including formation of neutrophil extracellular traps (NETs), is essential in host defense. However, NET formation has also been linked to inflammation and autoimmunity, such as systemic lupus erythematosus (SLE). We recently described that SLE-derived immune complexes (ICs) induced NET formation, with NETosis promoting lupus-like disease, including nephritis, in mice. However, whether these mechanisms would occur in human SLE, and the clinical implications of elevated levels of NETs in SLE has not been carefully addressed.

Methods: Levels of NETs (MPO-DNA complexes) were analyzed in plasma in four cross-sectional well-characterized SLE cohorts (n=44-142), and healthy individuals (n=100) using ELISA. Levels of immune complexes and calprotectin were determined by ELISA. Type I interferon (IFN) activity was analyzed using a cell reporter system. Endothelial-derived microparticles (EMPs) were measured by flow cytometry. To determine the capacity of NETs to predict disease flare, patients (n=47) were recruited at time-point of low disease activity and followed over three months, with 14 patients remaining in remission, and 33 patients developing worsening of disease. Statistical analyses were done using Mann-Whitney U test and logistic regression analysis.

Results: SLE patients had elevated levels of NETs in circulation as compared to healthy controls (p< 0.01). Consistent with prior in vitro studies, levels of NETs were associated with heightened type I IFN activity (p< 0.05) and levels of immune complexes (r=0.40, p< 0.0001). In contrast to the neutrophil activation marker calprotectin (p< 0.01), levels of NETs (a marker of neutrophil cell death) were not associated with current disease activity (p=0.20), nor individual SLEDAI items. However, of note, levels of NETs predicted future increase in disease activity. Thus, patients with elevated levels of NETs at time-point of clinical remission, were likely to develop disease flare within a few months (OR=13.8, p=0.002). These results are consistent with NET formation occurring early in the disease development, prior to clinical manifest disease. As such, NET formation may be a promising therapeutic target. Further, elevated levels of NETs identified patients with a severe disease phenotype involving nephritis, independent on presence of anti-dsDNA antibodies (OR=1.25 per 1 U/mL NETs, p=0.03). Finally, we made the intriguing observation that levels of NETs were elevated in patients with arterial thrombosis, including myocardial infarction (OR=9.55, p=0.01), and associated with endothelial damage and EMPs (p< 0.0001).

Conclusion: NET levels are elevated in SLE patients, and associated with immune complex-driven disease involving nephritis and arterial thrombosis. NET levels provide significant clinical value in identifying patients at risk of flare and/or severe disease, which may allow for early preventive treatment interventions, reducing overall disease morbidity and mortality.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/clinical-implications-of-neutrophil-extracellular-traps-in-systemic-lupus-erythematosus/