A2.8 Enhanced Neutrophil Extracellular Trap Formation in Rheumatoid Arthritis Patients is Correlated with High Levels of Rheumatoid Factor (RF)

Background/Objectives Neutrophils are the most abundant cell type identified in joints from patients with rheumatoid arthritis (RA), with a key role in inflammation and cartilage damage. Activated neutrophils form extracellular traps (NETs) with potent pro-inflammatory and immunostimulatory activity. Consequently, we sought to assess the role of NET release (NETosis) in RA pathogenesis and whether RA specific autoantibodies (rheumatoid factor [RF]) are correlated to this phenomenon. Materials/Methods Peripheral blood neutrophils were isolated from active RA patients (n = 6) (Disease activity score, DAS28 > 5.1) and healthy control subjects (n = 7). NET formation from neutrophils, both spontaneous and following incubation with RA serum (n = 7) or synovial fluid (n = 7), was assessed by immunofluoresence microscopy, using co-staining with myeloperoxidase and 4’,6-Diamidino-2-phenylindole dihydrochloride (DAPI). The percentage of NET releasing cells was determined by examining 200 cells per sample in a double blind fashion. Extracellular DNA content was quantified by fluorescence spectrometry (picogreen) and NET fold increase was calculated based on the extracellular DNA content produced by healthy unstimulated neutrophils. Results Freshly isolated neutrophils from the peripheral blood of RA patients underwent spontaneous NETosis at higher rates compared to healthy controls (12 ± 2.1% versus 3.2 ± 0.9%, p Conclusions We found that neutrophils from RA patients have enhanced NET formation, driven by soluble factors found in RA sera and synovial fluid, and this is associated with presence of RF. Further studies will address whether NETs are involved in the initiation of adaptive immune responses in humans and in mouse model of arthritis, and whether suppression of NETosis may ameliorate arthritis in RA mouse models.