Abstract
OBJECTIVES: Interferon (IFN) alpha is a key immunoregulatory cytokine secreted by activated plasmacytoid dendritic cells (PDC) that constitute less than 1% of leucocytes. IFNalpha plays an important role in the pathogenesis of systemic lupus erythematosus (SLE). Nevertheless, the natural IFNalpha inducers in SLE as well as the different IFNalpha secreting cell types are only partially characterised. METHODS: Chromatin was purified from calf thymus. Human peripheral blood mononuclear cells (PBMC), neutrophils and mouse bone marrow neutrophils were purified and cultured with different stimuli. IFNalpha production was estimated by flow cytometry, ELISA and a bioassay, and gene expression by quantitative real time PCR. Neutrophil activation and NETosis were analysed by flow cytometry, ELISA and confocal microscopy. RESULTS: Neutrophils produced a bioactive IFNalpha on stimulation with purified chromatin. IFNalpha secretion was observed with steady state neutrophils purified from 56 independent healthy individuals and autoimmune patients in response to free chromatin and not chromatin containing immune complexes. Chromatin induced IFNalpha secretion occurred independently of Toll-like receptor 9 (TLR9). Neutrophil priming by granulocyte-colony stimulating factor, granulocyte macrophage-colony stimulating factor or IFNalpha was not necessary but PBMC sustained IFNalpha secretion by neutrophils. PDC were 27 times more efficient than neutrophils but blood neutrophils were 100 times more frequent than PDC. Finally, neutrophil activation by chromatin was associated with NETosis and DNA sensor upregulation. CONCLUSIONS: Neutrophils have the capability of producing IFNalpha on selective triggering, and we identified a natural lupus stimulus involved, unveiling a new mechanism involved in SLE. Neutrophils represent another important source of IFNalpha and important targets for future therapies aimed at influencing IFNalpha levels.
