Cell-specific epigenetic landscapes drive differential IL-6 gene expression in human neutrophils and monocytes

IL-6 is a pleiotropic cytokine with a broad range of pro- and anti-inflammatory functions, currently representing a target to modulate in various inflammatory diseases. While monocytes are major source of IL-6, whether human neutrophils produce this cytokine remains still controversial. Human neutrophils and CD14+-monocytes, isolated from buffy coat by immunomagnetic beads at a purity of 99.7 ± 0.2 % and 98 ± 1 % respectively, were stimulated with TLR4 and TLR8 agonists, as well as with other ligands for up to 20 h. IL-6 mRNA expression and production were then measured by RT-qPCR and ELISA, respectively, while analysis of the chromatin status at the IL-6 genomic locus was investigated by chromatin immunoprecipitation assays for histone modifications or transcription factor binding. Our data prove that highly purified neutrophils display the capacity to produce biologically active quantities of IL-6, but only after a prolonged incubation with R848 or, less efficiently, LPS. Data also uncover that IL-6-induction by R848 requires PU.1 recruitment, H3 methylation (e.g., H3K4me1 and H3K4me3) and H3 and H4 acetylation (e.g., H3K27Ac and H4Ac) at the IL-6 locus. Analysis of the distribution of histone modifications within 64 kilobases upstream of the IL-6 transcription start site also revealed that neutrophils and monocytes utilize both common and cell-specific enhancer sites. Data clarify that, at least under TLR-stimulation, neutrophils may generate IL-6 by activating a cascade of events controlled at the epigenetic level.