LAUSR

Background Human plasmacytoid dendritic cells (pDCs) play a vital role in modulating immune responses. pDCs can produce massive amounts of type I IFNs in response to nucleic acids via toll-like receptors (TLRs) and they are known to possess weak antigen-presenting properties inducing CD4+ T cell activation. Previous data showed a cross-regulation between TNF-α and IFN-α but the effect of TNF-α on pDCs remains unclear. Objectives To investigate how TNF-α regulates the immune function of human pDCs. Methods Fresh PBMCs were treated with TNF-α, TLR7 and TLR9 synthetic agonists. pDCs were immunophenotyped using flow cytometry. RNA from sorted pDCs was extracted and sequenced using Smart-seq2 for sensitive full-length transcriptome profiling. For pDC/T cell co-culture, fresh or TNF-α-treated pDCs were cultured with na&x00EE;ve CD4+ T cells for 5 days. The production of cytokines was measured by intracellular staining and ELISA. Results Upon stimulation with TLR7 and TLR9 agonists, there were three main pDC populations: non-producers, TNF-α-producers, TNF-α/IFN-α-producers. Exogenous TNF-α significantly reduced the production of both IFN-α and TNF-α in TLR9-stimulated pDCs but only IFN-α in TLR7-stimulated pDCs. Neutralisation of autologous TNF-α with anti-TNF antibody partially sustained IFN-α secretion by TLR9-stimulated pDCs after 24 hours. Exogenous TNF-α significantly promoted pDC maturation by upregulation of costimulatory molecules and chemokine receptors such as CD80, CD86, HLA-DR, and CCR7. RNAseq data analysis suggested that TNF-α inhibits IFN-α/TNF-α production by interfering with the NFκB pathway, promotes antigen processing and presentation pathways as well as T cell activation and differentiation. Indeed, TNF-α-treated pDCs induced in vitro higher CD4+ T cell proliferation and favoured Th1/Th17 polarisation. Conclusions Although pDCs possess weak antigen-presenting properties, TNF-α can enhance pDC maturation by switching their main role as IFN-α-producing cells to a more conventional DC phenotype. The functional status of pDCs might be strongly influenced by overall inflammatory environment and TNF-α might regulate IFN-α-mediated aspects of a range of autoimmune and inflammatory diseases. Disclosure of Interest None declared