| Abstract |
Rheumatoid arthritis (RA) is a chronic immune inflammatory disease characterized by synovial hyperplasia, joint destruction and extra-articular manifestations. RA presents local inflammation in the joints, which subsequently develops into a systemic disorder as a result of the loss of immune tolerance. The inflammation in the joints is the result of a tight interaction between resident cells, such as fibroblast-like synoviocytes (FLS) and cells of the innate and adaptive immune system. Moreover, one of the major contributors in RA pathogenesis are inflammatory cytokines, mostly TNF-α and IL-6, which orchestrate innate and adaptive immune system activation, playing a key role in the tissue damage.
Plasmacytoid dendritic cells (pDCs) are a unique subset of dendritic cells (DCs), specialized in producing large amounts of type I IFNs following their recognition of viruses or self nucleic acids through Toll-like receptor 7 (TLR7) and TLR9. However, pDCs can also secrete other pro-inflammatory cytokines and chemokines, and function as antigen presenting cells (APCs). Interestingly, pDCs can participate in the priming of both immunogenic and tolerogenic immune responses. The role of pDCs in inflammatory arthritis has not been extensively investigated. Data from animal models have shown a rather immunoregulatory role. In addition, a previous study from our laboratory has demonstrated that RA patients in remission have higher number of pDCs in peripheral blood compared to active RA patients, which have a tolerogenic phenotype and induce IL-10 secreting Treg cells in vitro. In this study we investigated the molecular mechanism through which human pDCs contribute to restoration of tolerance in RA.
To address this question, we performed transcriptomic analysis (microarray) of pDCs of RA patients responding to anti-TNF therapy and pDCs derived from healthy donors. The results from microarrays revealed more than 6000 genes differentially regulated between RA and healthy pDCs. Interestingly, IL-6R was significantly upregulated on RA pDCs. Given that IL-6 has a major role in the pathogenesis of RA, we focused on its receptors and IL-6 signaling pathway. We firstly validated the result in a new cohort of RA patients, and we found increased IL-6R as well as pSTAT1 expression on pDCs of RA patients on remission as compared with healthy pDCs. These findings indicate that IL-6/IL-6R pathway is up-regulated and activated. Therefore, and based on the aforementioned data supporting a regulatory role of pDCs in the context of arthritis, we hypothesized that the increased IL-6 signaling pathway may enhance the tolerogenic phenotype of pDCs. In order to study our hypothesis, we performed in vitro experiments on isolated healthy pDCs treated with CpG-A, as a general stimulator of pDCs, and recombinant IL-6 (rIL6). Interestingly, we found that IL-6 signaling did not further induce IFN-a expression nor activation status of pDCs, but significantly decreased TFN-a production by pDCs. Considering that ADAM17 mediates the generation of soluble TNF, we assessed the effect of IL6 treatment on the function of ADAM17 in pDCs. We concluded with a trend towards decreased mature ADAM17 expression on the surface of pDCs upon IL-6 signaling. Moreover, IL-6R is another substrate cleaved by ADAM17. We thus assessed the effect of IL-6 signaling on soluble IL-6R secretion by pDCs. We found that IL-6 induces decreased production of soluble IL-6R in culture supernatants of pDCs, further supporting an inhibitory effect of IL-6 signaling in ADAM17 function. Collectively, we had strong evidence that IL-6 signaling on pDCs mediates impaired ADAM17 activity.
In summary, our findings delineated a novel role of pDCs in the induction of tolerance in RA and leaded us to the hypothesis that increased IL-6 signaling on pDCs inhibits ADAM17 activity, therefore limiting TNFa secretion and this process may enhance the tolerogenic phenotype of pDCs.
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Delineation of the tolerogenic function of plasmacytoid dendritic cells in Rheumatoid arthritis
