Objective: Tolerogenic dendritic cells (DCs) are antigen‐presenting cells with an immunosuppressive function. They are a promising immunotherapeutic tool for the attenuation of pathogenic T cell responses in autoimmune arthritis. The aims of this study were to determine the therapeutic action of tolerogenic DCs in a type II collagen–induced arthritis model and to investigate their effects on Th17 cells and other T cell subsets in mice with established arthritis.Methods: Tolerogenic DCs were generated by treating bone marrow–derived DCs with dexamethasone and vitamin D₃ during lipopolysaccharide‐induced maturation. Mice with established arthritis received 3 intravenous injections of tolerogenic DCs, mature DCs, or saline. Arthritis severity was monitored for up to 4 weeks after treatment. Fluorescence‐labeled tolerogenic DCs were used for in vivo trafficking studies. The in vivo effect of tolerogenic DCs on splenic T cell populations was determined by intracellular cytokine staining and flow cytometry.Results: Tolerogenic DCs displayed a semi‐mature phenotype, produced low levels of inflammatory cytokines, and exhibited low T cell stimulatory capacity. Upon intravenous injection into arthritic mice, tolerogenic DCs migrated to the spleen, liver, lung, feet, and draining lymph nodes. Treatment of arthritic mice with type II collagen–pulsed tolerogenic DCs, but not unpulsed tolerogenic DCs or mature DCs, significantly inhibited disease severity and progression. This improvement coincided with a significant decrease in the number of Th17 cells and an increase in the number of interleukin‐10–producing CD4+ T cells, whereas tolerogenic DC treatment had no detectable effect on Th1 cells or interleukin‐17–producing γ/δ T cells.Conclusion: Treatment with type II collagen–pulsed tolerogenic DCs decreases the proportion of Th17 cells in arthritic mice and simultaneously reduces the severity and progression of arthritis.

Autoimmune pathologies are caused by a breakdown in self-tolerance. Tolerogenic dendritic cells (tolDC) are a promising immunotherapeutic tool for restoring self-tolerance in an antigen-specific manner. Studies about tolDC have focused largely on generating stable maturation-resistant DC, but few have fully addressed questions about the antigen-presenting and migratory capacities of these cells, prerequisites for successful immunotherapy. Here, we investigated whether human tolDC, generated with dexamethasone and the active form of vitamin D3, maintained their tolerogenic function upon activation with LPS (LPS-tolDC), while acquiring the ability to present exogenous autoantigen and to migrate in response to the CCR7 ligand CCL19. LPS activation led to important changes in the tolDC phenotype and function. LPS-tolDC, but not tolDC, expressed the chemokine receptor CCR7 and migrated in response to CCL19. Furthermore, LPS-tolDC were superior to tolDC in their ability to present type II collagen, a candidate autoantigen in rheumatoid arthritis. tolDC and LPS-tolDC had low stimulatory capacity for allogeneic, naïve T cells and skewed T cell polarization toward an anti-inflammatory phenotype, although LPS-tolDC induced significantly higher levels of IL-10 production by T cells. Our finding that LPS activation is essential for inducing migratory and antigen-presenting activity in tolDC is important for optimizing their therapeutic potential.

Objective: Rheumatoid arthritis (RA) is considered to be a prototypical autoimmune disease. However, the autoantigens that play an important role in the development of RA remain unclear. The aim of this study was to investigate whether T cells specific for citrullinated epitopes from self proteins are present in patients with RA.Methods: Peripheral blood mononuclear cells (PBMCs) from 28 RA patients and 18 healthy controls were stimulated with citrullinated or noncitrullinated aggrecan peptide Agg^84–103, and proliferative and cytokine responses were assessed using ³H‐thymidine incorporation assay, enzyme‐linked immunosorbent assay, and intracellular cytokine analysis.Results: A proliferative response to the citrullinated aggrecan peptide was detected in >60% of RA patients but not in healthy controls. Furthermore, citrullinated aggrecan peptide–stimulated PBMCs from RA patients produced high levels of the proinflammatory cytokine interleukin‐17 (IL‐17), accompanied by an induction of IL‐17+CD4+ T cells. In contrast, PBMCs from RA patients and healthy controls exhibited no response to stimulation with the noncitrullinated aggrecan peptide.Conclusion: Proinflammatory T cell responses to stimulation with a citrullinated arthritogenic aggrecan peptide were detected in RA patients but not in healthy individuals, suggesting a role for these autoantigen‐specific T cells in the pathogenesis of RA. Our results suggest that the lack of response to the noncitrullinated analog peptide not only implicates the citrulline residue in T cell recognition but also highlights the potential value of citrullinated aggrecan peptide–specific responses as biomarkers of RA. To our knowledge, this is the first study to demonstrate the presence of citrullinated antigen–specific T cells in human RA.

Professional antigen-presenting cells, such as dendritic cells, macrophages and B cells have been implicated in the pathogenesis of rheumatoid arthritis, constituting a possible target for antigen-specific immunotherapy. We addressed the possibility of blocking antigen presentation of the type II collagen (CII)-derived immunodominant arthritogenic epitope CII_259–273 to specific CD4 T cells by inhibition of antigen uptake in HLA-DR1-transgenic mice in vitro and in vivo. Electron microscopy, confocal microscopy, subcellular fractionation and antigen presentation assays were used to establish the mechanisms of uptake, intracellular localization and antigen presentation of CII by dendritic cells and macrophages. We show that CII accumulated in membrane fractions of intermediate density corresponding to late endosomes. Treatment of dendritic cells and macrophages with cytochalasin D or amiloride prevented the intracellular appearance of CII and blocked antigen presentation of CII_259–273 to HLA-DR1-restricted T cell hybridomas. The data suggest that CII was taken up by dendritic cells and macrophages predominantly via macropinocytosis. Administration of amiloride in vivo prevented activation of CII-specific polyclonal T cells in the draining popliteal lymph nodes. This study suggests that selective targeting of CII internalization in professional antigen-presenting cells prevents activation of autoimmune T cells, constituting a novel therapeutic strategy for the immunotherapy of rheumatoid arthritis.