Destructive characteristics of Th17 cells. A joint venture

Contains fulltext : 112915.pdf (publisher's version ) (Open Access)Radboud Universiteit Nijmegen, 28 augustus 2013Promotores : Berg, W.B. van den, Boots, A.M.H. Co-promotores : Koenders, M.I., Joosten, L.A.B., Loo, A.A.J. van d

Method of the dosed application of a liquid onto a surface

The invention relates to a method of the dosed application of a liquid onto to selected portion of the surface of a substrate (A) by means of spraying under the influence of an electric current. According to the invention the liquid is fed at a flow rate between 0.01 pl/s and 1 ml/s to a distal tip ( 3 ) of a capillary ( 1 ) having an inside diameter of less than 150 mum, wherein the distance between the distal tip and the surface (A) is less than 2 mm. Surprisingly it has been shown that it is possible in this manner to apply liquid to a restricted surface of a defined size.Applied Science

Increased expression of interleukin-22 by synovial Th17 cells during late stages of murine experimental arthritis is controlled by interleukin-1 and enhances bone degradation

Item does not contain fulltextOBJECTIVE: Interleukin-22 (IL-22) is a mediator in antimicrobial responses and inflammatory autoimmune diseases. Although IL-22 and its receptor, IL-22R, have been identified in the synovium of rheumatoid arthritis patients, the source of IL-22 and its contribution to disease pathogenicity remain to be established. This study was undertaken to investigate the regulation of IL-22 by Th17 cells in vitro and to evaluate the potential for IL-22 depletion in an experimental arthritis model using mice deficient in the IL-1 receptor antagonist (IL-1Ra-/-). METHODS: Naive murine T cells were cultured under conditions leading to polarization of the cells into subsets of Th1, Th2, induced Treg, and Th17. Cytokines were measured in the culture supernatants, and the cells were analyzed by fluorescence-activated cell sorting. Tissue samples from the inflamed ankle synovium of IL-1Ra-/- mice were isolated, and messenger RNA levels of marker genes were quantified. IL-1Ra-/- mice were treated with neutralizing anti-IL-22 antibodies. Synovial cells were isolated from the inflamed tissue and sorted into fractions for analysis of cytokine production. RESULTS: In vitro tests showed that Th17 cells produced high levels of IL-22 after stimulation with IL-1 or IL-23. Interestingly, a synergistic increase in the production of IL-22 was observed after combining IL-1 and IL-23. In vivo, IL-1Ra-/- mice displayed a progressive erosive arthritis, characterized by up-regulation of IL-17 in mildly and severely inflamed tissue, whereas the levels of IL-22 and IL-22R were increased only in severely inflamed synovia. Anti-IL-22 treatment of IL-1Ra-/- mice significantly reduced the inflammation and bone erosion. Analysis of isolated single cells from the inflamed synovia revealed that IL-22 was mainly produced by IL-17-expressing T cells. CONCLUSION: These findings suggest that IL-22 plays an important role in IL-1-driven chronic joint destruction

Anti IL-17A therapy inhibits bone loss in TNF-α-mediated murine arthritis by modulation of the T-cell balance

Tumour necrosis factor alpha (TNF-α) is a major inducer for inflammation and bone loss. Here, we investigated whether interleukin (IL)-17 plays a role in TNF-α-mediated inflammation and bone resorption. Human TNF-α transgenic (hTNFtg) mice were treated with a neutralizing anti-IL-17A antibody and assessed for inflammation, cartilage and bone damage. T-cell transcription factors and lymphokine patterns were measured in the LNs. IL-17A inhibition in the absence of IL-1 was also evaluated by treating hTNFtg/IL-1−/− mice with an IL-17A neutralizing antibody. IL-17A neutralization had only minor effects on TNF-α-induced inflammation but effectively reduced local and systemic bone loss by blocking osteoclast differentiation in vivo. Effects were based on a shift to bone-protective T-cell responses such as enhanced Th2 differentiation, IL-4 and IL-12 expression and Treg cell numbers. Whereas inflammation in hTNFtg/IL-1−/− mice was highly sensitive to IL-17A blockade, no shift in the T-cell lineages and no additional benefit on bone mass were observed in response to IL-17A neutralization. We thus conclude that IL-17A is a key mediator of TNF-α-induced bone loss by closely interacting with IL-1 in blocking bone protective T-cell responses

IL-21R-deficiency increases the initial TLR2 response but protects against joint pathology by reducing Th1/17 cells during SCW-arthritis

Contains fulltext : 138199.pdf (publisher's version ) (Closed access)Objective: The cytokine IL-21 can have both proinflammatory and immunosuppressive effects. Purpose of this study was to investigate the potential dual role of IL-21 in experimental arthritis in relation to Th17-cells. Method: In IL-21R-deficient mice and wild-types, antigen-induced arthritis (AIA) and chronic Streptococcal Cell Wall (SCW) arthritis were induced. Knee joints, synovial tissue and serum were analyzed for arthritis pathology and inflammatory markers. Results: Our studies show that during AIA and chronic SCW-arthritis, IL-21R-deficiency protects against severe inflammation and joint destruction. This is accompanied by suppressed serum IgG1 levels and antigen-specific T cells responses. IL-17 was reduced during AIA, and synovial lymphocytes isolated during SCW-arthritis for flowcytometry demonstrated that mainly IL-17+ IFNgamma+ T cells were reduced in IL-21R-deficient mice. However, during the acute phases of SCW arthritis significantly higher joint swelling scores were observed, in line with enhanced TNF and IL-6 expression. Interestingly, IL-21R-/- mice were significantly less capable in upregulating SOCS 1/3 mRNA. IL-21 stimulation also affected the TLR2/NOD2 response to SCW fragments in vitro, indicating that impaired SOCS regulation in the absence of IL-21 signaling might contribute to the increased local activation during SCW arthritis. Conclusion: In contrast to the proinflammatory role of IL-21 in adaptive immunity, driving IL-17/IFN double-positive cells and joint pathology during chronic experimental arthritis, IL-21 also has an important immunosuppressive role, presumably by inhibiting TLR signaling via SOCS1/3. If this dual role of IL-21 in various immune processes is present in human disease, it could make IL-21 a difficult therapeutic target in RA. (c) 2013 American College of Rheumatology

The anti-CD20 antibody rituximab reduces the Th17 cell response

Contains fulltext : 98411.pdf (publisher's version ) (Closed access)OBJECTIVE: Rituximab has been shown to be successful in the treatment of rheumatoid arthritis (RA), and this unexpected finding indicates that B cells have an important role in this disease. The present study was undertaken to investigate the mechanism of action of rituximab in RA. METHODS: Twelve patients with active RA were treated with rituximab. Disease activity was evaluated using the 28-joint Disease Activity Score. Synovial biopsy samples obtained at baseline and 12 weeks after treatment initiation were analyzed by microarray, quantitative polymerase chain reaction, and immunohistochemistry. Peripheral blood mononuclear cells (PBMCs) from healthy volunteers and from 4 patients with X-linked agammaglobulinemia were stimulated with the Th17-inducing stimulus Candida albicans, and the response in the presence and absence of rituximab was examined. RESULTS: In RA patients, rituximab reduced expression of retinoic acid-related orphan receptor gammat and interleukin-22 (IL-22) and numbers of Th17-positive cells in synovial tissue, and this correlated with better clinical outcome. Rituximab did not affect tumor necrosis factor alpha (TNFalpha), Th1 cell, or Treg cell responses. Rituximab strongly reduced in vitro IL-17 and IL-22 production induced by C albicans. This effect was not observed in PBMCs from patients with X-linked agammaglobulinemia. CONCLUSION: Rituximab reduced the local Th17 response in RA patients, whereas it did not influence Th1 cell, Treg cell, or TNFalpha responses. The decreased Th17 response was associated with reduced inflammation and better clinical outcome. Moreover, inhibition of the Th17 response by rituximab was lost in the absence of B cells, providing evidence that the effects of rituximab are due to B cell depletion. These data demonstrate an unexpected role of B cells in the development of Th17 responses, which could possibly lead to B cell-based strategies for the treatment of Th17-related autoimmune diseases

Periodontal pathogens directly promote autoimmune experimental arthritis by inducing a TLR2- and IL-1-driven Th17 response

Contains fulltext : 136500.pdf (publisher's version ) (Closed access)Increasing epidemiologic evidence supports a link between periodontitis and rheumatoid arthritis. The actual involvement of periodontitis in the pathogenesis of rheumatoid arthritis and the underlying mechanisms remain, however, poorly understood. We investigated the influence of concomitant periodontitis on clinical and histopathologic characteristics of T cell-mediated experimental arthritis and evaluated modulation of type II collagen (CII)-reactive Th cell phenotype as a potential mechanism. Repeated oral inoculations of periodontal pathogens Porphyromonas gingivalis and Prevotella nigrescens induced periodontitis in mice, as evidenced by alveolar bone resorption. Interestingly, concurrent periodontitis induced by both bacteria significantly aggravated the severity of collagen-induced arthritis. Exacerbation of arthritis was characterized by increased arthritic bone erosion, whereas cartilage damage remained unaffected. Both P. gingivalis and P. nigrescens skewed the CII-specific T cell response in lymph nodes draining arthritic joints toward the Th17 phenotype without affecting Th1. Importantly, the levels of IL-17 induced by periodontal pathogens in CII-specific T cells directly correlated with the intensity of arthritic bone erosion, suggesting relevance in pathology. Furthermore, IL-17 production was significantly correlated with periodontal disease-induced IL-6 in lymph node cell cultures. The effects of the two bacteria diverged in that P. nigrescens, in contrast to P. gingivalis, suppressed the joint-protective type 2 cytokines, including IL-4. Further in vitro studies showed that the Th17 induction strongly depended on TLR2 expression on APCs and was highly promoted by IL-1. Our data provide evidence of the involvement of periodontitis in the pathogenesis of T cell-driven arthritis through induction of Ag-specific Th17 response

Proteome-wide analysis and CXCL4 as a biomarker in systemic sclerosis

Contains fulltext : 136617.pdf (publisher's version ) (Open Access)BACKGROUND: Plasmacytoid dendritic cells have been implicated in the pathogenesis of systemic sclerosis through mechanisms beyond the previously suggested production of type I interferon. METHODS: We isolated plasmacytoid dendritic cells from healthy persons and from patients with systemic sclerosis who had distinct clinical phenotypes. We then performed proteome-wide analysis and validated these observations in five large cohorts of patients with systemic sclerosis. Next, we compared the results with those in patients with systemic lupus erythematosus, ankylosing spondylitis, and hepatic fibrosis. We correlated plasma levels of CXCL4 protein with features of systemic sclerosis and studied the direct effects of CXCL4 in vitro and in vivo. RESULTS: Proteome-wide analysis and validation showed that CXCL4 is the predominant protein secreted by plasmacytoid dendritic cells in systemic sclerosis, both in circulation and in skin. The mean (+/-SD) level of CXCL4 in patients with systemic sclerosis was 25,624+/-2652 pg per milliliter, which was significantly higher than the level in controls (92.5+/-77.9 pg per milliliter) and than the level in patients with systemic lupus erythematosus (1346+/-1011 pg per milliliter), ankylosing spondylitis (1368+/-1162 pg per milliliter), or liver fibrosis (1668+/-1263 pg per milliliter). CXCL4 levels correlated with skin and lung fibrosis and with pulmonary arterial hypertension. Among chemokines, only CXCL4 predicted the risk and progression of systemic sclerosis. In vitro, CXCL4 down-regulated expression of transcription factor FLI1, induced markers of endothelial-cell activation, and potentiated responses of toll-like receptors. In vivo, CXCL4 induced the influx of inflammatory cells and skin transcriptome changes, as in systemic sclerosis. CONCLUSIONS: Levels of CXCL4 were elevated in patients with systemic sclerosis and correlated with the presence and progression of complications, such as lung fibrosis and pulmonary arterial hypertension. (Funded by the Dutch Arthritis Association and others.)