By using a state-of-the-art technique, we show for the first time that resolution pathways are present in OA patients. A better understanding of these pathways could guide us to more effective therapeutic approaches to inhibit inflammation and further structural damage in OA and RA.

, to target cells. Neutrophil EVs are remarkable in their antiinflammatory properties and were found to be abundant within the joints of RA patients. In addition, depletion of neutrophils within the joints of mice undergoing experimental arthritis, resulted in a reduced proteoglycan synthesis. We therefore investigated the role of Neutrophil EVs in RA and their effect on cartilage breakdown. Methods: Neutrophils were isolated from healthy volunteers and RA patient blood. EVs were isolated from these cells and tested in the K/ BxN serum transfer model of inflammatory arthritis in mice. EVs were administered intra-articularly, and joints were assessed for inflammatory outcomes and cartilage breakdown. Results: EVs were able to successfully enter the cartilage and deliver their cargo to the chondrocytes within. In murine inflammatory arthritis, neutrophil EVs displayed cartilage protective effects, resulting in a reduced loss of sulphated glycosaminoglycans in the treated joints compared to control paired joints. In vitro, neutrophil EVs were able to protect from IL-1 induced cartilage breakdown in 3D chondrocyte cultures and restored basal expression of cartilage specific genes, Sox9 and Collagen2a1. This protective effect occurred in part through the EV mediator Annexin A1 and its receptor Fpr2/3 on the chondrocyte. In addition, neutrophil EVs modulated the macrophage profile, polarizing the cells towards a more anti-inflammatory phenotype, characterized by a lower expression of MHCII and CD86 and higher levels of CD206 compared to contralateral controls. This was true for both healthy and RA derived vesicles. In vitro, co-culture of fibroblast like synoviocytes with classically activated macrophages resulted in upregulation of inflammatory markers in the fibroblasts, whereas pre-treatment of the macrophages with neutrophil EVs attenuated this effect. Conclusions: Neutrophil EVs exert powerful protective bioactions in inflammatory arthritis, displaying not only a modulation of the ongoing joint inflammation but also affording protection from cartilage breakdown. We propose these EVs provide a unique opportunity to target both the inflammatory component of RA and the resulting osteoarthritis with a single therapy.

Background and objectives The infrapatellar fat pad (IFP) is an adipose tissue organ present in the knee next to the synovium and cartilage, thereby constituting a potential player in the pathological processes in the osteoarthritic joint. Obesity-associated changes occur in IFP and this supports the hypothesis that IFP could mediate the association between obesity and the development and progression of osteoarthritis (OA). Interestingly, these changes were observed in the stromal vascular fraction (SVF) rather than adipocytes. As this fraction contain many different types of immune cells, we characterised the SVF of the IFP in OA patients phenotypically and functionally. Materials and methods IFP samples were obtained from knee OA patients (N = 43) undergoing joint replacement surgery (58.1% women; mean (SD) age 66.4 years (10.9); mean (SD) BMI 29.2 kg/m2 (5.7)). The SVF was isolated and cells were characterised based on surface markers expression and cytokine production using flow cytometry. Results Characterisation of the SVF of IFP showed the presence of various immune cells in this tissue, whereby macrophages and T cells were most abundant. Interestingly, flow cytometry analyses of ex vivo cytokine production by different cells revealed a subpopulation of CD4+ T cells that were able to produce IL-6 without further stimulation. These IL-6 producing CD4+ T cells expressed CD69, indicating recent activation. Upon polyclonal stimulation, CD4+ T cells were able to secrete IFNγ, TNFα, and IL-4. However, IL-6 producing CD4+ T cells did not secrete these cytokines. Furthermore, chemokine receptor expression revealed that these IL-6 producing T cells could not be categorised as conventional T helper 1 (Th1), Th2, Th17 or Tfh cells. These data indicate that IL-6-secreting T cells are a distinct population of T cells. Finally, we have also studied whether these IL-6 producing T cells are also present in other tissues. Indeed, we have found these cells also in sc adipose tissues and synovium of OA patients, but only at low frequencies in blood. Conclusion In conclusion, we have found a novel population of CD4+ T cells which secrete IL-6 directly ex vivo and are in an activated state, indicating that these CD4+ T cells might recognise adipose tissue antigens and could be involved in the inflammatory processes present in human adipose tissue. Moreover, they are a source of IL-6 in the OA joint, thereby potentially contributing to joint inflammation.

Ann Rheum Dis 2013;72(Suppl 1):A1-A88 A25 EWRR abstracts progression were determined using a multivariate normal regression model (EAC cohort) or by generalised estimated equations (GARP cohort). Adjustments were made for age, gender, treatment strategy and Body Mass Index (BMI). Results In RA patients totAPN associated positively with radiographic progression (Sharp van der Heijde scores) (association estimate 3.65, p = 0.002), whereas in patients with hand OA, totAPN associated negatively with radiographic progression (joint space narrowing (JSN)) (Odds 0.24/Odds 0.21, p = 0.002/p = 0.002 two highest tertiles compared to the lowest tertile). HmwAPN on the other hand, did not associate significantly with radiographic progression in patients with hand OA or RA, although in patients with RA we did observe a trend towards a positive association (association estimate 1.53 p = 0.07) upon correcting for age, gender and treatment strategy. This trend was lost after further adjustment for BMI. Similar results were obtained when joint space narrowing (JSN) was used as outcome measurement. Conclusions Our data further substantiate the connexion between APN-levels and radiographic progression in rheumatic disease and indicate that the differential effects associated between totAPN and radiographic progression in either in RA and hand OA is not mediated by (a selective effect of) hmwAPN. EFFECTS OF CHOLIC ACID AND ITS DERIVATIVES IN EXPERIMENTAL ARTHRITIS

Purpose: Mesenchymal Stem Cells (MSCs) are considered a promising cell type for the repair of damaged cartilage due to their chondrogenic differentiation potential. However, during cartilage repair in vivo, chondrogenically differentiating MSCs will be exposed to an inflammatory environment caused by osteoarthritis (OA) or as a response to trauma. The release of pro-inflammatory mediators by inflamed synovium may accelerate cartilage matrix degradation and impede cartilage repair. Osteoarthritic synovial fluid and medium conditioned by osteoarthritic synovium explants have been reported to inhibit the chondrogenic differentiation of MSCs. However, the mechanism responsible for this effect remains unclear. Synovium consists of synovial fibroblasts and macrophages. The aim of this study was to investigate how synovial macrophages and their specific polarisation state contribute to the inhibitory effect of OA synovium on MSC chondrogenesis. Methods: Human OA synovial tissue (n¼6) was cut in to pieces between 1-3 mm 2 , and 200 mg of tissue was cultured in 1 ml of serum-free medium. Conditioned medium (CM) was harvested following 3 days of culture. Additionally, synovial tissue was digested and fibroblast or macrophage-enriched fractions were isolated based on the rapid adhesion of macrophages to tissue culture plastic. Each fraction was cultured for 24 hr prior to harvesting CM. Immunohistochemical staining for CD11c as a marker of pro-inflammatory (M1) macrophages, and CD206 as a marker of anti-inflammatory (M2) macrophages, was performed on OA synovium sections (n¼6). Human peripheral blood derived monocytes (n¼3) were stimulated with IFN