Background New bone formation in spondyloarthritis involves myofibroblasts . IL-26 is part of the IL-10 cytokine family and was recently reported to increase Th17 differentiation . It was first discovered in herpesvirus saimiri–transformed T cells and thus suspected to be a T-cell cytokine . However, more recent studies of RA patients show IL-26 expression in synoviolin-postive cells harvested from the synovia . The knowledge about IL-26 is compromised by lack of IL-26 in mice and no studies of IL-26 in spondyloarthritis (SpA) and its regulation have been conducted.
Objectives In the present study we aim to characterize IL-26 producing cells in spondyloarthritis (SpA).
Methods IL-26 was measured by ELISA in plasma and synovial fluid (SF) of 14 SpA patients attending the clinic with a swollen joint and in plasma samples from 7 age and gender matched healthy controls. Synovial-fluid-mononuclear-cells (SFMC) and fibroblast-like-synoviocytes (FLS) were cultured and stimulated for 48 hours with IL-1β, TNFα, TGF-β, IL-17A and LPS. The supernatants were harvested and analyzed for IL-26 by ELISA. Tissue samples of inflamed synovial membrane were stained with a fluorochrome conjugated rabbit IL-26 antibody for immunofluorescence. Imaging of the sections was performed by confocal microscopy. For flow cytometry analysis, FLS were surface stained for CD90 and a live-dead marker, and subsequently permeabilized and stained intracellularly for α-Smooth-Muscle-Actin (αSMA) and IL-26. Samples were run on the flow cytometer within 24 hours of staining. Data are expressed as mean ± SD.
Results IL-26 levels were significantly higher in synovial fluid compared with plasma; this applied to plasma from the healthy controls (HC) as well as SpA patients. (HC plasma: 5.8 ± 3.3 pg/ml, SpA plasma: 6.6 ± 1.8 pg/ml, SpA SF: 41.6 ± 6.8 pg/ml - P-values HC vs. SF: 0.0020, P-value SP vs. SF: <0.0001.) Staining of synovial membrane revealed IL-26 presence in synovial cells with fibroblast-like morphology. ELISA analysis of the supernatants from culturing and stimulation of FLS and SFMC showed a high level of IL-26 in both cell populations as well as drastic increase in IL-26 production after stimulation with IL-1, TNF-α and LPS. The vast majority of the IL-26 producing FLS were αSMA+ and CD90+ myofibroblasts by flow cytometry.
Conclusions IL-26 levels in synovial fluid are significantly higher than in plasma, suggesting a local production of IL-26 in the inflamed joint. We found IL-26 positive cells in the synovial membrane with fibroblast morphology and FLS from the inflamed joints of SpA patients produce high amounts of IL-26. By flow cytometry we established that these IL-26 producing fibroblasts seem to be myofibroblasts primarily.
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Disclosure of Interest None declared