Effects of inflammatory cytokine IL-27 on the activation of fibroblast-like synoviocytes in rheumatoid arthritis

Abstract

IntroductionInterleukin (IL)-27 is a novel member of the IL-6/IL-12 family cytokines that are produced early by antigen-presenting cells in T helper (Th)1-mediated inflammation. Elevated expression of IL-27 has been detected in the synovial membranes and fluid of rheumatoid arthritis (RA).MethodsWe investigated the in vitro effects of IL-27, alone or in combination with inflammatory cytokine tumor necrosis factor (TNF)-α or IL-1 β on the pro-inflammatory activation of human primary fibroblast-like synoviocytes (FLS) from RA patients and normal control subjects, and the underlying intracellular signaling molecules were determined by intracellular staining using flow cytometry.ResultsSignificantly higher plasma concentration of IL-27 was found in RA patients (n = 112) than control subjects (n = 46). Both control and RA-FLS constitutively express functional IL-27 receptor heterodimer, gp130 and WSX-1, with more potent IL-27-mediated activation of signal transducers and activators of transcription (STAT)1 in RA-FLS. IL-27 was found to induce significantly higher cell surface expression of intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 and release of inflammatory chemokine IL-6, CCL2, CXCL9, CXCL10 and matrix metalloproteinase-1 of RA-FLS than that of control FLS (all P < 0.05). Moreover, an additive or synergistic effect was observed in the combined treatment of IL-27 and TNF-α or IL-1 β on the surface expression of ICAM-1 and VCAM-1 and the release of CXCL9 and CXCL10 of RA-FLS. Further investigations showed that the expression of ICAM-1, VCAM-1 and chemokines stimulated by IL-27 was differentially regulated by intracellular activation of phosphatidylinositol 3-OH kinase-AKT, c-Jun amino-terminal kinase and Janus kinase pathways.ConclusionsOur results therefore provide a new insight into the IL-27-activated immunopathological mechanisms mediated by distinct intracellular signal transductions in joint inflammation of RA.

Activation of human fibroblast-like synoviocytes by uric acid crystals in rheumatoid arthritis

“…27 We have recently reported that the inflammatory cytokine IL-27 can activate FLS in RA by activating distinct intracellular signaling pathways. 11 The present study further revealed that uric acid plays a crucial role in provoking inflammation by activating FLS through intracellular signal transduction mechanisms in RA.…”

“…11 Intracellular staining of activated (phosphorylated) signaling molecules The intracellular expression of phosphorylated signaling molecules was determined quantitatively using previously established intracellular staining methods using flow cytometry. 11,22 This quantitative flow cytometric method for the analysis of the activation of intracellular signaling molecules by intracellular staining of phosphorylated signaling molecules is less tedious than western blot, and the flow cytometric method requires fewer cells and a reduced assay time. Briefly, cells were fixed with pre-warmed BD Cytofix Buffer (4% paraformaldehyde) for 10 min at 37 uC following stimulation by monosodium urate crystals.…”

“…11,22,24 Following previous publications and toxicity threshold values from the methyl thiazolyl tetrazolium assay, we therefore used the following optimal concentrations of Janus kinase inhibitor AG490 (5 mM), IkB-a phosphorylation inhibitor BAY11-7082 (0.8 mM), phosphatidylinositol 3-OH kinase inhibitor LY294002 (5 mM), ERK inhibitor U0126 (5 mM), p38 MAPK inhibitor SB203580 (10 mM) and JNK inhibitor SP600125 (5 mM) to induce significant inhibitory effects on these signaling pathways without cell toxicity. As shown in Figure 6, the ERK inhibitor U0126 and the JNK inhibitor SP600125 were able to significantly suppress monosodium urate crystal-induced IL-6, CXCL8 and MMP-1 release from both normal and RA-FLS (P,0.05).…”

“…9 FLS can produce a broad range of pro-inflammatory molecules such as interleukin (IL)-1b, IL-6 and tumor-necrosis factor (TNF)-a, as well as the secretion of MMPs through the activation of multiple intracellular signal transduction pathways including extracellular signal-regulated protein kinase (ERK), c-Jun Nterminal kinase (JNK) and p38 mitogen activated protein kinase (MAPK). 10,11 These pro-inflammatory molecules lead to further FLS activation and the recruitment, accumulation and survival of a number of immune effector cells including leukocytes, endothelial cells, neutrophils and osteoclasts in the synovium. 9 IL-6, a key proinflammatory cytokine, was originally described as a T cell-derived B-cell differentiation factor that is essential for antibody production by activated B cells and may stimulate the production of autoantibodies such as rheumatoid factor in RA.…”

Interleukin‐27 in rheumatic diseases: Friend or foe? Comment on the article by Pickens et al

“…Consistent with these findings, anti-IL-27 antibody treatment or IL-27 receptor knockout has been found to suppress ongoing severe inflammation and proinflammatory cytokine production in AIA (5). Moreover, recently published data suggested that IL-27 could stimulate other nonimmune cell targets, such as fibroblast-like synoviocytes (FLS), in the joints and induce the expression of adhesion molecules on the cell surface of FLS and the release of chemokines and MMP-1 in RA FLS (6,7).…”