ABSTRACT
OBJECTIVES: TNF-like weak inducer of apoptosis (TWEAK), a member of the TNF superfamily, has been shown to increase cytokine production by rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS). In this study, we determined the effect of interaction between TWEAK and its receptor fibroblast growth factor-inducible-14 (Fn14) on cytokine expression in RAFLS. METHODS: RAFLS were obtained from surgical synovial specimens and used at passage 5-10. Cytokine protein and mRNA expression were measured with ELISA and real time-PCR, respectively. Apoptotic cells were detected by TUNEL assay. RelB activation was detected by Western blot analysis. RESULTS: TWEAK inhibited IL-6 production from total synovial cells from RA. TWEAK weakly induced FLS IL-6 and IL-8, but in contrast TWEAK dose-dependently inhibited IL-6 and IL-8 production by TNFα-activated FLS. TWEAK did not induce apoptosis in FLS but inhibited proliferation of TNFα-activated FLS. TWEAK induced RelB activation and suppressed IL-6 mRNA expression in TNFα-activated FLS and both of these phenomenon were abolished by inhibition of new protein synthesis with cycloheximide. CONCLUSIONS: TWEAK has a previously unsuspected inhibitory effect on cytokine production by TNFα-activated RAFLS. This observation suggests that the effects of TWEAK on cytokine expression varies with the pro-inflammatory context, and that in TNFα-activated states such as RA TWEAK may have a net inhibitory effect.
Subject(s)
Arthritis, Rheumatoid/metabolism , Interleukin-6/antagonists & inhibitors , Interleukin-6/metabolism , Receptors, Tumor Necrosis Factor/metabolism , Synovial Membrane/metabolism , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Tumor Necrosis Factors/metabolism , Apoptosis/genetics , Apoptosis/physiology , Arthritis, Rheumatoid/genetics , Arthritis, Rheumatoid/pathology , Cytokine TWEAK , Fibroblasts/metabolism , Fibroblasts/pathology , Humans , Interleukin-6/genetics , Interleukin-8/metabolism , RNA, Messenger/genetics , Recombinant Proteins/metabolism , Synovial Membrane/pathology , TWEAK Receptor , Transcription Factor RelB/metabolism , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/metabolismABSTRACT
CD16+ monocytes, identified as a minor population of monocytes in human peripheral blood, have been implicated in several inflammatory diseases, including rheumatoid arthritis (RA). Fractalkine (FKN, CX3CL1), a member of the CX3 C subfamily, is induced by pro-inflammatory cytokines, while a receptor for FKN, CX3CR1, is capable of mediating both leukocyte migration and firm adhesion. Here, we investigated the role of FKN and CX3CR1 in activation of CD16+ monocytes and their recruitment into synovial tissues in RA patients. High levels of soluble FKN were detected in the synovial fluid and sera of RA patients. Circulating CD16+ monocytes showed a higher level of CX3CR1 expression than CD16- monocytes in both RA patients and healthy subjects. High level expression of CX3CR1 was also seen in CD16+ monocytes localized to the lining layer in RA synovial tissue. In the in vitro culture experiments, IL-10 induced CX3CR1 expression on the surface of monocytes, and TNFalpha induced membrane-bound FKN as well as soluble FKN expression in synovial fibroblasts. Moreover, soluble FKN was capable of inducing IL-1beta and IL-6 by activated monocytes. These results suggest that FKN might preferentially mediate migration and recruitment of CD16+ monocytes, and might contribute to synovial tissue inflammation.
Subject(s)
Arthritis, Rheumatoid/metabolism , Arthritis, Rheumatoid/pathology , Chemokines, CX3C/metabolism , Membrane Proteins/metabolism , Monocytes/metabolism , Monocytes/pathology , Receptors, Chemokine/metabolism , Receptors, IgG/metabolism , Aged , Arthritis, Rheumatoid/blood , CX3C Chemokine Receptor 1 , Cell Membrane/metabolism , Cells, Cultured , Chemokine CX3CL1 , Chemokines, CX3C/blood , Chemokines, CX3C/pharmacology , Endothelial Cells/metabolism , Female , Humans , Interleukin-10/pharmacology , Interleukin-1beta/metabolism , Interleukin-6/metabolism , Macrophages/metabolism , Macrophages/pathology , Male , Membrane Proteins/blood , Membrane Proteins/pharmacology , Monocytes/drug effects , Osteoarthritis/blood , Osteoarthritis/metabolism , Osteoarthritis/pathology , Recombinant Proteins/pharmacology , Synovial Fluid/metabolism , Synovial Membrane/metabolism , Synovial Membrane/pathology , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
S100A8 and S100A9, two Ca2+-binding proteins of the S100 family, are secreted as a heterodimeric complex (S100A8/A9) from neutrophils and monocytes/macrophages. Serum and synovial fluid levels of S100A8, S100A9, and S100A8/A9 were all higher in patients with rheumatoid arthritis (RA) than in patients with osteoarthritis (OA), with the S100A8/A9 heterodimer being prevalent. By two-color immunofluorescence labeling, S100A8/A9 antigens were found to be expressed mainly by infiltrating CD68+ macrophages in RA synovial tissue (ST). Isolated ST cells from patients with RA spontaneously released larger amounts of S100A8/A9 protein than did the cells from patients with OA. S100A8/A9 complexes, as well as S100A9 homodimers, stimulated the production of proinflammatory cytokines, such as tumor necrosis factor alpha, by purified monocytes and in vitro-differentiated macrophages. S100A8/A9-mediated cytokine production was suppressed significantly by p38 mitogen-activated protein kinase (MAPK) inhibitors and almost completely by nuclear factor kappa B (NF-kappaB) inhibitors. NF-kappaB activation was induced in S100A8/A9-stimulated monocytes, but this activity was not inhibited by p38 MAPK inhibitors. These results indicate that the S100A8/A9 heterodimer, secreted extracellularly from activated tissue macrophages, may amplify proinflammatory cytokine responses through activation of NF-kappaB and p38 MAPK pathways in RA.
