Rheumatoid Synoviocytes Research Articles

Methyl salicylate lactoside inhibits inflammatory response of fibroblast‐like synoviocytes and joint destruction in collagen‐induced arthritis in mice

Methyl salicylate 2-O-β-d-lactoside (MSL), whose chemical structure is similar to that of salicylic acid, is a natural product derivative isolated from a traditional Chinese herb. The aim of this study was to investigate the therapeutic effect of MSL in mice with collagen-induced arthritis (CIA) and explore its underlying mechanism. The anti-arthritic effects of MSL were evaluated on human rheumatoid fibroblast-like synoviocytes (FLS) in vitro and CIA in mice in vivo by obtaining clinical scores, measuring hind paw thickness and inflammatory cytokine levels, radiographic evaluations and histopathological assessments. Treatment with MSL after the onset of arthritis significantly prevented the progression and development of rheumatoid arthritis (RA) in CIA mice without megascopic gastric mucosa damage. In addition, MSL inhibited the production of pro-inflammatory mediators, the phosphorylation and translocation of NF-κB, and cell proliferation induced by TNF-α in FLS. MSL non-selectively inhibited the activity of COX in vitro, but was a more potent inhibitor of COX-2 than COX-1. MSL also inhibited the phosphorylation of inhibitor of NF-κB kinase, IκBα and p65, thus blocking the nuclear translocation of NF-κB in TNF-α-stimulated FLS. MSL exerts therapeutic effects on CIA mice, suppressing the inflammatory response and joint destruction by non-selectively inhibiting the activity of COX and suppressing activation of the NF-κB signalling pathway, but without damaging the gastric mucosa. Therefore, MSL has great potential to be developed into a novel therapeutic agent for the treatment of RA.

AB0070 The Importance of Gliostatin as an Indicator of Disease Activity in Patients with Rheumatoid Arthritis

BackgroundGliostatin (GLS) plays an important role in joint destruction in rheumatoid arthritis (RA). GLS not only has an enzymatic activity as a thymidine phosphorylase but also has angiogenic and arthritogenic...

THU0553 Gliostatin Regulates Vascular Endothelial Growth-Factor Production in Human Fibroblast-Like Synoviocytes

BackgroundAngiogenesis, which is the growth of new capillaries from pre-existing blood vessels, is an essential feature of rheumatoid arthritis (RA). Neovascularisation might be triggered by several factors, such as cytokines,...

In vitro evaluation of effects of sustained anti-TNF release from MPEG-PCL-MPEG and PCL microspheres on human rheumatoid arthritis synoviocytes

Anti-tumor necrosis factor α (TNFα) drugs such as etanercept (ETN) have been mostly used in systemic treatment of rheumatoid arthritis. To eliminate the side effects in long-term treatments and to achieve a local sustained anti-inflammatory effect, a controlled drug delivery system is needed for anti-TNFα drugs. This study aims to develop novel injectable microcarriers of ETN that can provide long-term controlled release of this protein drug upon intra-articular application. In this study, poly(ε-caprolactone) (PCL) and its copolymer with poly(ethylene glycol), methoxypoly(ethylene glycol)-poly(ε-caprolactone)-methoxypoly(ethylene glycol) microspheres (MPEG-PCL-MPEG) were compared for their prospective success in rheumatoid arthritis treatment. Microspheres with smooth surface of a mean particle diameter of approximately 5 μm were prepared with both polymers. MPEG-PCL-MPEG microspheres had higher encapsulation efficiency than PCL microspheres. The activity of encapsulated ETN within MPEG-PCL-MPEG microspheres also retained while 90% of the activity of ETN within PCL microspheres could retain during 90-day release. MPEG-PCL-MPEG microspheres showed faster ETN release compared to PCL microspheres in various release media. Cumulative amounts of ETN released from both types of microspheres were significantly lower in cell culture medium and in synovial fluids than in phosphate buffered saline. This was mainly due to protein adsorption onto microspheres. Hydrophilic MPEG segment enhanced ETN release while preventing protein adsorption on microspheres compared to PCL. Sustained ETN release from microspheres resulted with a significant decrease in pro-inflammatory cytokines (TNFα, IFNγ, IL-6, IL-17) and MMP levels (MMP-3, MMP-13), while conserving viability of fibroblast-like synoviocytes compared to the free drug. Results suggest that MPEG-PCL-MPEG is a potential copolymer of PCL that can be used in development of biomedical materials for effective local treatment purposes in chronic inflammatory arthritis owing to enhanced hydrophilicity. Yet, PCL microspheres are also promising systems having good compatibility to synoviocytes and would be especially the choice for treatment approach requiring longer term and slower release.

PBMC and exosome-derived Hotair is a critical regulator and potent marker for rheumatoid arthritis.

Despite growing importance of long non-coding RNAs (lncRNAs) in normal physiological and disease conditions, our knowledge of RA-related lncRNAs remains limited. Therefore, we aimed to identify lncRNA signatures that have prognostic values in RA. There was a notably high expression level of Hotair in blood mononuclear cells and serum exosome of rheumatoid arthritis (RA) patients, leading the migration of active macrophage. In contrast, markedly lower level of Hotair was detected in differentiated osteoclasts and rheumatoid synoviocytes and enforced expression of Hotair led to significantly decreased levels of MMP-2 and MMP-13. This exploratory study provides novel empirical evidence that Hotair could be one of potential biomarkers for diagnosing RA.

A1.19 Zinc and cadmium homeostasis is altered in rheumatoid arthritis synoviocytes

Background and ObjectivesRheumatoid arthritis (RA) is a chronic inflammatory disease with uncontrolled proliferation of synoviocytes. Pro-inflammatory cytokines, notably IL-17 and TNF-α contribute to RA perpetuation. Zinc (Zn) is critical for...

A1.8 A dual role of MTOR in the rheumatoid mesenchymal tissue response to inflammation

BackgroundThe mechanistic target of rapamycin (mTOR) is known to couple energy and nutrient abundance to the execution of essential cellular processes, such as cellular growth and survival. As such, the...

Reduction of IL-20 Expression in Rheumatoid Arthritis by Linear Polarized Infrared Light Irradiation.

Low-level laser is being evaluated for treating rheumatoid arthritis (RA). Recently, the linear polarized infrared light (Super Lizer, SL) irradiation may also be useful for RA treatment. However, the molecular mechanism underlying the effectiveness of SL on RA is unclear. It has been IL-20 may involved in RA disease progression. To understand how SL action, we constructed the experimental model in vitro using human rheumatoid fibroblast-like synoviocyte (MH7A) and collagen induced (CIA) RA rat in vivo. We examined the effect of SL irradiation on IL-20 gene expression in MH7A and IL-20 protein production in CIA) rat joints. MH7A was cultured and challenged with IL-1ß, then examined IL-20 and IL-20R mRNA level by DNA microarray. IL-20 protein expression was examined by immunohistochemistry using a specific antibody against rat IL-20. Scatter plot analysis demonstrated that an increase in IL-20 gene expression by IL-1ß was reduced by SL irradiation, but IL-20R did not show a significant change. The Immunohistochemical analysis demonstrated a strong IL-20 staining in synovial membrane tissue of CIA rat joint, and SL irradiation significantly reduced the staining. Since IL-20 has been identified as an important cytokine in the pathogenesis of RA, the reduction of IL-20 expression by SL irradiation may be one of mechanisms in reduction of inflammation in RA joints by SL irradiation suggesting that SL irradiation may be useful for RA therapy.

TLR dependent XBP-1 activation induces an autocrine loop in rheumatoid arthritis synoviocytes

X-box binding protein 1 (XBP1) is a central regulator of the endoplasmic reticulum (ER) stress response. It is induced via activation of the IRE1 stress sensor as part of the unfolded protein response (UPR) and has been implicated in several diseases processes. XBP1 can also be activated in direct response to Toll-like receptor (TLR) ligation independently of the UPR but the pathogenic significance of this mode of XBP1 activation is not well understood. Here we show that TLR-dependent XBP1 activation is operative in the synovial fibroblasts (SF) of patients with active rheumatoid arthritis (RA). We investigated the expression of ER stress response genes in patients with active RA and also in patients in remission. The active (spliced) form of (s)XBP1 was significantly overexpressed in the active RA group compared to healthy controls and patients in remission. Paradoxically, expression of nine other ER stress response genes was reduced in active RA compared to patients in remission, suggestive of a UPR-independent process. However, sXBP1 was induced in SF by TLR4 and TLR2 stimulation, resulting in sXBP1-dependent interleukin-6 and tumour necrosis factor (TNF) production.We also show that TNF itself induces sXBP1 in SF, thus generating a potential feedback loop for sustained SF activation. These data confirm the first link between TLR-dependent XBP1 activation and human inflammatory disease. sXBP1 appears to play a central role in this process by providing a convergence point for two different stimuli to maintain activation of SF.

Identification of key regulators for the migration and invasion of rheumatoid synoviocytes through a systems approach

Rheumatoid synoviocytes, which consist of fibroblast-like synoviocytes (FLSs) and synovial macrophages (SMs), are crucial for the progression of rheumatoid arthritis (RA). Particularly, FLSs of RA patients (RA-FLSs) exhibit invasive characteristics reminiscent of cancer cells, destroying cartilage and bone. RA-FLSs and SMs originate differently from mesenchymal and myeloid cells, respectively, but share many pathologic functions. However, the molecular signatures and biological networks representing the distinct and shared features of the two cell types are unknown. We performed global transcriptome profiling of FLSs and SMs obtained from RA and osteoarthritis patients. By comparing the transcriptomes, we identified distinct molecular signatures and cellular processes defining invasiveness of RA-FLSs and proinflammatory properties of RA-SMs, respectively. Interestingly, under the interleukin-1β (IL-1β)-stimulated condition, the RA-FLSs newly acquired proinflammatory signature dominant in RA-SMs without losing invasive properties. We next reconstructed a network model that delineates the shared, RA-FLS-dominant (invasive), and RA-SM-dominant (inflammatory) processes. From the network model, we selected 13 genes, including periostin, osteoblast-specific factor (POSTN) and twist basic helix-loop-helix transcription factor 1 (TWIST1), as key regulator candidates responsible for FLS invasiveness. Of note, POSTN and TWIST1 expressions were elevated in independent RA-FLSs and further instigated by IL-1β. Functional assays demonstrated the requirement of POSTN and TWIST1 for migration and invasion of RA-FLSs stimulated with IL-1β. Together, our systems approach to rheumatoid synovitis provides a basis for identifying key regulators responsible for pathological features of RA-FLSs and -SMs, demonstrating how a certain type of cells acquires functional redundancy under chronic inflammatory conditions.

AB0046 The sp1 transcription factor is essential for the expression of gliostatin/thymidine phosphorylase in rheumatoid fibroblast-like synoviocytes

BackgroundGliostatin/thymidine phosphorylase (GLS) has angiogenic and arthritogenic activities, and GLS production has been observed in the active synovial membranes of rheumatoid arthritis (RA) patients. The human GLS gene promoter contains...

THU0077 Multimeric Adiponectin Isoforms in Rheumatoid Arthritis: Local and Systemic Effects

BackgroundMultimeric adiponectin isoforms of high-, middle-, and low-molecular weight (HMW, MMW and LMW) exist in body fluids. Their implication in rheumatoid arthritis (RA) pathogenesis is suggested but not fully confirmed.Objectives(1)...

THU0004 The Role of Histone Deacetylases in Rheumatoid Arthritis Fibroblasts-Like Synoviocytes

BackgroundRheumatoid arthritis (RA) is a chronic, autoimmune, inflammatory disease that affects synovial joints. A key characteristic of RA is hyperplasia of the synovial lining layer, which includes fibroblast-like synoviocytes (FLS)....

SAT0047 The origin and biological activities of classical adipokines in rheumatoid joint

BackgroundClassical adipokines, leptin and adiponectin, are thought to originate principally from white adipose tissue. Although both of them have been suggested to participate in pathogenesis of rheumatoid arthritis (RA), their...

SAT0060 Alpha-defensin-1 induces IL-6 and MMP-1 expression in rheumatoid fibroblast-like synoviocytes via ERK and/ORJNK signaling pathway

BackgroundFibroblast-like synoviocytes (FLS) play an essential role in the pathophysiology of rheumatoid arthritis (RA). Neutrophils play an important role in overt inflammation in RA and are the most abundant cell...

SeMet Inhibits IL-1β-Induced Rheumatoid Fibroblast-Like Synoviocytes Proliferation and the Production of Inflammatory Mediators

Previous studies demonstrated that Se has anti-inflammatory activities and that it plays an important role in maintaining normal cartilage metabolism. Nevertheless, little is known about the effects of Se on the production of inflammatory mediators in rheumatoid fibroblast-like synoviocytes (FLSs). The objective of this study was to determine the effects of Se on the interleukin-1β (IL-1β)-induced proliferation of FLSs and production of matrix metalloproteinases (MMPs) and inflammatory mediators by FLSs. In this study, the proliferation of FLSs was assessed using the MTT assay after cultured with/without the presence of IL-1β and SeMet. Human FLSs were pretreated with SeMet (0.5μM) and subsequently stimulated with IL-1β (5ng/ml) for 24h. Production of NO and PGE2 were evaluated by the Griess reaction and ELISA. Gene expression of MMP-3, MMP-13, iNOS, and COX-2 was measured by real-time PCR. MMP-3 and MMP-13 proteins in culture medium were determined using cytokine-specific ELISA. Western immunoblotting was used to analyze the iNOS and COX-2 protein production in the culture medium and the activity of phosphorylation of P38 MAPK pathways. We found that SeMet significantly inhibits IL-1β-induced proliferation of FLSs. SeMet also inhibited the production of PGE2 and NO induced by IL-1β. SeMet significantly decreased IL-1β-stimulated gene expression and production of MMP-3, MMP-13, iNOS, and COX-2 in human FLSs. In addition, we found SeMet partly inhibited the IL-1β-induced activation of p38 MAPK pathways. The present report is first to demonstrate that SeMet inhibits IL-1β-induced expression of MMPs and production of inflammatory factors in cultured FLSs, indicating that SeMet may be a potential agent in the treatment of rheumatoid arthritis.

Selective Inhibition of Human Group IIA-secreted Phospholipase A2 (hGIIA) Signaling Reveals Arachidonic Acid Metabolism Is Associated with Colocalization of hGIIA to Vimentin in Rheumatoid Synoviocytes

Human group IIA secreted phospholipase A2 (hGIIA) promotes tumor growth and inflammation and can act independently of its well described catalytic lipase activity via an alternative poorly understood signaling pathway. With six chemically diverse inhibitors we show that it is possible to selectively inhibit hGIIA signaling over catalysis, and x-ray crystal structures illustrate that signaling involves a pharmacologically distinct surface to the catalytic site. We demonstrate in rheumatoid fibroblast-like synoviocytes that non-catalytic signaling is associated with rapid internalization of the enzyme and colocalization with vimentin. Trafficking of exogenous hGIIA was monitored with immunofluorescence studies, which revealed that vimentin localization is disrupted by inhibitors of signaling that belong to a rare class of small molecule inhibitors that modulate protein-protein interactions. This study provides structural and pharmacological evidence for an association between vimentin, hGIIA, and arachidonic acid metabolism in synovial inflammation, avenues for selective interrogation of hGIIA signaling, and new strategies for therapeutic hGIIA inhibitor design.

The role of -defensin-1 and related signal transduction mechanisms in the production of IL-6, IL-8 and MMPs in rheumatoid fibroblast-like synoviocytes

To investigate the effect of α-defensin-1 on the expression of IL-6, IL-8 and MMPs as well as the signal transduction mechanisms responsible for their expression in RA fibroblast-like synoviocytes (FLS). The concentrations of α-defensin-1 in SF were measured by ELISA. In RA FLS, mRNA expression of IL-6, IL-8 and MMPs and activation of signalling molecules were examined by real-time PCR, western blotting and electrophoretic mobility shift assay. Concentrations of SF α-defensin-1 were significantly increased in RA patients compared with OA patients. The levels of mRNA expression of IL-6, IL-8, MMP-1 and MMP-3 were significantly increased in RA FLS treated with α-defensin-1 compared with controls. Furthermore, α-defensin-1 activated JNK and ERK in RA FLS, respectively. Treatment of RA FLS with ERK or JNK inhibitors prior to α-defensin-1 treatment resulted in reduced expression of IL-6, IL-8, MMP-1, and MMP-3 compared with controls. Remarkably, treatment of RA FLS with an ERK inhibitor prior to α-defensin-1 stimulation significantly reduced production of IL-6 and MMP-1 by approximately 71% and 98% compared with controls, respectively. The JNK inhibitor significantly suppressed α-defensin-1-induced MMP-1 production by approximately 73% compared with controls. Finally, there was a significant induction of NF-κB DNA binding activity in response to α-defensin-1. Our results suggest that α-defensin-1 may play a role in RA pathogenesis by regulating the production of MMPs as well as IL-6 and IL-8. These processes were dependent on the regulation of the JNK and/or ERK and NF-κB pathways.

Regulation of DNA Methylation in Rheumatoid Arthritis Synoviocytes

Rheumatoid arthritis (RA) is a chronic inflammatory disease in which fibroblast-like synoviocytes (FLS) exhibit an aggressive phenotype. Although the mechanisms responsible are not well defined, epigenetic determinants such as DNA methylation might contribute. DNA methyltransferases (DNMTs) are critical enzymes that establish and maintain DNA methylation. We evaluated whether proinflammatory cytokines might contribute to differential DNA methylation previously described in RA FLS through altered DNMT expression. FLS were obtained from RA and osteoarthritis (OA) synovium at the time of total joint replacement. Gene expression was determined by quantitative real-time PCR and protein expression by Western blot analysis. DNMT activity was measured with a functional assay, and global methylation was determined by an immunoassay that detects methylcytosine. Resting expression of DNMT1, -3a, and -3b mRNA were similar in RA and OA FLS. Western blot showed abundant DNMT1 and DNMT3a protein. Exposure to IL-1 decreased DNMT1 and DNMT3a mRNA expression in FLS. Dose responses demonstrated decreased DNMT expression at concentrations as low as 1 pg/ml of IL-1. DNMT mRNA levels decreased rapidly, with significant suppression after 2-8 h of IL-1 stimulation. IL-1 stimulation of OA FLS did not affect methylation of LINE1 sites but led to demethylation of a CHI3L1 locus that is hypomethylated in RA FLS. Chronic IL-1 stimulation also mimicked the effect of a DNMT inhibitor on FLS gene expression. Exposure to proinflammatory mediators reversibly alters DNA methylation in FLS by decreasing DNMT expression and function. These data suggest that IL-1 can potentially imprint cells in chronic inflammatory diseases.

Serum amyloid A (SAA) induces pentraxin 3 (PTX3) production in rheumatoid synoviocytes

Objective: Pentraxin 3 (PTX3) is an acute-phase reactant that is involved in amplification of the inflammatory response and innate immunity. In the present study, we evaluated the relationship between PTX3 and serum amyloid A (SAA), another acute-phase reactant, in rheumatoid synoviocytes.Methods: PTX3 mRNA expression was examined by reverse transcription polymerase chain reaction, and PTX3 protein was measured by enzyme-linked immunosorbent assay.Results: SAA induced PTX3 mRNA and PTX3 protein expression in rheumatoid synoviocytes. SAA-induced PTX3 expression was attenuated when rheumatoid synoviocytes were nucleofected with N-formyl peptide receptor ligand-1 (FPRL-1)-specific siRNA, suggesting the involvement of FPRL-1. Furthermore, SAA-induced PTX3 expression was inhibited by NF-κB or mitogen-activated protein kinase-specific inhibitors. Neither soluble TNF receptor (etanercept) nor recombinant IL-1 receptor antagonist affected PTX3 production by SAA-stimulated synoviocytes, suggesting that SAA directly induces PTX3.Conclusion: Our data suggest that SAA plays a role in the proinflammatory and immune responses in rheumatoid synovium by inducing PTX3. We provide the first evidence that the acute-phase reactant SAA, which is produced systemically by hepatocytes, perpetuates the rheumatoid inflammatory processes by inducing another proinflammatory molecule, PTX3, locally in rheumatoid synovial tissues.