Potential Therapeutic Target For Rheumatoid Arthritis Research Articles

Germinal Center Kinase–like Kinase Overexpression in T Cells as a Novel Biomarker in Rheumatoid Arthritis

Germinal center kinase-like kinase (GLK; also called MAPKKKK-3) activates protein kinase Cθ (PKCθ) during T cell activation and controls autoimmunity in lupus patients. Intracellular kinases are involved in the pathogenesis of rheumatoid arthritis (RA). We undertook this study to determine the role of GLK in RA. The severity of collagen-induced arthritis (CIA) was studied in GLK-deficient mice. Expression levels of GLK from RA patients were determined by Western blotting, flow cytometry, real-time polymerase chain reaction, and immunohistochemical staining. Localization of GLK in T cells was identified by confocal microscopy. RA disease activity was assessed using the Disease Activity Score in 28 joints. GLK-deficient mice displayed impaired CIA development and decreased inflammatory cytokine levels. Local T cell infiltration and collagen restimulation responses were impaired by GLK deficiency. RA patients showed significantly higher GLK protein and messenger RNA levels in peripheral blood T cells than did healthy controls. GLK-overexpressing T cells in synovial fluid and synovial tissue samples from RA patients were increased compared with those from osteoarthritis patients. Confocal microscopy and flow cytometry showed that GLK colocalized and coexisted with phosphorylated PKCθ in T cells from RA patients. Frequencies of GLK-expressing T cells were significantly correlated with RA disease activity. GLK overexpression in T cells contributes to the pathogenesis of RA, indicating that GLK is a novel biomarker for autoimmune disease severity and a potential therapeutic target for RA.

Interleukin-27 as a potential therapeutic target for rheumatoid arthritis: has the time come?

Interleukin (IL)-27 is a novel member of the IL-6/IL-12 family of cytokines with a broad range of pro- and anti-inflammatory properties. Recently, accumulating evidence has shown that IL-27 can play either a pathogenic or a protective role in animal models of inflammatory arthritis, depending upon the model and underlying pathogenic mechanisms. As to human system, elevated expression of IL-27 has clearly been detected in the synovial membranes and fluid from patients with rheumatoid arthritis (RA). Moreover, stimulation of IL-27 receptor with IL-27 of fibroblast-like synoviocytes from RA had a suppressive effect on the production of proinflammatory cytokines in vitro. All these findings suggest that IL-27 may have promise as a potential therapeutic target for RA. In this review, we will discuss the biological features of IL-27 and summarize recent advances on both pathogenic and protective roles of IL-27 in RA.

Deficient or abundant but unable to fight? Estimation of circulating FoxP3+ T regulatory cells and their counteracting FoxP3− in rheumatoid arthritis and correlation with disease activity

Aim of the workPersistent inflammation and recurring activity in rheumatoid arthritis (RA) daring a quarterback of T regulatory cells (Tregs) intrigued rheumatologists. Tregs’ most specific marker is the forkhead box P3 (FoxP3) denoting FoxP3+ cells as suppressors whereas FoxP3− as effectors. This study evaluates subset distribution of peripheral blood (PB) CD4+CD25+ Tregs according to FoxP3 expression in RA to better understand its role in pathogenesis. Patients and methodsIn our observational cross-sectional study PB Tregs from 40 RA patients and 20 age and sex matched healthy controls (HC) were characterized and quantified by flow cytometry. Disease activity was evaluated by DAS28. Patients were divided into: active RA group (ARA) and remission RA group (RRA). ResultsSignificantly higher CD4+CD25+FoxP3+ Tregs were found on comparing RRA, ARA patients and HC (mean 153.25±6.29, 136.3±3.27 and 97.25±6.25, respectively) with a statistically highly significant difference (F=553.13, p<0.001). CD4+CD25+FoxP3− increased 3-folds in RRA and 4-folds in ARA compared to HC and CD4+CD25+FoxP3+ increased 1.6-folds in RRA and 1.5-folds in ARA. Thus the ratio of FoxP3−/FoxP3+ cells is altered from 1:3 in HC to 2:3 in RRA and 1:1 in ARA. CD4+CD25+FoxP3+ had a highly significant negative correlation with disease activity score DAS28 (r=−0.86, p<0.001). ConclusionThough CD4+CD25+FoxP3+ and CD4+CD25+FoxP3− are both increased in RA, their balance is more important; abundant FoxP3− cells with effector function have the upper hand over suppressive FoxP3+ cells. This imbalance relates to RA presence and activity. Reinforcing FoxP3 expression might be a good potential therapeutic target in RA.

Interleukin 22, a Potential Therapeutic Target for Rheumatoid Arthritis

To the Editor: A recent report by Da Rocha, et al indicated that levels of interleukin 22 (IL-22) were increased in patients with rheumatoid arthritis (RA) compared with controls; levels of IL-22 correlated with Disease Activity Score (DAS28) and Clinical Disease Activity Index measures, rheumatoid factor positivity was correlated with higher levels of IL-22 in patients with RA, and the presence of bone erosions was associated with high IL-22 levels1. These findings suggest that IL-22 may be a good biomarker for assessment of activity in RA, and IL-22 seems to be a potential therapeutic target for RA. Other studies have indicated the similar relationship between IL-22 and RA, … Address correspondence to Dr. Li; E-mail: lijun{at}ahmu.edu.cn

Gene expression profiling and functional analysis of angiogenic markers in murine collagen-induced arthritis

IntroductionDysregulated angiogenesis is implicated in the pathogenesis of rheumatoid arthritis (RA). To provide a more profound understanding of arthritis-associated angiogenesis, we evaluated the expression of angiogenesis-modulating genes at onset, peak and declining phases of collagen-induced arthritis (CIA), a well-established mouse model for RA.MethodsCIA was induced in DBA/1 mice with type II collagen. Functional capillary density in synovial tissue of knee joints was determined by intravital fluorescence microscopy. To assess the ability of arthritic joint homogenates to induce angiogenesis, an endothelial chemotaxis assay and an in vivo matrigel plug assay were employed. The temporal expression profile of angiogenesis-related genes in arthritic paws was analysed by quantitative real-time RT-PCR using an angiogenesis focused array as well as gene specific PCR. Finally, we investigated the therapeutic effect of a monoclonal antibody specifically blocking the binding of VEGF to neuropilin (NRP)-1.ResultsAlthough arthritic paw homogenates displayed angiogenic activity in vitro and in vivo, and synovia of arthritic paws appeared highly vascularised on histological examination, the functional capillary density in arthritic knee synovia was significantly decreased, whereas capillary diameter was increased. Of the 84 genes analysed, 41 displayed a differential expression in arthritic paws as compared to control paws. Most significant alterations were seen at the peak of clinical arthritis. Increased mRNA expression could be observed for VEGF receptors (Flt-1, Flk-1, Nrp-1, Nrp-2), as well as for midkine, hepatocyte growth factor, insulin-like growth factor-1 and angiopoietin-1. Signalling through NRP-1 accounted in part for the chemotactic activity for endothelial cells observed in arthritic paw homogenates. Importantly, therapeutic administration of anti-NRP1B antibody significantly reduced disease severity and progression in CIA mice.ConclusionsOur findings confirm that the arthritic synovium in murine CIA is a site of active angiogenesis, but an altered balance in the expression of angiogenic factors seems to favour the formation of non-functional and dilated capillaries. Furthermore, our results validate NRP-1 as a key player in the pathogenesis of CIA, and support the VEGF/VEGF receptor pathway as a potential therapeutic target in RA.

Adiponectin is a predictor for radiographic progression in early RA patients, independently of anti-CCP antibodies

Background and objectivesAdipose tissue can secrete soluble mediators (adipokines) with potent immune regulatory functions. Some adipokines have been previously associated with radiographic damage in rheumatoid arthritis (RA) in cross-sectional studies....

REDUCTION OF CXCR4 EXPRESSION IN RHEUMATOID ARTHRITIS RAT JOINTS BY LOW LEVEL DIODE LASER IRRADIATION

Rheumatoid arthritis (RA) is an inflammatory joint disorder, whose progression leads to the destruction of cartilage and bone. Chemokines and their receptors are potential therapeutic targets in RA. Among these, it has been suggested that CXC chemokine 4 (CXCR4) and its ligand CXC ligand 12 (CXCL12) are involved in RA pathogenesis. Low-level laser irradiation (LLLI) is currently being evaluated for the treatment of RA; however, the molecular mechanisms underlying its effectiveness remain unclear. To understand the anti-inflammatory effect of LLLI, we used the collagen-induced arthritis (CIA) rat as RA model and analyzed the gene expression profile in synovial membrane in the hindpaw joints of control, CIA and CIA + LLLI. Expression of CXCR4 and CXCL12 genes were also studied. Total RNA was isolated from the synovial membrane tissue of CIA rat joints or CIA joints treated with LLLI (830 nm Ga-Al-As diode), and gene expression profiles were analyzed by DNA microarray (41,000 rat genes). The mRNA levels were confirmed by reverse transcription polymerase chain reaction (RT-PCR) and real-time PCR. Immunohistochemical examination to examine CXCR4 protein expression was also carried out. DNA microarray analysis showed that CXCR4 gene expression was increased in CIA tissue and LLLI treatment significantly decreased CIA-induced CXCR4 mRNA levels. In contrast, CXCL12 did not show any significant changes. The microarray data of CXCR4 mRNA levels were further validated using RT-PCR and real-time PCR. Increased CXCR4 mRNA levels by CIA and its reduction following LLLI was successfully confirmed. CXCR4 production was increased in CIA joints and its production was decreased by LLLI. Decreased CXCR4 expression may be one of the mechanisms in LLLI-mediated reduction of RA inflammation.

The adaptor protein CIKS/ACT1 is necessary for collagen‐induced arthritis, and it contributes to the production of collagen‐specific antibody

CIKS/ACT1 is an adaptor molecule that is necessary for signaling by members of the interleukin-17 cytokine family. The aim of this study was to determine whether this adaptor is required for the initiation of collagen-induced arthritis (CIA). If it is required, then CIKS-mediated signaling could be a potential target for therapeutic intervention in patients with rheumatoid arthritis (RA). CIA model studies were performed with CIKS-deficient and CIKS-sufficient mice on an otherwise wild-type (WT) C57BL/6 background or on a C57BL/6 background lacking Fcγ receptor IIb (FcγRIIb). In addition, collagen antibody-induced arthritis (CAIA) studies were performed in WT and CIKS-deficient mice. Pathologic changes of arthritis were evaluated by visual inspection of the paws, by histochemical analysis of tissue sections, and by measurements of collagen-specific antibodies. Pathologic changes of CIA were readily induced in WT mice, with exacerbation of the changes in FcγRIIb-deficient mice. In contrast, CIKS-deficient mice were protected from all aspects of CIA pathology, even on an FcγRIIb-deficient background. The absence of CIKS completely prevented neutrophil infiltration into joints, bone erosion, and cartilage damage; furthermore, the production of type II collagen (CII)-specific antibodies was reduced. In contrast to the CIA model, CIKS-deficient mice in the CAIA model remained susceptible to arthritis. CIKS-mediated signaling is necessary for the pathogenesis of CIA, but not CAIA. These findings suggest critical functions of CIKS during the development of arthritis in the CIA model, including in the formation of CII antibodies, and they mark the CIKS adaptor as a potential therapeutic target in RA.

SIGIRR/TIR-8 is an inhibitor of toll-like receptor signaling in primary human cells and regulates inflammation in models of rheumatoid arthritis

Single-immunoglobulin interleukin-1 receptor-related (SIGIRR), which is also known as Toll/interleukin-1 receptor 8 (TIR-8), is a member of the TIR domain-containing family of receptors and was first characterized as an inhibitor of interleukin-1 receptor (IL-1R) and Toll-like receptor (TLR) signaling. In the Dextran sulfate sodium-induced colitis model, SIGIRR(-/-) mice were shown to have increased inflammation and to be more susceptible to endotoxin challenge. Increasing evidence implicates TLR and IL-1R signaling in the pathology of rheumatoid arthritis (RA). Therefore, the purpose of this study was to investigate the involvement of SIGIRR in regulating inflammation in disease-relevant models. Primary human monocyte-derived macrophages and dendritic cells (DCs) were used to overexpress SIGIRR as well as to knock down endogenously expressed SIGIRR using small interfering RNAs. SIGIRR was also overexpressed in synovial cells derived from RA patients. To investigate the role of SIGIRR in vivo, zymosan-induced arthritis (ZIA) and collagen antibody-induced arthritis (CAIA) were induced in SIGIRR-knockout mice. SIGIRR overexpression inhibited TLR-induced cytokine production in macrophages and DCs, while SIGIRR knockdown resulted in increased cytokine production following TLR stimulation. Moreover, SIGIRR overexpression inhibited the spontaneous release of cytokines by human RA synovial cells. The role of SIGIRR as an inhibitor of inflammation was confirmed in vivo, since SIGIRR(-/-) mice developed a more severe disease in both the ZIA and CAIA models. Our study is the first to show the expression pattern and function of SIGIRR in primary human cells. Furthermore, this investigation defines the role of SIGIRR in disease-relevant cell types and demonstrates that SIGIRR is a potential therapeutic target for RA.

CIKS/Act1 is required for collagen-induced arthritis (93.3)

Abstract Collagen-induced arthritis (CIA) is a widely used animal model for human rheumatoid arthritis (RA). Th17 cells are thought to be critical for development of RA and CIA, mediated in part via production of IL-17. CIKS/Act1 is an important adaptor molecule for signaling by IL-17 and related cytokines. Although evidence suggests a role for IL-17 in RA, the physiologic relevance of CIKS/Act1 in RA and the CIA model has not been evaluated. We have applied the CIA model to CIKS deficient and wild-type (WT) B6 mice. Based on clinical scores and histochemical analyses, arthritis could be induced in WT, but not CIKS deficient mice. The production of collagen type 2-specific antibodies (CII-Abs) and neutrophil recruitment to the joint were also blocked in CIKS deficient mice. We also performed CIA model studies in mice with compound deficiency in CIKS and FcgammaRIIb (B6). Although the loss of FcgammaRIIb greatly exacerbated arthritis in the CIA model in CIKS-sufficient mice, arthritis and production of CII-Abs were still blocked in the absence of CIKS. In addition we determined that CIKS is not required in the collagen antibody-induced arthritis (CAIA) model, in contrast to this adaptor’s role in CIA. These data suggest a crucial role of CIKS during the generation of CII-Abs and clearly implicate the CIKS adaptor as essential for development of CIA and thus a potential therapeutic target in RA.

Slug suppression induces apoptosis via Puma transactivation in rheumatoid arthritis fibroblast-like synoviocytes treated with hydrogen peroxide.

Inadequate apoptosis contributes to synovial hyperplasia in rheumatoid arthritis (RA). Recent study shows that low expression of Puma might be partially responsible for the decreased apoptosis of fibroblast-like synoviocytes (FLS). Slug, a highly conserved zinc finger transcriptional repressor, is known to antagonize apoptosis of hematopoietic progenitor cells by repressing Puma transactivation. In this study, we examined the expression and function of Slug in RA FLS. Slug mRNA expression was measured in the synovial tissue (ST) and FLS obtained from RA and osteoarthritis patients. Slug and Puma mRNA expression in FLS by apoptotic stimuli were measured by real-time PCR analysis. FLS were transfected with control siRNA or Slug siRNA. Apoptosis was quantified by trypan blue exclusion, DNA fragmentation and caspase-3 assay. RA ST expressed higher level of Slug mRNA compared with osteoarthritis ST. Slug was significantly induced by hydrogen peroxide (H2O2) but not by exogenous p53 in RA FLS. Puma induction by H2O2 stimulation was significantly higher in Slug siRNA-transfected FLS compared with control siRNA-transfected FLS. After H2O2 stimulation, viable cell number was significantly lower in Slug siRNA-transfected FLS compared with control siRNA-transfected FLS. Apoptosis enhancing effect of Slug siRNA was further confirmed by ELISA that detects cytoplasmic histone-associated DNA fragments and caspase-3 assay. These data demonstrate that Slug is overexpressed in RA ST and that suppression of Slug gene facilitates apoptosis of FLS by increasing Puma transactivation. Slug may therefore represent a potential therapeutic target in RA.

Akt activity protects rheumatoid synovial fibroblasts from Fas-induced apoptosis by inhibition of Bid cleavage

IntroductionSynovial hyperplasia is a main feature of rheumatoid arthritis pathology that leads to cartilage and bone damage in the inflamed joints. Impaired apoptosis of resident synoviocytes is pivotal in this process. Apoptosis resistance seems to involve defects in the extrinsic and intrinsic apoptotic pathways. The aim of this study was to investigate the association of PI3Kinase/Akt and the mitochondrial apoptotic pathway in the resistance of rheumatoid arthritis (RA) fibroblast like synovial cells (FLS) to Fas-mediated apoptosis.MethodsApoptosis was assessed by ELISA quantification of nucleosomal release, Hoechst staining and activated caspase-3/7 measure in cultured RA FLS stimulated with anti-Fas antibody. Two Phosphoinositol-3-kinase/protein Kinase B (PI3 Kinase) inhibitors, Wortmannine and LY294002, were used before anti-Fas stimulation. Proapoptotic BH3 interacting domain death agonist (Bid) was suppressed in RA FLS by small interfering RNA (siRNA) transfection. Bid was overexpressed by transfection with the pDsRed2-Bid vector. Phosphorylated Akt, caspase-9, and Bid expression were analysed by western blot.ResultsPI3 kinase inhibition sensitizes RA FLS to Fas-induced apoptosis by increasing cleavage of Bid protein. Bid suppression completely abrogated Fas-induced apoptosis and Bid overexpression highly increased apoptotic rate of RA FLS in association with cleavage of caspase-9.ConclusionsIn RA FLS, phosphorylation of Akt protects against Fas-induced apoptosis through inhibition of Bid cleavage. The connection between the extrinsic and the intrinsic apoptotic pathways are critical in this Fas- mediated apoptosis and points to PI3Kinase as potential therapeutic target for RA.

Visfatin: A Potential Therapeutic Target for Rheumatoid Arthritis

Rheumatoid arthritis (RA) is usually a chronic, systemic inflammatory disorder primarily targeting the synovium and articular cartilage. It is incurable, costly and responds poorly to treatment. Methotrexate alone or in combination with conventional and/or biological disease-modifying antirheumatic drugs (DMARDs) is often used to induce remission of active disease. The effectiveness of treatment is, however, limited and most patients develop chronic disability and require total knee arthroplasty or total hip replacement. Emerging therapies targeting specific cytokines and growth factors in the RA inflammatory cascade offer potent new means of modifying disease activity. Recently, increased concentrations of adipokines, including visfatin, mainly produced by adipocytes in serum and joint synovial fluid, were found in RA patients. Visfatin has important pro-inflammatory and catabolic roles in RA pathogenesis and is now being studied as a potential therapeutic target for RA. Here we discuss the relationship between visfatin and RA and its potential as a therapeutic target for RA.

Altered collagen II 263–272 peptide immunization induces inhibition of collagen‐induced arthritis through a shift toward Th2‐type response

To investigate the effects of altered collagen II (CII) peptide ligands in collagen-induced arthritis (CIA), CIA rats were subcutaneously injected with an altered CII263-272 peptide ligand (APL, 100, 10, and 1 microg/dose, six dose each, twice a week), which had been identified by us as an inhibitory effect on CII-specific T-cell activation in vitro in rheumatoid arthritis (RA). Clinical, radiographic, and histologic scores were evaluated. Serum level of interferon (IFN)-gamma was assessed by enzyme-linked immunosorbent assay, and the numbers of interleukin (IL)-4-producing cells in vitro in memory responses to the APL were determined by enzyme-linked immunospot. The results showed significantly reduced arthritis scores in CIA rats treated with 100 microg/dose of APL compared with those treated with phosphate buffered solution (PBS) and control peptide. The mean radiographic and histologic scores were also markedly lower in APL-treated CIA rats. On day 35 after immunization, a significantly lower level of IFN-gamma in serum was found in APL-treated rats, accompanied by increased numbers of IL-4-producing cells, indicating that APL treatment skewed the T helper (Th)1/Th2 balance toward Th2-type response in vivo. Altered CII263-272 peptide ligand immunization induces inhibition of CIA through a shift toward Th2-type response, suggesting that altered CII peptide might be a potential therapeutic target for RA.

Ectopic lymphoid structures support ongoing production of class-switched autoantibodies in rheumatoid synovium.

BackgroundFollicular structures resembling germinal centres (GCs) that are characterized by follicular dendritic cell (FDC) networks have long been recognized in chronically inflamed tissues in autoimmune diseases, including the synovium of rheumatoid arthritis (RA). However, it is debated whether these ectopic structures promote autoimmunity and chronic inflammation driving the production of pathogenic autoantibodies. Anti-citrullinated protein/peptide antibodies (ACPA) are highly specific markers of RA, predict a poor prognosis, and have been suggested to be pathogenic. Therefore, the main study objectives were to determine whether ectopic lymphoid structures in RA synovium: (i) express activation-induced cytidine deaminase (AID), the enzyme required for somatic hypermutation and class-switch recombination (CSR) of Ig genes; (ii) support ongoing CSR and ACPA production; and (iii) remain functional in a RA/severe combined immunodeficiency (SCID) chimera model devoid of new immune cell influx into the synovium.Methods and FindingsUsing immunohistochemistry (IHC) and quantitative Taqman real-time PCR (QT-PCR) in synovial tissue from 55 patients with RA, we demonstrated that FDC+ structures invariably expressed AID with a distribution resembling secondary lymphoid organs. Further, AID+/CD21+ follicular structures were surrounded by ACPA+/CD138+ plasma cells, as demonstrated by immune reactivity to citrullinated fibrinogen. Moreover, we identified a novel subset of synovial AID+/CD20+ B cells outside GCs resembling interfollicular large B cells. In order to gain direct functional evidence that AID+ structures support CSR and in situ manufacturing of class-switched ACPA, 34 SCID mice were transplanted with RA synovium and humanely killed at 4 wk for harvesting of transplants and sera. Persistent expression of AID and Iγ-Cμ circular transcripts (identifying ongoing IgM-IgG class-switching) was observed in synovial grafts expressing FDCs/CD21L. Furthermore, synovial mRNA levels of AID were closely associated with circulating human IgG ACPA in mouse sera. Finally, the survival and proliferation of functional B cell niches was associated with persistent overexpression of genes regulating ectopic lymphoneogenesis.ConclusionsOur demonstration that FDC+ follicular units invariably express AID and are surrounded by ACPA-producing plasma cells provides strong evidence that ectopic lymphoid structures in the RA synovium are functional and support autoantibody production. This concept is further confirmed by evidence of sustained AID expression, B cell proliferation, ongoing CSR, and production of human IgG ACPA from GC+ synovial tissue transplanted into SCID mice, independently of new B cell influx from the systemic circulation. These data identify AID as a potential therapeutic target in RA and suggest that survival of functional synovial B cell niches may profoundly influence chronic inflammation, autoimmunity, and response to B cell–depleting therapies.

Cross-talk between MEK1/2-ERK1/2 signaling and G protein-couple signaling in synoviocytes of collagen-induced arthritis rats

Signaling pathways that regulate the production of cytokines and destructive enzymes have been implicated in rheumatoid arthritis (RA) pathogenesis. There are co-relations between signaling pathways. The aim of this study was to investigate interactions and cross-talks between MEK1/2-extracellular signal-related kinase (ERK1/2) signaling and G protein-couple signaling in synoviocytes of collagen-induced arthritis (CIA) rats by the stimulation of interleukin-1 (IL-1), U0126, isoprenaline hydrochloride and aminophylline respectively. Twenty Sprague-Dawley (SD) rats were induced by chicken type II collagen. Synoviocytes of CIA rats were isolated and cultured. The expressions of Gi, phosphorylated MEK1/2 (p-MEK1/2) and phosphorylated ERK1/2 (p-ERK1/2) were detected by Western blotting. cAMP level and protein kinase A (PKA) activity were measured by radioimmunoassay and kinase-glo luminescent kinase assay respectively. There was remarkable inflammation in CIA rats accompanied by swelling paws, hyperplastic synovium, pannus and cartilage erosion. cAMP level and PKA activity of synoviocytes decreased. Gi, p-ERK1/2 and p-MEK1/2 increased. rIL-1alpha improved the expression of Gi, p-ERK1/2 and p-MEK1/2. cAMP and PKA increased with stimulation of rIL-1alpha. U0126 inhibited Gi, cAMP and PKA of synoviocytes stimulated by rIL-1alpha. Isoprenaline hydrochloride enhanced Gi, cAMP and PKA, but had no effects on p-MEK1/2 and p-ERK1/2. Aminophylline increased cAMP and PKA, but inhibited p-MEK1/2 and p-ERK1/2. Mitogen-activated protein kinases (MAPKs) and G protein-couple signaling are associated with synovitis. There are cross talks between MAPKs and G protein-couple signaling. The two signaling pathways represent potential therapeutic targets for RA.

MAPK phosphatases as novel targets for rheumatoid arthritis

Background: Rheumatoid arthritis (RA) represents a challenge for therapeutic interventions due to complex inflammatory signalling pathways underlying its pathogenesis. The MAPK signalling network, a major effector limb of the inflammatory lesion, is an attractive therapeutic target. MAPK phosphatases (MKPs), endogenous negative regulators of MAPK signalling, have received increasing recognition as modulators of inflammatory and immune responses, and hence as a potential therapeutic avenue for RA. Objective: To present the rationale for therapeutically targeting MAPK signalling and explore the case for addressing MKP1 as a novel therapeutic strategy for RA. Methods: We summarise literature describing the importance of MAPK signalling in RA and review reports describing the roles of MKPs in modulating innate and adaptive immune responses. Finally we expand on the role of MKP1 in RA pathogenesis and explore data defining MKP1 as a mediator of glucocorticoid action. Conclusion: MKP1 constitutes an exciting, novel potential therapeutic target for RA.

Light up‐regulated on B lymphocytes and monocytes in rheumatoid arthritis mediates cellular adhesion and metalloproteinase production by synoviocytes

To study the expression of LIGHT (tumor necrosis factor superfamily 14) and herpesvirus entry mediator (HVEM; tumor necrosis factor receptor superfamily 14) in rheumatoid arthritis (RA) and to determine the regulatory role of LIGHT on the effector functions of fibroblast-like synoviocytes (FLS). The expression of LIGHT and HVEM was assessed by immunohistochemical staining of synovial tissue and by flow cytometric analysis of mononuclear cells. The presence of HVEM and lymphotoxin beta receptor was measured by reverse transcriptase-polymerase chain reaction and by flow cytometry. The regulation of effector molecules, including matrix metalloproteinases (MMPs) and adhesion molecules, was evaluated. The adhesiveness of FLS was determined by adhesion assay. HVEM was detected in most cell types within rheumatoid synovial tissue, while only a few cells were positive for LIGHT. In RA patients, LIGHT expression was significantly up-regulated only in CD20+ B cells and monocytes, whereas the mean fluorescence intensity of HVEM was down-regulated in mononuclear cells. The stimulation of FLS with LIGHT resulted in the production of MMPs and the expression of adhesion molecules, which were efficiently inhibited by dexamethasone. LIGHT-mediated up-regulation of MMPs and intercellular adhesion molecule 1 was blocked by inhibitors of NF-kappaB and JNK, whereas up-regulation of vascular cell adhesion molecule 1 was blocked by inhibitors of phosphatidylinositol 3-kinase, as well as NF-kappaB. These data suggest that binding of LIGHT with its receptors may play a role in the progression of inflammation within rheumatoid synovium, especially by mediating the interactions between infiltrating inflammatory cells and stromal cells. These findings thus emphasize the relevance of LIGHT as a potential therapeutic target in RA.

The functional variant of the inhibitory Fcγ receptor IIb (CD32B) is associated with the rate of radiologic joint damage and dendritic cell function in rheumatoid arthritis

Fcgamma receptors (FcgammaRs) recognize immune complexes (ICs) and coordinate the immune response by modulating the functions of dendritic cells (DCs). The purpose of this study was to unravel the role of the inhibitory FcgammaRIIb in rheumatoid arthritis (RA) by studying the effect of the FCGR2B 695T>C polymorphism on susceptibility to RA, severity of the disease, and DC function. Genotyping was performed in RA patients (n = 246) and healthy blood donors (n = 269). The patients' demographic data, disease severity, and disease progression were assessed over a followup of 6 years. DCs were cultured for flow cytometry to determine the expression of FcgammaRs. For detection of FcgammaRIIb (CD32B), a unique anti-FcgammaRIIb antibody (2B6-fluorescein isothiocyanate [FITC]) was used. The capacity for antigen uptake by DCs was studied by assessing the uptake of FITC-labeled ICs. Levels of cytokine production by DCs were measured during lipopolysaccharide-mediated cell activation in the presence and absence of ICs. Although no role of the FCGR2B variant in RA susceptibility was demonstrated, this variant was associated with a nearly doubled rate of radiologic joint damage during the first 6 years of RA. Multiple regression analysis showed that FCGR2B was by far the strongest predictor of joint damage identified to date. DCs from patients carrying this variant failed to display the inhibitory phenotype normally observed upon IC-mediated triggering of inflammation and displayed diminished FcgammaRII-mediated antigen uptake compared with wild-type DCs. However, the levels of FcgammaRs were not affected, suggesting that the FCGR2B variant alters the function rather than regulation of proteins. This study is the first to show that a single genetic variant, the FCGR2B 695T>C polymorphism, is a critical determinant of disease severity in RA and radically changes DC behavior. Our results underscore the key role of DCs in the progression of RA and reveal FcgammaRIIb as an important potential therapeutic target in RA and other autoimmune conditions.

Interleukin-18 induces the production of vascular endothelial growth factor (VEGF) in rheumatoid arthritis synovial fibroblasts via AP-1-dependent pathways

Interleukin-18 (IL-18) is a novel pro-inflammatory cytokine which has been implicated to play a pathogenic role in rheumatoid arthritis (RA). Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis in rheumatoid synoviocytes. In present study, we examined the effect of IL-18 on VEGF production in fibroblast-like synoviocytes (FLS) isolated from the patients with RA. FLS were prepared from the synovial tissues of patients with RA and osteoarthritis (OA) and cultured in the presence of IL-18. The production of VEGF from FLS was measured in culture supernatants by enzyme-linked immunosorbent assay (ELISA). The VEGF messenger RNA (mRNA) expression and AP-1 binding activity of VEGF transcript were determined by reverse transcription-polymerase chain reaction (RT-PCR) and electrophoretic mobility shift assay (EMSA). IL-18 and VEGF levels of sera and synovial fluids (SF) of RA patients ( n = 30) were significantly higher than those of OA patients ( n = 20). IL-18 dose-dependently increased the production of VEGF. The effect of IL-18 on VEGF production appeared to be as potent as IL-1β, whereas tumor necrosis factor (TNF)-α and interferon (IFN)-γ showed little effects on VEGF production. AP-1-specific inhibitor Curcumin dose-dependently abrogated the effect of IL-18 on VEGF production. The VEGF enhancement of IL-18 was associated with increased AP-1 binding activity to the VEGF promoter site. These findings suggest IL-18 as an angiogenic factor in RA and down-regulation of IL-18 activity or AP-1 signal pathway can be potential therapeutic targets for RA.