Plasmin immunization preferentially induces potentially prothrombotic IgG anticardiolipin antibodies in MRL/MpJ mice

Objective

To test the hypothesis, utilizing 2 experimental mouse models, that plasmin is an important autoantigen that drives the production of certain IgG anticardiolipin (aCL) antibodies in patients with the antiphospholipid syndrome.

Methods

BALB/cJ and MRL/MpJ mice were immunized with Freund's complete adjuvant in the presence or absence of human plasmin. The mouse sera were analyzed for production of IgG antiplasmin, IgG aCL, and IgG anti–β₂‐glycoprotein I (anti‐β₂GPI) antibodies. IgG monoclonal antibodies (mAb) were generated from the plasmin‐immunized MRL/MpJ mice with high titers of aCL, and these 10 mAb were studied for their binding properties and functional activity in vitro.

Results

Plasmin‐immunized BALB/cJ mice produced high titers of IgG antiplasmin only, while plasmin‐immunized MRL/MpJ mice produced high titers of IgG antiplasmin, IgG aCL, and IgG anti‐β₂GPI. Both strains of mice immunized with the adjuvant alone did not develop IgG antiplasmin or IgG aCL. All 10 of the IgG mAb bound to human plasmin and cardiolipin, while 4 of 10 bound to β₂GPI, 3 of 10 bound to thrombin, and 4 of 10 bound to the activated coagulation factor X (FXa). Functionally, 4 of the 10 IgG mAb inhibited plasmin activity, 1 of 10 hindered inactivation of thrombin by antithrombin III, and 2 of 10 inhibited inactivation of FXa by antithrombin III.

Conclusion

Plasmin immunization leads to production of IgG antiplasmin, aCL, and anti‐β₂GPI in MRL/MpJ mice, but leads to production of only IgG antiplasmin in BALB/cJ mice. IgG mAb generated from plasmin‐immunized MRL/MpJ mice bind to various antigens and exhibit procoagulant activity in vitro. These results suggest that plasmin may drive potentially prothrombotic aCL in genetically susceptible individuals.

     

Interleukin‐17 receptor deficiency results in impaired synovial expression of interleukin‐1 and matrix metalloproteinases 3, 9, and 13 and prevents cartilage destruction during chronic reactivated streptococcal cell wall–induced arthritis

Objective

To examine the role of interleukin‐17 receptor (IL‐17R) signaling in cartilage destruction and its interrelationship with synovial IL‐1 expression during chronic reactivated streptococcal cell wall (SCW)–induced arthritis.

Methods

SCW arthritis was repeatedly induced in wild‐type (WT) and IL‐17R–deficient (IL‐17R^–/–) mice. At different time points, joint inflammation was assessed by using calipers to measure joint swelling. On day 42, mice were killed, and knee joints were removed for histologic analysis. Quantitative polymerase chain reaction (PCR) analyses for different proinflammatory mediators and matrix metalloproteinases (MMPs) were performed on inflamed synovium from WT and IL‐17R^–/– mice after 5 repeated injections of SCW fragments.

Results

IL‐17R signaling did not play a significant role in acute joint swelling induced by a single injection of SCW fragments directly into the joint. However, repeated local injections of SCW fragments into the knee joints of IL‐17R^–/– mice resulted in fewer infiltrating cells in the joint compared with WT mice. Moreover, histologic analysis on day 42 revealed a significant suppression of the degree of chondrocyte death and an absence of cartilage surface erosion in IL‐17R^–/– mice. Quantitative PCR analysis revealed impaired synovial expression of IL‐1, IL‐6, cyclooxygenase 2, stromelysin (MMP‐3), gelatinase B (MMP‐9), and collagenase 3 (MMP‐13) in IL‐17R^–/– mice.

Conclusion

These data show a critical role of IL‐17R signaling in driving the synovial expression of proinflammatory and catabolic mediators, such as IL‐1 and different MMPs, during progression from an acute, macrophage‐driven joint inflammation to a chronic, cartilage‐destructive, T cell–mediated synovitis. Prevention of IL‐17R signaling warrants consideration as a therapeutic target in chronic destructive arthritis.

     

Inhibition of interleukin‐33 signaling attenuates the severity of experimental arthritis

Objective

Interleukin‐33 (IL‐33; or, IL‐1F11) was recently identified as the ligand of the IL‐1 family receptor T1/ST2. The aim of this study was to examine IL‐33 production in human and mouse joints and to investigate the role of IL‐33 and T1/ST2 in experimental arthritis.

Methods

IL‐33 expression was examined in human synovial tissue, rheumatoid arthritis (RA) synovial fibroblasts, and arthritic mouse joints. Mice with collagen‐induced arthritis (CIA) were treated with blocking anti‐ST2 antibody or control antibody beginning at the onset of disease. Arthritis severity was assessed by clinical and histologic scoring. Draining lymph node (LN) cell responses were examined ex vivo, and joint messenger RNA (mRNA) was used for expression profiling.

Results

IL‐33 was highly expressed in human RA synovium. In cultured synovial fibroblasts, IL‐33 expression was strongly induced by IL‐1β and/or tumor necrosis factor α. Furthermore, IL‐33 mRNA was detected in the joints of mice with CIA and increased during the early phase of the disease. Administration of a blocking anti‐ST2 antibody at the onset of disease attenuated the severity of CIA and reduced joint destruction. Anti‐ST2 antibody treatment was associated with a marked decrease in interferon‐γ production as well as with a more limited reduction in IL‐17 production by ex vivo–stimulated draining LN cells. Finally, RANKL mRNA levels in the joint were reduced by anti‐ST2 treatment.

Conclusion

IL‐33 is produced locally in inflamed joints, and neutralization of IL‐33 signaling has a therapeutic effect on the course of arthritis. These observations suggest that locally produced IL‐33 may contribute to the pathogenesis of joint inflammation and destruction.

     

Regulation of angiogenesis by the C‐X‐C chemokines interleukin‐8 and epithelial neutrophil activating peptide 78 in the rheumatoid joint

Objective

Angiogenesis, the growth of new blood vessels, is vital to the ingress of inflammatory leukocytes in rheumatoid arthritis (RA) synovial tissue and to the growth and proliferation of RA pannus. The factors that mediate the growth of new blood vessels have not been completely defined. This study examined the ability of Glu‐Leu‐Arg (ELR)–containing chemokines to induce angiogenesis in the RA joint.

Methods

To reflect angiogenic activity in vivo, we selected a model using whole human synovial tissue rather than isolated cells. Tissues were examined by immunohistochemistry and enzyme‐linked immunosorbent assay, and tissue homogenates were immunoneutralized and assayed for their ability to induce endothelial cell chemotaxis and rat corneal neovascularization.

Results

Cells expressing interleukin‐8 (IL‐8) and epithelial neutrophil activating peptide 78 (ENA‐78) were located in proximity to factor VIII–related antigen–immunopositive endothelial cells. RA homogenates produced more IL‐8 and ENA‐78 compared with normal synovial tissue homogenates. Moreover, homogenates from RA synovial tissue produced significantly more chemotactic activity for endothelial cells in vitro and angiogenic activity in the rat cornea in vivo than did normal synovial tissue homogenates. The effects of IL‐8 and ENA‐78 accounted for a significant proportion of the chemotactic activity of endothelial cells and angiogenic activity found in RA synovial tissue homogenates.

Conclusion

These results indicate that the ELR‐containing chemokines IL‐8 and ENA‐78 are important contributors to the angiogenic activity found in the inflamed RA joint. It is possible that efforts aimed at down‐regulating these chemokines offer a novel targeted therapy for the treatment of RA.

     

Elevated expression of interleukin‐7 receptor in inflamed joints mediates interleukin‐7–induced immune activation in rheumatoid arthritis

Objective

To evaluate the expression and functional ability of the high‐affinity interleukin‐7 receptor (IL‐7Rα) in patients with rheumatoid arthritis (RA).

Methods

Expression of IL‐7Rα and IL‐7 was determined in synovial tissue from RA patients and was compared with that in synovial tissue from patients with undifferentiated arthritis (UA) and osteoarthritis (OA). IL‐7Rα expression on CD4 T cells, CD19 B cells, and CD14 monocyte/macrophages from RA synovial tissue, synovial fluid, and peripheral blood was also assessed. The proliferative capacity of IL‐7Rα^bright and IL‐7Rα^dim/− T cells was measured. In addition, we examined IL‐7R blockade with soluble human IL‐7Rα (hIL‐7Rα) in the prevention of immune activation of peripheral blood mononuclear cells.

Results

We found significantly higher IL‐7Rα expression in RA and UA synovial tissue than in OA synovial tissue, and the level of IL‐7Rα expression correlated significantly with the levels of CD3 and IL‐7 expression. CD4 T cells from RA synovial fluid and synovial tissue strongly expressed IL‐7Rα. A substantial percentage of B cells and macrophages from RA synovial fluid and synovial tissue also expressed IL‐7Rα, although less prominently than T cells. We found that peripheral blood IL‐7Rα^bright T cells that did not express FoxP3 were highly proliferative as compared with IL‐7Rα^dim/− T cells that did express high levels of FoxP3. Soluble hIL‐7Rα inhibited IL‐7–induced proliferation and interferon‐γ production by mononuclear cells from RA patients.

Conclusion

Our data suggest that enhanced expression of IL‐7Rα and IL‐7 in RA patients contributes significantly to the joint inflammation by activating T cells, B cells, and macrophages. The inhibition of IL‐7R–mediated immune activation by soluble hIL‐7Rα further indicates an important role of IL‐7Rα in inflammatory responses in RA, suggesting IL‐7Rα as a therapeutic target for immunotherapy in RA.

     

Increased intraarticular interleukin‐7 in rheumatoid arthritis patients stimulates cell contact–dependent activation of CD4+ T cells and macrophages

Objective

To determine the level of intraarticular expression of interleukin‐7 (IL‐7) in patients with rheumatoid arthritis (RA) and to investigate the mechanisms by which IL‐7 facilitates activation of CD4+ T cells and monocyte/macrophages in RA.

Methods

IL‐7 levels were measured in synovial fluid obtained from patients with RA and patients with osteoarthritis (OA). Immunohistologic analysis was used to assess the expression of IL‐7 in synovial tissue from patients with RA. Proliferation and activation markers were determined in order to measure the effect of IL‐7 on mononuclear cells, isolated CD4+ T cells, and monocyte/macrophages from the peripheral blood and synovial fluid. Cocultures of CD4+ T cells and monocytic cells in the absence or presence of a semipermeable membrane were performed to assess the extent to which IL‐7 induces its effects, either contact dependently or via soluble mediators.

Results

IL‐7 levels were increased in synovial fluid from patients with RA compared with the levels in synovial fluid from patients with OA. Macrophages, fibroblasts, and endothelial cells in the joint lining tissue expressed abundant IL‐7. In vitro, synovial fluid CD4+ T cells and macrophages were hyperresponsive to IL‐7 when compared with peripheral blood cells. Furthermore, IL‐7 enhanced cell contact–dependent activation of CD4+ T cells and monocyte/macrophages.

Conclusion

The abundant intraarticular expression of IL‐7 and the stimulation by IL‐7 of contact‐dependent activation of CD4+ T cells and monocytic cells indicate that this cytokine plays an important proinflammatory role in RA synovitis. Further identification of IL‐7–induced pathways may improve understanding of the important interactive role of CD4+ T cells and monocytic cells in RA.

     

Diet‐induced obesity significantly increases the severity of posttraumatic arthritis in mice

Objective

Obesity and joint injury are primary risk factors for osteoarthritis (OA) that involve potential alterations in the biomechanical and inflammatory environments of the joint. Posttraumatic arthritis is a frequent long‐term complication of intraarticular fractures. Obesity has been linked to primary OA and may potentially contribute to the development of posttraumatic arthritis by a variety of mechanisms. The objectives of this study were to determine whether diet‐induced obesity influences the severity of posttraumatic arthritis in mice and to examine the interrelationships between joint degeneration and serum levels of the inflammatory cytokines and adipokines that are involved in this response.

Methods

C57BL/6 mice were fed either normal chow (13% fat) or a high‐fat diet (60% fat) starting at 4 weeks of age. At 16 weeks of age, half of the mice in each group were subjected to a closed intraarticular fracture of the left knee. At 8 weeks postfracture, knee OA was assessed by cartilage and synovium histology in addition to bone morphology. Serum cytokine concentrations were determined with multiplex assays.

Results

Fractured knee joints of mice receiving a high‐fat diet showed significantly increased OA degeneration compared with nonfractured contralateral control knees, while fractured knee joints of mice receiving a low‐fat diet did not demonstrate significant differences from nonfractured contralateral control knees. A high‐fat diet increased serum concentrations of interleukin‐12p70 (IL‐12p70), IL‐6, and keratinocyte‐derived chemokine while decreasing adiponectin concentrations. Joint injury also increased IL‐12p70 concentrations in mice receiving a high‐fat diet. Systemic levels of adiponectin were inversely correlated with synovial inflammation in control limbs.

Conclusion

Diet‐induced obesity significantly increased the severity of OA following intraarticular fracture. Obesity and joint injury together can alter systemic levels of inflammatory cytokines such as IL‐12p70.

     

Inhibition of interleukin‐6 receptor directly blocks osteoclast formation in vitro and in vivo

Objective

To investigate the efficacy of a murine anti–interleukin‐6 receptor (anti–IL‐6R) antibody in directly blocking tumor necrosis factor (TNF)– and RANKL‐mediated osteoclastogenesis in vitro and in vivo.

Methods

The efficacy of a murine antibody against IL‐6R in blocking osteoclast differentiation of mononuclear cells stimulated with RANKL was tested. In addition, arthritic human TNFα–transgenic mice were treated with anti–IL‐6R antibody, and osteoclast formation and bone erosion were assessed in arthritic paws.

Results

Blockade of IL‐6R dose dependently reduced osteoclast differentiation and bone resorption in monocyte cultures stimulated with RANKL or RANKL plus TNF. In human TNFα–transgenic mice, IL‐6R blockade did not inhibit joint inflammation, but it strongly reduced osteoclast formation in inflamed joints as well as bone erosion in vivo. Neither the cell influx into joints nor the synovial expression of IL‐6 and RANKL changed with IL‐6R blockade, while the synovial expression of IL‐1 was significantly reduced. In contrast, TNF‐mediated systemic bone loss was not inhibited by IL‐6R blockade.

Conclusion

These data suggest that blockade of IL‐6R directly affects osteoclast formation in vitro and in vivo, suggesting a direct and specific effect of anti–IL‐6R therapy on osteoclasts independently of its antiinflammatory effects. This effect adds significantly to the structure‐sparing potential of pharmacologic blockade of IL‐6R in arthritis.

     

Endothelial nitric oxide synthase inhibits the development of autoimmune‐mediated vasculitis in mice

Objective

Many different genes or mediators have been implicated in promoting the development of vasculitis, although little is known regarding the mechanisms that normally act to suppress lesion formation. Endothelial nitric oxide synthase (eNOS) has been shown to inhibit vascular inflammation in many different model systems, but its roles in the pathogenesis of vasculitis have not been elucidated. This study was undertaken to determine the functions of eNOS in the initiation and progression of vasculitic lesion formation.

Methods

MRL/MpJ‐Fas^lpr mice lacking the gene for eNOS (Nos3^−/−) were generated and comprehensively evaluated and compared to controls with regard to the development of autoimmune disease, including vasculitic lesion formation and glomerulonephritis.

Results

Nos3^−/− MRL/MpJ‐Fas^lpr mice exhibited accelerated onset and increased incidence of renal vasculitis compared to Nos3^+/+ controls. In contrast, no significant differences in severity of glomerulonephritis were observed between groups. Vasculitis was also observed in other organs of eNOS‐deficient mice, including in the lungs of several of these animals. Ultrastructural analyses of renal lesions revealed the presence of electron‐dense deposits in affected arteries, and IgG, IgA, and C3 deposition was observed in some vessels in the kidneys of Nos3^−/− mice. In addition, Nos3^−/− MRL/MpJ‐Fas^lp mice showed increased levels of circulating IgG–IgA immune complexes at 20 weeks of age, compared to Nos3^+/+ MRL/MpJ‐Fas^lpr and Nos3^−/− C57BL/6 mice.

Conclusion

These findings strongly indicate that eNOS serves as a negative regulator of vasculitis in MRL/MpJ‐Fas^lpr mice and further suggest that NO produced by this enzyme may be critical for inhibiting lesion formation and vascular damage in human vasculitic diseases.

     

Interleukin‐15 inhibits sodium nitroprusside–induced apoptosis of synovial fibroblasts and vascular endothelial cells

Objective

One of the pathologic hallmarks of juvenile rheumatoid arthritis (JRA) is a tumor‐like expansion of inflamed synovial tissue, or pannus, which causes much of the joint damage in this disease. The expansion of pannus is supported by extensive formation of new blood vessels. We have previously shown that revascularization of minced JRA synovial tissues engrafted into SCID mice correlated with the intensity of inflammatory activity in the tissues and with interleukin‐15 (IL‐15) expression. Since synovial vascular endothelial cells (VECs) expressed IL‐15 receptors, the present study was undertaken to investigate the hypothesis that IL‐15 might play a role in neovascularization of the pannus.

Methods

To evaluate IL‐15 for possible angiogenic activity, we assessed the ability of recombinant human IL‐15 (rHuIL‐15) to induce VEC growth directly and to stimulate synovial cells to produce endothelial growth factors. Since IL‐15 had been shown to inhibit apoptosis of certain immune cells, we were also interested in whether it might have similar effects on VECs. Apoptosis was induced by addition of sodium nitroprusside (SNP) at 1–2 mM to >80% confluent primary VECs, and numbers of apoptotic cells were determined by annexin V assay.

Results

Addition of rHuIL‐15 at 10–100 ng/ml to primary synovial fibroblast cultures failed to up‐regulate expression of vascular endothelial growth factor and angiopoietin 1 by these cells. Although rHuIL‐15 failed to induce a mitogenic response of VECs, it promoted survival of these cells on Matrigel. Preincubation of VECs with rHuIL‐15 at 50 ng/ml significantly reduced the proportion of VECs undergoing apoptosis.

Conclusion

IL‐15 promotes survival of VECs on Matrigel and inhibits SNP‐induced apoptosis of endothelial cells. We hypothesize that this mechanism may be relevant to the stabilization of newly formed vascular structures in JRA synovium.

     

Tumor necrosis factor–interleukin‐17 interplay induces S100A8, interleukin‐1β, and matrix metalloproteinases,and drives irreversible cartilage destruction in murine arthritis: Rationale for combination treatment during arthritis

Objective

To examine whether synovial interleukin‐17 (IL‐17) expression promotes tumor necrosis factor (TNF)–induced joint pathologic processes in vivo, and to analyze the surplus ameliorative value of neutralizing IL‐17 in addition to TNF during collagen‐induced arthritis (CIA).

Methods

Adenoviral vectors were used to induce overexpression of IL‐17 and/or TNF in murine knee joints. In addition, mice with CIA were treated, at different stages of arthritis, with soluble IL‐17 receptor (sIL‐17R), TNF binding protein (TNFBP), or the combination.

Results

Overexpression of IL‐17 and TNF resulted in joint inflammation and bone erosion in murine knees. Interestingly, IL‐17 strikingly enhanced both the joint‐inflammatory and joint‐destructive capacity of TNF. Further analysis revealed a strongly enhanced up‐regulation of S100A8, IL‐1β, and matrix metalloproteinase (MMP) messenger RNA, only when both TNF and IL‐17 were present. Moreover, the increase in irreversible cartilage destruction was not merely the result of enhanced inflammation, but also was associated with a direct synergistic effect of these cytokines in the joint. S100A9 deficiency in mice protected against IL‐17/TNF–induced expression of cartilage NITEGE neoepitopes. During established arthritis, the combination of sIL‐17R and TNFBP was more effective than the anticytokine treatments alone, and significantly inhibited further joint inflammation and cartilage destruction.

Conclusion

Local synovial IL‐17 expression enhances the role of TNF in joint destruction. Synergy between TNF and IL‐17 in vivo results in striking exaggeration of cartilage erosion, in parallel with a synergistic up‐regulation of S100A8, IL‐1β, and erosive MMPs. Moreover, neutralizing IL‐17 in addition to TNF further improves protection against joint damage and is still effective during late‐stage CIA. Therefore, compared with anti‐TNF alone, combination blocking of TNF and IL‐17 may have additional therapeutic value for the treatment of destructive arthritis.

     

Exacerbation of antigen‐induced arthritis in inducible nitric oxide synthase–deficient mice

Objective

Inhibition of nitric oxide (NO) produced by inducible NO synthase (iNOS) is suggested to be beneficial in experimental arthritis. Although NO is important for the integrity of the microcirculation, the effects of inhibition of iNOS on the synovial microcirculation are not currently known. This study investigated the synovial microcirculation and leukocyte–endothelial cell interactions in iNOS‐deficient mice with antigen‐induced arthritis (AIA) and compared these findings with disease severity.

Methods

Fourteen homozygous iNOS−/− and 14 iNOS+/+ mice were used. The severity of AIA was assessed by measuring knee joint swelling and by histologic scoring. The number of rolling and adherent leukocytes was quantitatively analyzed in synovial microvessels using intravital microscopy of intraarticular synovial tissue. Nitrite/nitrate concentrations were measured, and the expression of iNOS, E‐ and P‐selectin, intercellular adhesion molecule 1, and vascular cell adhesion molecule 1 (VCAM‐1) was assessed by immunohistochemistry.

Results

In iNOS+/+ animals with AIA, the plasma concentration of nitrite/nitrate was increased 3‐fold and iNOS expression was detected in cells of the joint. Swelling of the knee joint as well as leukocyte infiltration were enhanced in the iNOS−/− arthritic animals compared with iNOS+/+ mice with AIA. AIA‐associated leukocyte–endothelial cell interaction in synovial postcapillary venules was more pronounced in iNOS−/−, compared with iNOS+/+, arthritic mice. A strong expression of P‐selectin and VCAM‐1 was observed in the iNOS−/− arthritic mice only.

Conclusion

These data suggest that NO production by iNOS in vivo has antiinflammatory effects in experimental arthritis, by mediating a reduction in leukocyte adhesion and infiltration.

     

Leptin exacerbates collagen‐induced arthritis via enhancement of Th17 cell response

Objective

To determine the role of leptin in modulating Th17 cell response and joint inflammation in mice with collagen‐induced arthritis (CIA).

Methods

Leptin receptor expression on T cells was examined by polymerase chain reaction (PCR) analysis, immunofluorescence microscopy, and flow cytometry. Effects of leptin on Th17 cell differentiation and proliferation were evaluated by quantitative PCR, carboxyfluorescein diacetate succinimidyl ester proliferation assay, and flow cytometry. Dynamic changes in leptin concentrations in the joint tissue and synovial fluid of mice with CIA were determined by immunohistochemistry analysis and enzyme‐linked immunosorbent assay (ELISA). Arthritis symptoms and joint pathology in mice with CIA were assessed after injection of leptin into the knee joint. Th1 and Th17 cell populations in the spleen, draining lymph nodes, and joint tissue were analyzed by flow cytometry and enzyme‐linked immunospot assay. Interleukin‐17 messenger RNA and protein levels in the joint tissue were measured by PCR analysis and ELISA.

Results

In culture, leptin treatment significantly increased Th17 cell generation from naive CD4+ T cells. During CIA development, markedly elevated levels of leptin were detected in the joint tissue and synovial fluid. Moreover, injection of leptin into the knee joint of collagen‐immunized mice resulted in an early onset of arthritis and substantially increased the severity of clinical symptoms, accompanied by more pronounced synovial hyperplasia and joint damage. Further examination by immunofluorescence microscopy confirmed the presence of significantly increased numbers of Th17 cells in the joint tissue and draining lymph nodes of leptin‐treated mice with CIA.

Conclusion

The results of this study identify a previously undescribed function of leptin in enhancing Th17 cell response and exacerbating joint inflammation in mice with CIA.

     

Interleukin‐17–producing T cells are enriched in the joints of children with arthritis,but have a reciprocal relationship to regulatory T cell numbers

Objective

To identify interleukin‐17 (IL‐17)–producing T cells from patients with juvenile idiopathic arthritis (JIA), and investigate their cytokine production, migratory capacity, and relationship to Treg cells at sites of inflammation, as well as to test the hypothesis that IL‐17+ T cell numbers correlate with clinical phenotype in childhood arthritis.

Methods

Flow cytometry was used to analyze the phenotype, cytokine production, and chemokine receptor expression of IL‐17–producing T cells in peripheral blood and synovial fluid mononuclear cells from 36 children with JIA, in parallel with analysis of forkhead box P3 (FoxP3)–positive Treg cells. Migration of IL‐17+ T cells toward CCL20 was assessed by a Transwell assay. Synovial tissue was analyzed by immunohistochemistry for IL‐17 and IL‐22.

Results

IL‐17+ T cells were enriched in the joints of children with JIA as compared with the blood of JIA patients (P = 0.0001) and controls (P = 0.018) and were demonstrated in synovial tissue. IL‐17+ T cell numbers were higher in patients with extended oligoarthritis, the more severe subtype of JIA, as compared with patients with persistent oligoarthritis, the milder subtype (P = 0.046). Within the joint, there was an inverse relationship between IL‐17+ T cells and FoxP3+ Treg cells (r = 0.61, P = 0.016). IL‐17+,CD4+ T cells were uniformly CCR6+ and migrated toward CCL20, but synovial IL‐17+ T cells had variable CCR4 expression. A proportion of IL‐17+ synovial T cells produced IL‐22 and interferon‐γ.

Conclusion

This study is the first to define the frequency and characteristics of “Th17” cells in JIA. We suggest that these highly proinflammatory cells contribute to joint pathology, as indicated by relationships with clinical phenotypes, and that the balance between IL‐17+ T cells and Treg cells may be critical to outcome.

     

The response of T cells to interleukin‐6 is differentially regulated by the microenvironment of the rheumatoid synovial fluid and tissue

Objective

Interleukin‐6 (IL‐6) is a proinflammatory cytokine with regulatory effects on the survival and differentiation of T cells. It exerts its biologic function in 2 ways: by directly binding to the IL‐6 receptor (IL‐6R; CD126) or via trans‐signaling, in which soluble IL‐6R/IL‐6 complexes bind to the signaling component CD130. This study was undertaken to assess the expression and regulation of CD126 and CD130 and determine how these affect the response of CD4+ T cells to IL‐6 in the joints of patients with rheumatoid arthritis (RA).

Methods

Flow cytometry and immunofluorescence microscopy were used to determine the expression, function, and regulation of CD126 and CD130 in CD4+ T cells from the peripheral blood (PB), synovial fluid (SF), and synovial tissue of RA patients.

Results

Compared to the findings in RA PB, CD4+ T cells in the SF and synovial tissue expressed low levels of CD126. In contrast, whereas CD4+ T cell expression of CD130 was minimal in the SF, its level in the synovial tissue was high. Consistent with this phenotype, synovial tissue T cells responded to trans‐signaling by soluble IL‐6R/IL‐6 complexes, whereas no response was evident in CD4+ T cells from the SF. Down‐regulation of both receptor components in SF T cells could be explained by exposure to high levels of IL‐6. Increased levels of CD130 messenger RNA and protein in synovial tissue CD4+ T cells suggested that CD130 is up‐regulated locally. Among a range of cytokines tested, only IL‐10 induced CD130 expression in T cells.

Conclusion

The inflamed microenvironment in the synovial tissue maintains responsiveness to IL‐6 trans‐signaling through the up‐regulation of CD130 expression in CD4+ T cells, and this process may be driven by IL‐10.