Yojiro Kawabe

Primary Sjögren's syndrome with antibodies to HTLV-I: clinical and laboratory features.

The prevalence of antibodies to human T lymphotropic virus type I (HTLV-I) was studied in patients with primary Sjögrens syndrome. Thirteen of 36 serum samples were positive by enzyme linked immunosorbent assay (ELISA) and particle agglutination assay for antibodies to HTLV-I and were confirmed by western blotting. The presence of antibodies to HTLV-I may signify an HTLV-I carrier state. These patients had a high occurrence of extraglandular manifestations such as uveitis, myopathy, and recurrent high fever compared with patients who did not have antibodies to HTLV-I. Patients with antibodies to HTLV-I had an increased spontaneous proliferation of peripheral blood mononuclear cells compared with those without the antibodies. The proportions of activated and memory T cells (HLA-DR+ CD3+, CD25+ CD3+, and CD29+ CD4+ cells) were higher in HTLV-I carriers than in non-carriers. The presence of antibodies to HTLV-I in some patients with primary Sjögrens syndrome suggests that HTLV-I may cause primary Sjögrens syndrome or its extraglandular manifestations, or both.

Importance of NF-κB in rheumatoid synovial tissues: in situ NF-κB expression and in vitro study using cultured synovial cells

OBJECTIVES To examine whether inhibition of NF-κB induces apoptosis of human synovial cells stimulated by tumour necrosis factor α (TNFα), interleukin 1β (IL1β), and anti-Fas monoclonal antibody (mAb). METHODS The expression of proliferating cell nuclear antigen (PCNA), NF-κB, and the presence of apoptotic synovial cells were determined in synovial tissues. Apoptosis of cultured synovial cells was induced by inhibition of NF-κB nuclear translocation by Z-Leu-Leu-Leu-aldehyde (LLL-CHO). The activation of caspase-3 and expression of XIAP and cIAP2 in synovial cells in LLL-CHO induced apoptosis was also examined. RESULTS Abundant PCNA+ synovial cells were found in rheumatoid arthritis (RA) synovial tissue, though a few apoptotic synovial cells were also detected in the RA synovial tissues. Nuclear NF-κB was expressed in RA synovial cells. Electrophoretic mobility shift assay showed that treatment of cells with TNFα or IL1β significantly stimulated nuclear NF-κB activity. A small number of apoptotic synovial cells expressing intracellular active caspase-3 were found after treatment of cells with LLL-CHO. Although treatment of RA synovial cells with TNFα or IL1β alone did not induce apoptosis, apoptosis induced by LLL-CHO and caspase-3 activation were clearly enhanced in TNFα or IL1β stimulated synovial cells compared with unstimulated synovial cells. Furthermore, induction of apoptosis of synovial cells with caspase-3 activation by anti-Fas mAb was clearly increased by LLL-CHO. The expression of cIAP2 and XIAP in synovial cells may not directly influence the sensitivity of synovial cells to apoptosis induced by LLL-CHO. CONCLUSION The results suggest that NF-κB inhibition may be a potentially important therapeutic approach for RA by correcting the imbalance between apoptosis and proliferation of synovial cells in RA synovial tissue.

Expression of basic fibroblast growth factor in synovial tissues from patients with rheumatoid arthritis: detection by immunohistological staining and in situ hybridisation.

OBJECTIVE--The distribution and production of basic fibroblast growth factor (bFGF) was examined on the synovium from patients with rheumatoid arthritis (RA) and osteoarthritis (OA). METHODS--The localisation of bFGF was determined by an immunohistochemical staining procedure using anti-bFGF monoclonal antibody. The expression of bFGF mRNA was detected by nonradioactive in situ hybridisation using bFGF antisense oligo DNA. RESULTS--The bFGF was found in the synovial lining cell, sublining stromal fibroblast-like cells, and vascular endothelial cells from patients with RA and OA. Little or no bFGF was found in non-inflamed synovium. Immunostaining of bFGF in the synovial cells was more extensive and intense in synovium of patients with RA than that of patients with OA. The nuclei of the synovial lining cell layer were also immunostained. These nuclear staining were more intense in the lining cell layer from RA patients with moderate or severe proliferation of synovial cells than in RA patients with mild proliferation. The bFGF mRNA was also detected in the synovial lining cell layer of the inflamed synovium. CONCLUSION--The synovial lining cells produced bFGF. The proliferation of synovial cells in the inflamed joints may be the results of stimulation by the bFGF in autocrine manner.

Suppressive effect of leflunomide metabolite (A77 1726) on metalloproteinase production in IL-1β stimulated rheumatoid synovial fibroblasts

Leflunomide, an isoxazol derivative structurally unrelated to other immunomodulatory drugs, has proven to be efficacious in the treatment of rheumatoid arthritis (RA). This study was conducted to elucidate the mechanism by which leflunomide mediated antirheumatic effects. We investigated the effects of A77u20031726, leflunomides active metabolite, on mitogen‐activated protein kinase (MAPK) activation in IL‐1β‐stimulated rheumatoid synovial fibroblasts. The effects of A77u20031726 on the secretion of matrix metalloproteinases (MMPs) from rheumatoid synovial fibroblasts were also examined. A77u20031726 partially suppressed IL‐1β‐induced ERK1/2 and p38 kinase activation. In contrast, A77u20031726 efficiently suppressed IL‐1β‐stimulated JNK1/2 kinase activation. Although no suppressive effect was demonstrated on MMP‐2, A77u20031726 markedly inhibited MMP‐1, 3, and 13 secretions from IL‐1β‐stimulated rheumatoid synovial fibroblasts. Tissue inhibitor of metalloproteinases‐1 (TIMP‐1) was constitutively produced from rheumatoid synovial fibroblasts and the suppressive effects of A77u20031726 on TIMP‐1 production were minimal. Our results suggest that the suppression of the MAPK signalling pathway and MMP synthesis in rheumatoid synovial fibroblasts is a possible mechanism for the inhibitory activity of leflunomide against rheumatoid arthritis.

Apoptosis induction in synovial fibroblasts by ceramide: In vitro and in vivo effects

Several lipid second messengers are important mediators of extracellular signals. Among them ceramide, which is formed by cell membrane sphingomyelin, influences the apoptotic signal pathway through Fas antigen. We examined the apoptotic effect of cell-permeable C2-ceramide on rheumatoid synovial fibroblasts in vitro and in vivo. Exposure of cultured human rheumatoid synovial fibroblasts to C2-ceramide for 24 hours produced internucleosomal DNA fragmentation and morphologic changes characteristic of apoptosis. This C2-ceramide-mediated apoptosis was dose dependent as confirmed by analysis of cytosolic oligonucleosome-bound DNA of treated synovial fibroblasts. We also demonstrated that intra-articular administration of C2-ceramide into Fas-deficient MRL lpr/lpr mice produced a profound reduction of synovial hyperplasia within 24 hours. In situ nick end labeling analysis confirmed the induction of apoptosis in synovial lining cells. Our results indicate that C2-ceramide may function as a potent inducer of apoptosis in the synovium and suggest that pharmacologically-induced apoptosis may be useful as a new therapeutic modality for rheumatoid arthritis.

Increased levels of serum IgM antibody to staphylococcal enterotoxin B in patients with rheumatoid arthritis.

OBJECTIVE--To investigate the role of superantigen in rheumatoid arthritis (RA) by assaying the serum levels of staphylococcal enterotoxin B (SEB) antibodies. METHODS--Serum IgG and IgM SEB antibodies were measured using an enzyme linked immunosorbent assay (ELISA), and confirmed by Western blot analysis. The T cell receptor V beta (TCR V beta) repertoire was analysed using the reverse transcriptase polymerase chain reaction. RESULTS--RA patients had increased levels of serum IgM SEB antibody compared with normal subjects, patients with systemic lupus erythematosus, Sjögrens syndrome, and Behçets disease. The titres of rheumatoid factor (RF) showed no correlation with the levels of IgM SEB antibodies, and the levels of SEB antibodies were not inhibited by the addition of human immunoglobulin, or after absorption of RF. RA patients whose disease duration was less than 10 years had greater levels of serum IgM SEB antibodies than those with disease duration more than 10 years. The levels of IgM and IgG SEB antibodies in synovial fluid from RA patients were correlated with those in their sera. Western blot analysis detected IgM and IgG SEB antibodies as a band of approximately 30 kDa molecular size. The percentage of TCR V beta 2, V beta 5.2, and V beta 12 in phytohaemagglutinin stimulated peripheral T cells correlated significantly with the levels of serum IgM SEB antibody in RA patients. CONCLUSION--These results suggest that SEB, one of the superantigens, may have a critical role in the pathogenesis of RA.

Prediction of DAS28-ESR remission at 6 months by baseline variables in patients with rheumatoid arthritis treated with etanercept in Japanese population

We tried to determine which baseline variables are responsible for remission induction at 6xa0months in unselected rheumatoid arthritis (RA) patients of Japanese population treated with etanercept. One hundred forty-one patients with RA who were administered etanercept were registered. Thirty-four patients were started on etanercept monotherapy, 60 patients on cotherapy with methotrexate (MTX) (MTX cotherapy), and 47 patients on cotherapy with other non-MTX nonbiologic disease-modifying antirheumatic drugs (DMARDs) (non-MTX cotherapy). None of the patients were treated with both MTX and non-MTX nonbiologic DMARDs at entry. Outcome was set as achievement of disease activity scorexa028 (DAS28)-ESR remission at 6xa0months. We examined association of gender, DAS at baseline, MTX cotherapy at baseline, non-MTX cotherapy at baseline, and prednisolone use at baseline with achievement of remission at 6xa0months by logistic regression analysis. All subjects were classified as having high (Nxa0=xa0109) or moderate disease activity (Nxa0=xa032) at entry. One hundred twenty out of 141 patients (85.1%) continued treatment with etanercept at 6xa0months. Continuation rate was statistically higher in MTX cotherapy (93.3%) compared with etanercept monotherapy (73.5%), and tended to be higher than with non-MTX cotherapy (85.1%). Logistic regression analysis identified that MTX cotherapy at entry and moderate disease activity at entry were independent variables for remission induction at 6xa0months. Accordingly, DAS28-ESR at 6xa0months was significantly lower with MTX cotherapy as compared with etanercept monotherapy or non-MTX cotherapy. To a lesser extent, DAS28-ESR with non-MTX cotherapy at 6xa0months was lower than with etanercept monotherapy. In this study of unselected patients, use of MTX and moderate disease activity at entry were associated with higher likelihood of response to etanercept. Non-MTX nonbiologic DMARDs may be an alternative in RA patients administrated etanercept who are intolerant to MTX.

Cytokine production by endothelial cells infected with human T cell lymphotropic virus type I.

OBJECTIVE: To investigate the ability of human T cell lymphotropic virus type I (HTLV-I) to infect endothelial cells and induce cytokine production by these cells. METHODS: Human umbilical vein endothelial cells (HUVEC) were cocultured with HTLV-I infected T cell line (MT-2 cells) or uninfected T cell line (CEM cells). RESULTS: Following coculture with MT-2 cells, endothelial cells expressed HTLV-I specific core antigens. Endothelial cells cocultured with MT-2 cells produced significant amounts of several cytokines, including interleukin (IL)-1 alpha, IL-6, granulocyte colony stimulating factor (G-CSF), and granulocyte/macrophage colony stimulating factor (GM-CSF), compared with endothelial cells cocultured with CEM cells. Coculturing of endothelial cells with MT-2 and CEM cells failed to produce detectable amounts of IL-1 beta and tumour necrosis factor alpha (TNF-alpha). The production of cytokines by endothelial cells cocultured with MT-2 cells was more persistent than that by endothelial cells cocultured with CEM cells after several passages. Furthermore, the production was blocked by cocultivation of endothelial cells and MT-2 cells using the Millicell system. Finally, after cocultivation of endothelial cells and MT-2 cells, endothelial cells positive for HTLV-I antigen were stained by anti-GM-CSF antibody. CONCLUSIONS: HTLV-I can infect endothelial cells, resulting in their active production of several cytokines, such as IL-1 alpha, IL-6, G-CSF, and GM-CSF. These findings strongly suggest that the excess production of these cytokines by HTLV-I infected endothelial cells may be involved in the pathogenesis of HTLV-I associated inflammatory diseases.

Interleukin 4 increases human synovial cell expression of VCAM-1 and T cell binding.

OBJECTIVE--The effects were studied of interleukin 4 (IL-4) on T cell-synovial cell adhesion and on the expression of adhesion molecules on the surface of synovial fibroblast-like cells. METHODS--The adhesion of T cells toward the synovial cells were measured by 51chromium-labelled adhesion assay. The expression of adhesion molecules on synovial cells were analysed by flowcytometry. RESULTS--Stimulation of synovial cells with IL-4 increased T cell-synovial cells adhesion in a time- and dose-dependent manner. IL-4 considerably enhanced the expression of VCAM-1 on the surface of synovial cells, but not the expression of ICAM-1 and ELAM-1. The combination of IL-1 beta and IL-4 had no effect on the expression of ICAM-1 or VCAM-1 on the surface of synovial cells. The increased adhesion of T cells to IL-4 stimulated synovial cells was inhibited significantly by adding anti-VCAM-1 or anti-CD29 monoclonal antibody. Furthermore, anti-VLA-4 alpha or the combination of anti-VLA-4 alpha and anti-VCAM-1 antibodies blocked completely T-cell binding to IL-4 stimulated synovial cells. CONCLUSIONS--These results suggest that the increased adhesion of T cells to IL-4-stimulated synovial cells is mediated by VLA-4/VCAM-1 pathway.

SAT0533 Early Diagnosis is Associated with the Less Flair in Patients with Remitting Seronegative Symmetrical Synovitis with Pitting Edema (RS3PE) Syndrome

Background Remitting Seronegative Symmetrical Synovitis with Pitting Edema (RS3PE) syndrome is first described by McCarty et al. in 1985. We had previously analyzed the characteristics of RS3PE syndrome and found that the diagnosis is sometimes delayed and the flair is often found during the first year of follow-up1. Objectives To investigate whether the delay of diagnosis is associated with the flair in patients with RS3PE syndrome. Methods A retrospective rheumatology multicenter study has been conducted in Nagasaki prefecture, Japan, toward patients with RS3PE syndrome fulfilling the following criteria: (1) bilateral pitting edema of both hands or feet, or both, (2) sudden onset of polyarthritis, (3) age >50 years, (4) seronegative for rheumatoid factor. The patients, diagnosed as RS3PE syndrome, were consecutively enrolled between November 2005 and October 2014, and subjected to investigate the association of the delay of diagnosis with the flair during the first year of follow-up from medical records. The date of diagnosis was defined as the introduction of prednisolone and we have divided the 39 patients according to the duration from disease onset to the diagnosis: early diagnosis group less than 1 month whereas delayed diagnosis group required more than 1 month until diagnosis. The flair is defined as the increment of prednisolone dosages. Results Fifty-three patients fulfilled the above criteria of RS3PE syndrome. Of these, 14 patients were excluded owing to a lack of clinical data and 39 patients were included. They were 22 male (55.2%) and 17 female (44.8%) patients. The mean age was 78.9±6.3 years (65-89). Fourteen patients were classified as the early diagnosis group whereas the remaining 25 patients as the delayed diagnosis group, respectively. All the 39 patients showed an initial good response toward prednisolone, however, 12 patients (30.1%) experienced the flair during the first year. No significant differences were noted in C-reactive protein and erythrocyte sedimentation rate between the 2 groups. However, there was a significant difference in the rate of flair between the 2 groups since no flair in the early diagnosis group whereas 12 out of 25 patients (48.0%) in the delayed diagnosis group (p<0.001). Conclusions Our present data suggest that early diagnosis is associated with the less flair in patients with RS3PE syndrome. Physicians are required to recognize this rare but important inflammatory condition found in elderly subjects. References T. Origuchi, K. Arima, S.-Y. Kawashiri, M. Tamai, H. Nakamura, A. Kawakami, T. Tsukada, T. Miyashita, T. Aramaki, M. Furuyama, Y. Kawabe, N. Iwanaga, Y. Ueki, T. Fukuda, K. Eguchi: Surveillance of the Outcome of Remitting Seronegative Symmetrical Synovitis with Pitting Edema (RS3PE) Syndrome. Ann Rheum Dis 2014;73:Suppl 2 105 Disclosure of Interest None declared