RESUMO
OBJECTIVES: This study aimed to identify therapeutic predictors of abatacept (ABT) treatment in rheumatoid arthritis (RA) in vitro and in patients. METHODS: T cell cytokine, monokine, and chemokine levels in culture supernatants or serum were determined using flow cytometry bead-based immunoassays. CXCL10 mRNA and protein expressions were also assessed using qPCR and ELISA analyses, respectively. In the patient study, 25 ABT-treated patients were analysed retrospectively. The patients were divided into low disease activity (LDA) or non-low disease activity (non-LDA) groups at 24 weeks of ABT treatment. Seven T cell cytokines and CXCL10 levels were compared in these two groups. RESULTS: Peripheral blood mononuclear cells (PBMC) from healthy donors were stimulated by immobilised anti-CD3 with or without ABT for three days, and the levels of 13 T cell cytokines in culture supernatants were determined. ABT significantly inhibited anti-CD3-induced production of IFN-γ. To examine the effect of these T cell cytokines in rheumatoid synovial cells (RSC), RSCs were stimulated with 10% of culture supernatants from anti-CD3-stimulated PBMCs with or without ABT, and the levels of 23 cytokines were determined. Only CXCL10 was significantly reduced by ABT-treated supernatants. In the patient study, CXCL10 levels at baseline were not different between the LDA and non-LDA groups, whereas CXCL10 levels at 24 weeks were significantly decreased in the LDA group only. CONCLUSIONS: ABT treatment significantly affected IFN-γ and CXCL10 cytokine levels in vitro. In addition, serum CXCL10 levels were associated with better responses in ABT treatment.
Assuntos
Artrite Reumatoide , Leucócitos Mononucleares , Abatacepte/farmacologia , Artrite Reumatoide/tratamento farmacológico , Quimiocina CXCL10 , Quimiocinas , Humanos , Estudos RetrospectivosRESUMO
BACKGROUND/OBJECTIVES: Immunosuppressant medications (ISPs) increase the occurrence of Pneumocystis jirovecii pneumonia (PCP) in rheumatoid arthritis (RA) patients. The prophylactic administration of trimethoprim/sulfamethoxazole (TMP/SMX) for PCP is effective but has serious adverse effects and so should be selectively used for patients at high risk. The aims of this study were to clarify the risk factors for PCP in RA patients and to establish the indications for administering TMP/SMX. METHODS: This retrospective cohort study analyzed data from 2640 patients (2010-2014) diagnosed as having RA who had not received a prophylactic administration of TMP/SMX. The risk factors for PCP were evaluated by comparing the clinical parameters between patients with PCP (PCP group, n = 19) and those without (non-PCP group, n = 2621). RESULTS: The PCP group was older (70 vs. 64 years), received higher doses of prednisolone (6.2 vs. 2.4 mg/d) and methotrexate (7.7 vs. 5.2 mg/wk), and had a greater number of ISPs (1.3 vs. 0.8) (p < 0.05). We stratified the PCP risk using a scoring system based on odds ratios (ORs) calculated for these parameters (methotrexate ≥6 mg/wk OR = 4.5, 1 point; age ≥65 years, OR = 3.7, 1 point; ≥2 ISPs, OR = 3.7, 1 point; prednisolone ≥5 mg/d, OR = 12.4, 3 points). The incidence of PCP among patients scoring 0 to 2 points was 0.04%; 3 to 4 points, 2.3%; and 5 points or more, 5.8%. CONCLUSIONS: The prophylactic administration of TMP/SMX for PCP is recommended for RA patients who score at least 5 points with our system.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Imunossupressores/efeitos adversos , Pneumonia por Pneumocystis/imunologia , Pneumonia por Pneumocystis/prevenção & controle , Combinação Trimetoprima e Sulfametoxazol/uso terapêutico , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pneumocystis carinii , Estudos Retrospectivos , Fatores de Risco , Resultado do TratamentoRESUMO
OBJECTIVES: To investigate the duration of remission and low disease activity (LDA) after cessation of tocilizumab (TCZ) treatment in rheumatoid arthritis patients who showed remission or LDA as assessed by DAS28 in response to preceding TCZ monotherapy, and to explore the factors contributing to prolonged efficacy duration. METHODS: Disease activity was monitored for 56 weeks. The rate of continued efficacy was estimated by Kaplan-Meier curves. RESULTS: A total of 187 patients were eligible. At baseline of this study, median disease duration was 7.8 years, preceding TCZ treatment period was 4.0 years and DAS28 was 1.5. The rate of continued LDA at 52 weeks was 13.4 % according to the Kaplan-Meier estimate. 19 patients (10 %) were completely drug-free and 17 patients (9.1 %) fulfilled DAS28 remission at 52 weeks. Multivariate Cox regression analysis identified low serum IL-6 and normalisation of MMP-3 levels at cessation of TCZ as independent predictive markers for longer duration of LDA. In patients with low serum IL-6 (<12.9 pg/mL) and normal MMP-3 levels, the rate of continued LDA reached 38.0 % at 52 weeks. CONCLUSIONS: TCZ monotherapy may induce biologics-free remission or LDA without concomitant use of synthetic DMARDs. Serum levels of IL-6 and MMP-3 are useful markers for identifying patients who could discontinue TCZ without acute disease flare.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Adulto , Idoso , Artrite Reumatoide/sangue , Feminino , Humanos , Interleucina-6/sangue , Masculino , Metaloproteinase 3 da Matriz/sangue , Pessoa de Meia-Idade , Indução de Remissão/métodos , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
OBJECTIVES: To evaluate the safety and efficacy of retreatment with tocilizumab (TCZ) in patients who had participated in the DREAM study (Drug free remission/low disease activity after cessation of tocilizumab [Actemar] monotherapy study) and had experienced loss of efficacy. METHODS: Patients were retreated with TCZ or other disease modifying antirheumatic drugs (DMARDs). Disease activity was measured using the 28-joint disease activity score (DAS28) for 12 weeks. RESULTS: A total of 164 eligible patients, including 161 who experienced loss of efficacy within 52 weeks of the DREAM study, resumed treatment: 157 with TCZ and 7 with DMARDs and/or infliximab. Of TCZ-treated patients, 88.5 % (139 patients) achieved DAS28 <2.6 within 12 weeks, whereas among patients treated with DMARDs and/or infliximab only 14.3 % (1 patient) achieved DAS28 <2.6. Adverse events were observed in 70 TCZ-treated patients (44.0 %), but no serious infusion reactions were observed. CONCLUSIONS: Retreatment with TCZ was well-tolerated and effective in patients who had responded to the preceding TCZ monotherapy but had experienced loss of efficacy after cessation of TCZ.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Adulto , Idoso , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/efeitos adversos , Antirreumáticos/administração & dosagem , Antirreumáticos/efeitos adversos , Quimioterapia Combinada , Feminino , Humanos , Infliximab , Masculino , Pessoa de Meia-Idade , Recidiva , Retratamento , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
OBJECTIVES: TNF-like weak inducer of apoptosis (TWEAK), a member of the TNF superfamily, has been shown to increase cytokine production by rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS). In this study, we determined the effect of interaction between TWEAK and its receptor fibroblast growth factor-inducible-14 (Fn14) on cytokine expression in RAFLS. METHODS: RAFLS were obtained from surgical synovial specimens and used at passage 5-10. Cytokine protein and mRNA expression were measured with ELISA and real time-PCR, respectively. Apoptotic cells were detected by TUNEL assay. RelB activation was detected by Western blot analysis. RESULTS: TWEAK inhibited IL-6 production from total synovial cells from RA. TWEAK weakly induced FLS IL-6 and IL-8, but in contrast TWEAK dose-dependently inhibited IL-6 and IL-8 production by TNFα-activated FLS. TWEAK did not induce apoptosis in FLS but inhibited proliferation of TNFα-activated FLS. TWEAK induced RelB activation and suppressed IL-6 mRNA expression in TNFα-activated FLS and both of these phenomenon were abolished by inhibition of new protein synthesis with cycloheximide. CONCLUSIONS: TWEAK has a previously unsuspected inhibitory effect on cytokine production by TNFα-activated RAFLS. This observation suggests that the effects of TWEAK on cytokine expression varies with the pro-inflammatory context, and that in TNFα-activated states such as RA TWEAK may have a net inhibitory effect.
Assuntos
Artrite Reumatoide/metabolismo , Interleucina-6/antagonistas & inibidores , Interleucina-6/metabolismo , Receptores do Fator de Necrose Tumoral/metabolismo , Membrana Sinovial/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fatores de Necrose Tumoral/metabolismo , Apoptose/genética , Apoptose/fisiologia , Artrite Reumatoide/genética , Artrite Reumatoide/patologia , Citocina TWEAK , Fibroblastos/metabolismo , Fibroblastos/patologia , Humanos , Interleucina-6/genética , Interleucina-8/metabolismo , RNA Mensageiro/genética , Proteínas Recombinantes/metabolismo , Membrana Sinovial/patologia , Receptor de TWEAK , Fator de Transcrição RelB/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/metabolismoRESUMO
IFN-gamma has significant immunoregulatory activity and plays an important role in both innate and adaptive immunity. Additive effects of IFN-gamma and the Toll-like receptor ligand LPS has been investigated in macrophages, but in fibroblasts is incompletely understood. IFN-gamma and LPS synergistically induced MCP-1 and NO release in primary murine dermal fibroblasts. IFN-gamma enhanced LPS-induced JNK and p38 MAPK phosphorylation but had no effect on NF-kappaB activity. The induction of both MCP-1 and NO was attenuated by inhibition of JNK but not p38 MAPK. Serine 727 STAT1 phosphorylation by IFN-gamma was increased by LPS, and this was also attenuated by inhibition of JNK but not p38 MAPK. IFN-gamma inhibited the basal expression of MAPK phosphatase-1, a negative regulator of MAPK signaling pathway. These results suggest that enhancement of LPS-induced JNK activation by IFN-gamma associated with inhibition of MAPK phosphatase-1 may be one of the mechanisms of additive effects between IFN-gamma and LPS in fibroblasts.
Assuntos
Quimiocina CCL2/imunologia , Fosfatase 1 de Especificidade Dupla/imunologia , Fibroblastos/imunologia , Interferon gama/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Lipopolissacarídeos/imunologia , Animais , Antracenos/metabolismo , Células Cultivadas , Ativação Enzimática , Inibidores Enzimáticos/metabolismo , Fibroblastos/citologia , Humanos , Imidazóis/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Piridinas/metabolismo , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismoRESUMO
CD16+ monocytes, identified as a minor population of monocytes in human peripheral blood, have been implicated in several inflammatory diseases, including rheumatoid arthritis (RA). Fractalkine (FKN, CX3CL1), a member of the CX3 C subfamily, is induced by pro-inflammatory cytokines, while a receptor for FKN, CX3CR1, is capable of mediating both leukocyte migration and firm adhesion. Here, we investigated the role of FKN and CX3CR1 in activation of CD16+ monocytes and their recruitment into synovial tissues in RA patients. High levels of soluble FKN were detected in the synovial fluid and sera of RA patients. Circulating CD16+ monocytes showed a higher level of CX3CR1 expression than CD16- monocytes in both RA patients and healthy subjects. High level expression of CX3CR1 was also seen in CD16+ monocytes localized to the lining layer in RA synovial tissue. In the in vitro culture experiments, IL-10 induced CX3CR1 expression on the surface of monocytes, and TNFalpha induced membrane-bound FKN as well as soluble FKN expression in synovial fibroblasts. Moreover, soluble FKN was capable of inducing IL-1beta and IL-6 by activated monocytes. These results suggest that FKN might preferentially mediate migration and recruitment of CD16+ monocytes, and might contribute to synovial tissue inflammation.
Assuntos
Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Quimiocinas CX3C/metabolismo , Proteínas de Membrana/metabolismo , Monócitos/metabolismo , Monócitos/patologia , Receptores de Quimiocinas/metabolismo , Receptores de IgG/metabolismo , Idoso , Artrite Reumatoide/sangue , Receptor 1 de Quimiocina CX3C , Membrana Celular/metabolismo , Células Cultivadas , Quimiocina CX3CL1 , Quimiocinas CX3C/sangue , Quimiocinas CX3C/farmacologia , Células Endoteliais/metabolismo , Feminino , Humanos , Interleucina-10/farmacologia , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Proteínas de Membrana/sangue , Proteínas de Membrana/farmacologia , Monócitos/efeitos dos fármacos , Osteoartrite/sangue , Osteoartrite/metabolismo , Osteoartrite/patologia , Proteínas Recombinantes/farmacologia , Líquido Sinovial/metabolismo , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
The inflamed synovial tissue (ST) of rheumatoid arthritis (RA) is characterized by the selective accumulation of interferon gamma-producing Th1-type CD4+ T cells. In this study, we investigated whether the predominance of Th1-type CD4+ cells in the ST lesion is mediated by their selective recruitment through Th1 cell-associated chemokine receptors CXCR3 and CCR5. The lymphocyte aggregates in the ST of RA contained a large number of CD4+ T cells, which mostly expressed both CXCR3 and CCR5, but not CCR4. In contrast, the frequencies of CD4+ and CD8+ T cells expressing CXCR3 and CCR5 in the blood were significantly decreased in RA patients, compared with healthy controls (HC), although there was no difference in the frequencies of CCR4-expressing CD4+ and CD8+ T cells between RA and HC. CXCR3, CCR5, and CCR4 expression in blood CD4 + T cells and CXCR3 expression in CD8+ T cells were increased after interleukin-15 (IL-15) stimulation. Therefore, the distribution of Th1-type CD4+ T cells into the ST from the blood in RA may be associated with the local expression of chemokines, both CXCR3 and CCR5 ligands, and IL-15 may play a role in enhancing these chemokine receptors on CD4+ T cell infiltrates.
Assuntos
Artrite Reumatoide/imunologia , Artrite Reumatoide/fisiopatologia , Linfócitos T CD4-Positivos/química , Linfócitos T CD4-Positivos/patologia , Receptores CCR5/metabolismo , Receptores de Quimiocinas/metabolismo , Membrana Sinovial/imunologia , Idoso , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/química , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/patologia , Estudos de Casos e Controles , Movimento Celular/fisiologia , Feminino , Citometria de Fluxo , Humanos , Interleucina-15/fisiologia , Ligantes , Masculino , Pessoa de Meia-Idade , Receptores CCR4 , Receptores CXCR3 , Receptores de Quimiocinas/análise , Membrana Sinovial/fisiopatologia , Células Th1/imunologia , Células Th1/patologiaRESUMO
UNLABELLED: The inability to match rheumatoid arthritis (RA) patients with the anti-cytokine agent most efficacious for them is a major hindrance to patients' speedy recovery and to the clinical use of anti-cytokine therapy. Identifying predictive biomarkers that can assist in matching RA patients with more suitable anti-cytokine treatment was our aim in this report. The sample consisted of 138 RA patients (naïve and non-naïve) who were administered tocilizumab or etanercept for a minimum of 16 weeks as a prescribed RA treatment. Pretreatment serum samples were obtained from patients and clinical measures of their disease activity were evaluated at baseline and 16 weeks after treatment commenced. Using patients' pretreatment serum, we measured 31 cytokines/chemokines/soluble receptors and used multiple linear regression analysis to identify biomarkers that correlated with patients' symptom levels (DAS28-CRP score) at week 16 and multiple logistic analyses for biomarkers that correlated with patients' final outcome. The results revealed that sgp130, logIL-6, logIL-8, logEotaxin, logIP-10, logVEGF, logsTNFR-I and logsTNFR-II pretreatment serum levels were predictive of the week 16 DAS28-CRP score in naïve tocilizumab patients while sgp130, logGM-CSF and logIP-10 were predictive in non-naïve patients. Additionally, we found logIL-9, logVEGF and logTNF-α to be less reliable at predicting the week 16 DAS28-CRP score in naïve etanercept patients. Multiple linear regression and multiple logistic regression analyses identified biomarkers that were predictive of remission/non-remission in tocilizumab and etanercept therapy. Although less reliable than those for tocilizumab, we identified a few possible biomarkers for etanercept therapy. The biomarkers for these two therapies differ suggesting that their efficacy will vary for individual patients. We discovered biomarkers in RA pretreatment serum that predicted their week 16 DAS28-CRP score and clinical outcome to tocilizumab therapy. Most of these biomarkers, especially sgp130, are involved in RA pathogenesis and IL-6 signal transduction, which further suggests that they are highly reliable. TRIAL REGISTRATION: UMIN-CTR Clinical Trial UMIN000016298.
Assuntos
Antirreumáticos/uso terapêutico , Artrite Reumatoide/sangue , Artrite Reumatoide/tratamento farmacológico , Biomarcadores/sangue , Quimiocinas/sangue , Receptores de Quimiocinas/sangue , Anticorpos Monoclonais Humanizados/uso terapêutico , Receptor gp130 de Citocina/sangue , Etanercepte/uso terapêutico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Curva ROC , Análise de Regressão , Indução de Remissão , Solubilidade , Resultado do TratamentoRESUMO
High levels of soluble CD30 (sCD30) were detected in the serum and synovial fluid of patients with rheumatoid arthritis (RA), indicating the involvement of CD30+ T cells in the pathogenesis. We investigated the induction of CD30 and its functions in CD4+T cells from patients with established RA (disease duration >_2 years). CD4+ T cells from both the peripheral blood (PB) and synovial tissue (ST) of RA patients expressed surface CD30 when stimulated with anti-CD3 antibody (Ab) and anti-CD28 Ab, but their CD30 induction was slower and weaker compared with PB CD4+ T cells of healthy controls (HC). Immunohistochemical analysis showed that only a small proportion of lymphocytes expressed CD30 in the ST (-1%). RA PB CD4+ T cells, after recovery from 6-day stimulation with anti-CD3 Ab and anti-CD28 Ab, showed in intracellular cytokine staining that CD30+ T cells could produce more interleukin-4 (IL-4) but less interferon-gamma. In the culture of RA PB CD4+ T Cells with anti-CD3 Ab and anti-CD28 Ab, blocking anti-CD30 Ab similarly inhibited the cell proliferation and activation of nuclear factor-kappaB on day 4 in RA and HC, but inhibited the apoptotic cell death on day 6 only in RA. These results indicate that despite high-level expression of sCD30, the anti-inflammatory activity of IL-4-producing CD30+ CD4+ T cells may be limited in the ST due to a poor induction of surface CD30 and a susceptibility to CD30-mediated cell death.
Assuntos
Artrite Reumatoide/imunologia , Artrite Reumatoide/fisiopatologia , Linfócitos T CD4-Positivos/imunologia , Antígeno Ki-1/sangue , Antígeno Ki-1/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose/imunologia , Antígenos CD4/metabolismo , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Divisão Celular/imunologia , Feminino , Expressão Gênica/imunologia , Humanos , Interleucina-4/metabolismo , Masculino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Transdução de Sinais/imunologia , Solubilidade , Membrana Sinovial/citologia , Membrana Sinovial/imunologia , Membrana Sinovial/metabolismoRESUMO
INTRODUCTION: Anemia of inflammation (AI) is a common complication of rheumatoid arthritis (RA) and has a negative impact on RA symptoms and quality of life. Upregulation of hepcidin by inflammatory cytokines has been implicated in AI. In this study, we evaluated and compared the effects of IL-6 and TNF-α blocking therapies on anemia, disease activity, and iron-related parameters including serum hepcidin in RA patients. METHODS: Patients (n = 93) were treated with an anti-IL-6 receptor antibody (tocilizumab) or TNF-α inhibitors for 16 weeks. Major disease activity indicators and iron-related parameters including serum hepcidin-25 were monitored before and 2, 4, 8, and 16 weeks after the initiation of treatment. Effects of tocilizumab and infliximab (anti-TNF-α antibody) on cytokine-induced hepcidin expression in hepatoma cells were analyzed by quantitative real-time PCR. RESULTS: Anemia at base line was present in 66% of patients. Baseline serum hepcidin-25 levels were correlated positively with serum ferritin, C-reactive protein (CRP), vascular endothelial growth factor (VEGF) levels and Disease Activity Score 28 (DAS28). Significant improvements in anemia and disease activity, and reductions in serum hepcidin-25 levels were observed within 2 weeks in both groups, and these effects were more pronounced in the tocilizumab group than in the TNF-α inhibitors group. Serum hepcidin-25 reduction by the TNF-α inhibitor therapy was accompanied by a decrease in serum IL-6, suggesting that the effect of TNF-α on the induction of hepcidin-25 was indirect. In in vitro experiments, stimulation with the cytokine combination of IL-6+TNF-α induced weaker hepcidin expression than did with IL-6 alone, and this induction was completely suppressed by tocilizumab but not by infliximab. CONCLUSIONS: Hepcidin-mediated iron metabolism may contribute to the pathogenesis of RA-related anemia. In our cohort, tocilizumab was more effective than TNF-α inhibitors for improving anemia and normalizing iron metabolism in RA patients by inhibiting hepcidin production.
Assuntos
Anemia/prevenção & controle , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Hepcidinas/sangue , Adulto , Idoso , Anemia/sangue , Antirreumáticos/uso terapêutico , Artrite Reumatoide/sangue , Artrite Reumatoide/patologia , Proteína C-Reativa/metabolismo , Linhagem Celular Tumoral , Feminino , Ferritinas/sangue , Expressão Gênica/efeitos dos fármacos , Hepcidinas/genética , Humanos , Infliximab , Interleucina-6/sangue , Interleucina-6/genética , Interleucina-6/farmacologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Células U937 , Fator A de Crescimento do Endotélio Vascular/sangueRESUMO
OBJECTIVE: The accumulation of advanced glycation end products (AGEs), S100A12, and high mobility group box chromosomal protein 1 has been associated with joint inflammation in rheumatoid arthritis (RA). This study was undertaken to determine the induction of the receptor for these proteins, termed receptor for AGEs (RAGE), in synovial tissue (ST) macrophages from RA patients. METHODS: RAGE and CD68 expression in ST were determined by 2-color immunofluorescence labeling. Cell surface and messenger RNA (mRNA) expression of RAGE were examined by flow cytometry and reverse transcriptase-polymerase chain reaction (PCR) or real-time PCR, respectively. RESULTS: CD68+ lining macrophages, like the vasculature, expressed high levels of RAGE in inflamed ST from RA patients. RAGE mRNA expression was significantly higher in RA ST than in ST from patients with osteoarthritis. RAGE mRNA levels were significantly higher in ST macrophages and normal endothelial cells than in ST CD4+ T cells and synovial fibroblasts stimulated with tumor necrosis factor alpha and interleukin-1beta (IL-1beta). Cell surface RAGE was highly induced on normal monocytes after a 24-hour incubation with a 20% concentration of RA ST cell culture supernatants. RAGE mRNA expression in adherent monocytes was augmented by various cytokines, most potently by IL-1beta. CONCLUSION: These results indicate that RAGE overexpression in lining macrophages may be induced, at least in part, by cytokines such as IL-1, leading to the amplification of inflammatory responses mediated by RAGE ligands that are abundant in RA joints.
Assuntos
Artrite Reumatoide/imunologia , Artrite Reumatoide/metabolismo , Macrófagos/metabolismo , Receptores Imunológicos/biossíntese , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Receptor para Produtos Finais de Glicação Avançada , Membrana Sinovial/imunologiaRESUMO
Despite its potent ability to inhibit proinflammatory cytokine synthesis, interleukin (IL)-10 has a marginal clinical effect in rheumatoid arthritis (RA) patients. Recent evidence suggests that IL-10 induces monocyte/macrophage maturation in cooperation with macrophage-colony stimulating factor (M-CSF). In the present study, we found that the inducible subunit of the IL-10 receptor (IL-10R), type 1 IL-10R (IL-10R1), was expressed at higher levels on monocytes in RA than in healthy controls, in association with disease activity, while their expression of both type 1 and 2 tumour necrosis factor receptors (TNFR1/2) was not increased. The expression of IL-10R1 but not IL-10R2 was augmented on monocytes cultured in the presence of RA synovial tissue (ST) cell culture supernatants. Cell surface expression of TNFR1/2 expression on monocytes was induced by IL-10, and more efficiently in combination with M-CSF. Two-color immunofluorescence labeling of RA ST samples showed an intensive coexpression of IL-10R1, TNFR1/2, and M-CSF receptor in CD68+ lining macrophages. Adhered monocytes, after 3-day preincubation with IL-10 and M-CSF, could produce more IL-1beta and IL-6 in response to TNF-alpha in the presence of dibutyryl cAMP, as compared with the cells preincubated with or without IL-10 or M-CSF alone. Microarray analysis of gene expression revealed that IL-10 activated various genes essential for macrophage functions, including other members of the TNFR superfamily, receptors for chemokines and growth factors, Toll-like receptors, and TNFR-associated signaling molecules. These results suggest that IL-10 may contribute to the inflammatory process by facilitating monocyte differentiation into TNF-alpha-responsive macrophages in the presence of M-CSF in RA.
Assuntos
Artrite Reumatoide/metabolismo , Interleucina-10/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/metabolismo , Receptores do Fator de Necrose Tumoral/metabolismo , Adulto , Idoso , Artrite Reumatoide/sangue , Artrite Reumatoide/fisiopatologia , Extratos Celulares/farmacologia , Células Cultivadas , Citocinas/biossíntese , Citocinas/sangue , Combinação de Medicamentos , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-10/genética , Interleucina-10/farmacologia , Subunidade alfa de Receptor de Interleucina-10/metabolismo , Fator Estimulador de Colônias de Macrófagos/farmacologia , Masculino , Análise em Microsséries , Pessoa de Meia-Idade , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Receptor de Fator Estimulador de Colônias de Macrófagos/sangue , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/biossíntese , Receptores Tipo I de Fatores de Necrose Tumoral/sangue , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Receptores Tipo II do Fator de Necrose Tumoral/biossíntese , Receptores Tipo II do Fator de Necrose Tumoral/sangue , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Proteínas Recombinantes/farmacologia , Membrana Sinovial/química , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
S100A8 and S100A9, two Ca2+-binding proteins of the S100 family, are secreted as a heterodimeric complex (S100A8/A9) from neutrophils and monocytes/macrophages. Serum and synovial fluid levels of S100A8, S100A9, and S100A8/A9 were all higher in patients with rheumatoid arthritis (RA) than in patients with osteoarthritis (OA), with the S100A8/A9 heterodimer being prevalent. By two-color immunofluorescence labeling, S100A8/A9 antigens were found to be expressed mainly by infiltrating CD68+ macrophages in RA synovial tissue (ST). Isolated ST cells from patients with RA spontaneously released larger amounts of S100A8/A9 protein than did the cells from patients with OA. S100A8/A9 complexes, as well as S100A9 homodimers, stimulated the production of proinflammatory cytokines, such as tumor necrosis factor alpha, by purified monocytes and in vitro-differentiated macrophages. S100A8/A9-mediated cytokine production was suppressed significantly by p38 mitogen-activated protein kinase (MAPK) inhibitors and almost completely by nuclear factor kappa B (NF-kappaB) inhibitors. NF-kappaB activation was induced in S100A8/A9-stimulated monocytes, but this activity was not inhibited by p38 MAPK inhibitors. These results indicate that the S100A8/A9 heterodimer, secreted extracellularly from activated tissue macrophages, may amplify proinflammatory cytokine responses through activation of NF-kappaB and p38 MAPK pathways in RA.
Assuntos
Artrite Reumatoide/fisiopatologia , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Citocinas/biossíntese , Macrófagos/imunologia , NF-kappa B/metabolismo , Líquido Sinovial/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Idoso , Artrite Reumatoide/imunologia , Dimerização , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Líquido Sinovial/citologiaRESUMO
Systemic lupus erythematosus (SLE) is a serious systemic autoimmune disease of unknown etiology. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that is operative in innate and adaptive immunity and important in immune-mediated diseases such as rheumatoid arthritis and atherosclerosis. The functional relevance of MIF in systemic autoimmune diseases such as SLE is unknown. Using the lupus-prone MRL/lpr mice, we aim to examine the expression and function of MIF in this murine model of systemic autoimmune disease. These experiments revealed that renal MIF expression was significantly higher in MRL/lpr mice compared with nondiseased control mice (MRL/MpJ), and MIF was also markedly up-regulated in skin lesions of MRL/lpr mice. To examine the effect of MIF on development of systemic autoimmune disease, we generated MRL/lpr mice with a targeted disruption of the MIF gene (MIF(-/-)MRL/lpr), and compared their disease manifestations to MIF(+/+)MRL/lpr littermates. MIF(-/-)MRL/lpr mice exhibited significantly prolonged survival, and reduced renal and skin manifestations of SLE. These effects occurred in the absence of major changes in T and B cell markers or alterations in autoantibody production. In contrast, renal macrophage recruitment and glomerular injury were significantly reduced in MIF(-/-)MRL/lpr mice, and this was associated with reduction in the monocyte chemokine MCP-1. Taken together, these data suggest MIF as a critical effector of organ injury in SLE.
Assuntos
Quimiotaxia , Glomerulonefrite/imunologia , Fatores Inibidores da Migração de Macrófagos/deficiência , Fatores Inibidores da Migração de Macrófagos/fisiologia , Macrófagos/fisiologia , Animais , Rim/química , Rim/patologia , Lúpus Eritematoso Sistêmico/patologia , Fatores Inibidores da Migração de Macrófagos/análise , Camundongos , Camundongos Endogâmicos MRL lpr , Mortalidade , Pele/química , Pele/patologiaRESUMO
A 58-year-old Japanese woman developed spiking fever, polyarthralgia and hepatosplenomegaly, with highly elevated levels of c-creactive protein (CRP) and ferrtin, and elevated erythrocyte sedimentation rate (ESR). The AOSD was diagnosed according to the Yamaguchi-criteria of 1992. She was first treated with a combination of prednisolone (20 mg/day) and oral methotrexate (MTX) (7.5 mg/week). This combination, however, was not effective with tapering the dose of prednisolone. When a high dose of cyclosporin A (CyA) (5.5 mg/kg/day) was then added to MTX (5 mg/week), the patient's fever and polyarthralgia decreased, and her elevated serological parameters such as CRP and ESR also gradually declined. Finally, the dose of prednisolone was tapered to 10 mg/day. Only a few cases of AOSD treated with CyA plus MTX have been reported. Thus, further careful observation will be needed to establish the usefulness of this drug combination as a therapy for AOSD.
Assuntos
Ciclosporina/administração & dosagem , Imunossupressores/administração & dosagem , Metotrexato/administração & dosagem , Anti-Inflamatórios/administração & dosagem , Quimioterapia Combinada , Feminino , Humanos , Pessoa de Meia-Idade , Prednisolona/administração & dosagem , Resultado do TratamentoRESUMO
IL-10 has been shown to block the antigen-specific T-cell cytokine response by inhibiting the CD28 signaling pathway. We found that peripheral blood CD4+ T cells from patients with active rheumatoid arthritis (RA) were able to produce greater amounts of interferon gamma after CD3 and CD28 costimulation in the presence of 1 ng/ml IL-10 than were normal control CD4+ T cells, although their surface expression of the type 1 IL-10 receptor was increased. The phosphorylation of signal transducer and activator of transcription 3 was sustained in both blood and synovial tissue CD4+ T cells of RA, but it was not augmented by the presence of 1 ng/ml IL-10. Sera from RA patients induced signal transducer and activator of transcription 3 phosphorylation in normal CD4+ T cells, which was mostly abolished by neutralizing anti-IL-6 antibody. Preincubation of normal CD4+ T cells with IL-6 reduced IL-10-mediated inhibition of interferon gamma production. Blood CD4+ T cells from RA patients contained higher levels of suppressor of cytokine signaling 1 but lower levels of suppressor of cytokine signaling 3 mRNA compared with control CD4+ T cells, as determined by real-time PCR. These results indicate that RA CD4+ T cells become resistant to the immunosuppressive effect of IL-10 before migration into synovial tissue, and this impaired IL-10 signaling may be associated with sustained signal transducer and activator of transcription 3 activation and suppressor of cytokine signaling 1 induction.
Assuntos
Artrite Reumatoide/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Interferon gama/biossíntese , Interleucina-10/farmacologia , Proteínas Repressoras/biossíntese , Proteínas Supressoras da Sinalização de Citocina/biossíntese , Adulto , Idoso , Antígenos CD28/fisiologia , Complexo CD3/fisiologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas/efeitos dos fármacos , Células Cultivadas/imunologia , Células Cultivadas/metabolismo , Feminino , Humanos , Interferon gama/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Pessoa de Meia-Idade , Fosforilação , Processamento de Proteína Pós-Traducional , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores de Interleucina/biossíntese , Receptores de Interleucina/genética , Receptores de Interleucina-10 , Proteínas Repressoras/genética , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina/genéticaRESUMO
OBJECTIVE: Interleukin 12 (IL-12) and IL-18 synergistically induce interferon-gamma (IFN-gamma) production by T cell infiltrates in rheumatoid arthritis (RA). To investigate this synergism, we examined the expression and regulation of IL-12 receptor (IL-12R) and IL-18R on peripheral blood (PB) and synovial tissue (ST) CD4+ T cells from patients with RA. METHODS: The mRNA and cell surface expression of IL-12R and IL-18R in CD4+ T cells were determined by reverse transcriptase-polymerase chain reaction and flow cytometry, respectively. IFN-gamma and IL-4 production by CD4+ T cells stimulated with phorbol myristate acetate (PMA) and calcium ionophore A23187 was measured by intracellular cytokine staining and flow cytometry. RESULTS: Despite the negligible expression of IL-12R on fresh cells, PB CD4+ T cells from RA patients expressed higher levels of both IL-12Rbeta1 and beta2 subunits after stimulation with anti-CD3 antibody (Ab) than the cells of healthy controls. ST CD4+ T cells contained mRNA transcripts encoding IL-12Rbeta1 and beta2, and expressed detectable levels of these 2 subunits on the cell surface. Their IL-12R expression was markedly augmented by costimulation with anti-CD3 Ab and IL-18. In contrast, IL-18Ralpha was expressed on freshly isolated PB CD4+ T cells from RA patients and controls, and the level of expression was higher in RA. IL-18Ralpha+ CD4+ T cells were further increased in the ST lesion, where IL-18Rbeta mRNA was constitutively detected. IL-12Rbeta1 and beta2 were induced mainly on IL-18Ralpha+ CD4+ T cells after anti-CD3 Ab stimulation. PMA and A23187-activated ST CD4+ T cells mostly expressed IL-18Ralpha and produced high levels of IFN-gamma. CONCLUSION: These results indicate that IL-18R-expressing CD4+ T cells are accumulated in the ST of patients with RA, where the functional IL-12R is locally induced by stimuli such as CD3 activation and IL-18. Activation of both cytokine receptors may be necessary for the IFN-gamma-dominant cytokine response.
Assuntos
Artrite Reumatoide/imunologia , Linfócitos T CD4-Positivos/imunologia , Receptores de Interleucina/biossíntese , Adulto , Idoso , Feminino , Humanos , Interferon gama/biossíntese , Subunidade alfa de Receptor de Interleucina-18 , Masculino , Pessoa de Meia-Idade , Receptores de Interleucina/análise , Receptores de Interleucina-12 , Receptores de Interleucina-18 , Membrana Sinovial/química , Células Th1/imunologiaRESUMO
OBJECTIVE: To examine whether depsipeptide (FK228), a histone deacetylase (HDA) inhibitor, has inhibitory effects on the proliferation of synovial fibroblasts from rheumatoid arthritis (RA) patients, and to examine the effects of systemic administration of FK228 in an animal model of arthritis. METHODS: Autoantibody-mediated arthritis (AMA) was induced in 19 male DBA/1 mice (6-7 weeks old); 10 of them were treated by intravenous administration of FK228 (2.5 mg/kg), and 9 were used as controls. The effects of FK228 were examined by radiographic, histologic, and immunohistochemical analyses and arthritis scores. RA synovial fibroblasts (RASFs) were obtained at the time of joint replacement surgery. In vitro effects of FK228 on cell proliferation were assessed by MTT assay. Cell morphology was examined by light and transmission electron microscopy. The effects on the expression of the cell cycle regulators p16INK4a and p21(WAF1/Cip1) were examined by real-time polymerase chain reaction and Western blot analysis. The acetylation status of the promoter regions of p16INK4a and p21(WAF1/Cip1) were determined by chromatin immunoprecipitation assay. RESULTS: A single intravenous injection of FK228 (2.5 mg/ml) successfully inhibited joint swelling, synovial inflammation, and subsequent bone and cartilage destruction in mice with AMA. FK228 treatment induced histone hyperacetylation in the synovial cells and decreased the levels of tumor necrosis factor alpha and interleukin-1beta in the synovial tissues of mice with AMA. FK228 inhibited the in vitro proliferation of RASFs in a dose-dependent manner. Treatment of cells with FK228 induced the expression of p16INK4a and up-regulated the expression of p21(WAF1/Cip1). These effects of FK228 on p16INK4a and p21(WAF1/Cip1) were related to the acetylation of the promoter region of the genes. CONCLUSION: Our findings strongly suggest that systemic administration of HDA inhibitors may represent a novel therapeutic target in RA by means of cell cycle arrest in RASFs via induction of p16INK4a expression and increase in p21(WAF1/Cip1) expression.