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1.
Life Sci ; 269: 118987, 2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33417958

RESUMO

AIMS: To explore the therapeutic effect of miR-129-5p carried by exosomes from Human Synovial Mesenchymal Stem Cell (HS-MSC) on osteoarthritis(OA). MATERIALS AND METHODS: The levels of miR-129-5p and high mobility group protein -1 (HMGB1) and interleukin-1ß (IL-1ß) in the joint fluid of OA patients were respectively detected via real-time quantitative reverse transcription-PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). IL-1ß was taken to act on chondrocytes for the establishment of OA model in vitro. Ultracentrifugation was conducted to isolate HS-MSC exosomes (HS-MSC-Exo) from the supernatant. Western blot and ELISA were carried out to measure the expression of iNOS, COX2, MMP13, Collagen 2, TLR4, NF-κB, Caspase3, Bcl-2, HMGB1 in chondrocytes. Flow cytometry was conducted to detect the apoptosis of chondrocytes. Besides, bioinformatics was employed to predict the targeted relationship between miR-129-5p and HMGB1, which was further verified via dual luciferase activity experiments. KEY FINDINGS: The results illustrated that miR-129-5p was decreased in OA patients and IL-1ß-induced chondrocytes, while HMGB1 was notably upregulated. HS-MSC-Exo rich in miR-129-5p remarkably declined the inflammatory response and apoptosis of chondrocytes, while HS-MSC-Exo deficient in miR-129-5p increased the IL-1ß-mediated inflammatory response and apoptosis of chondrocytes. In terms of mechanism, miR-129-5p targets the 3'UTR end of HMGB1 and inhibits IL-1ß-mediated upregulation of HMGB1. SIGNIFICANCE: In a word, this paper proved that miR-129-5p, existing in HS-MSC-Exo, can suppress the IL-1ß-mediated OA by inhibiting HMGB1 release.


Assuntos
Exossomos/metabolismo , Proteína HMGB1/metabolismo , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Osteoartrite/metabolismo , Membrana Sinovial/metabolismo , Sequência de Bases , Condrócitos/metabolismo , Condrócitos/patologia , Feminino , Regulação da Expressão Gênica , Proteína HMGB1/genética , Humanos , Interleucina-1beta/metabolismo , Articulações/patologia , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Líquido Sinovial/metabolismo
2.
Gene ; 766: 145149, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-32971185

RESUMO

BACKGROUND: Crosstalk between posterior cruciate ligament fibroblasts (PCLfs) and synoviocytes (SCs) significantly modifies the homeostatic balance of the extracellular matrix (ECM) and appears to post a prominent affection for wound healing of PCL. Interleukin-1ß (IL-1ß) is regarded as a critical factor in acute inflammatory events during ligament injury. METHODS: In order to confirm the capability of SCs the response of lysyl oxidases (LOXs) and matrix metalloproteinases (MMPs) to IL-1ß, the complex cues of the joint cavity following PCL injury were simulated and the effect of IL-1ß on the expression of LOXs and MMPs in PCLfs were investigated. PCLfs in both the mono- and co-culture conditions were treated with IL-1ß. Cell lysates were collected from the PCLfs and LOXs and MMP-1, 2, 3 expression quantified using quantitative real-time PCR and western bolting. RESULTS: The results indicated that injury alone elevated the expression of LOXs and MMP-1, 2 and 3. But IL-1ß significantly decreased the LOX, LOXL1, and LOXL3 expression, and simultaneously increased MMP-1, 2 and 3 expressions in injured PCLfs. Furthermore, co-culture further suppressed LOXs, but stimulated MMP-1, 2 and 3 expressions when subjected to both mechanical injury and IL-1ß treatment. This possibly suggests that a number of soluble factors are secreted that act as mediators that amplify the response of SCs. CONCLUSION: The results indicated that the SCs could affect the IL-1ß-induction of LOXs inhibition and MMPs accumulation, which may be the underlying mechanism of the the poor healing response following PCL injury.


Assuntos
Fibroblastos/metabolismo , Interleucina-1beta/metabolismo , Metaloproteinases da Matriz/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Sinoviócitos/metabolismo , Células Cultivadas , Técnicas de Cocultura/métodos , Matriz Extracelular/metabolismo , Humanos , Membrana Sinovial/metabolismo , Cicatrização/fisiologia
3.
PLoS One ; 15(12): e0242868, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33382721

RESUMO

Rheumatoid arthritis (RA) and periodontitis (PD) are chronic inflammatory diseases that appear to occur in tandem. However, the mutual impact PD exerts on RA and vice versa has not yet been defined. To address this issue, we set up an animal model and analyzed how two prime inducers of periodontitis-Porphyromonas gingivalis (Pg) and Aggregatibacter actinomycetemcomitans (Aa)-differ in their pathogenic potential. Our experimental setup included collagen induced arthritis (CIA) in the mouse, oral inoculation with Pg or Aa to induce alveolar bone loss and the combination of both diseases in inverted orders of events. Neither pathobiont impacted on macroscopic arthritis and arthritis did not exacerbate alveolar bone loss. However, there were subtle differences between Pg and Aa with the former inducing more alveolar bone loss if PD was induced before CIA. On a molecular level, Pg and Aa led to differential expression patterns in the synovial membranes that were reminiscent of cellular and humoral immune responses, respectively. The Pg and Aa specific signatures in the synovial proteomes suggest a role for oral pathogens in shaping disease subtypes and setting the stage for subsequent therapy response.


Assuntos
Aggregatibacter actinomycetemcomitans/fisiologia , Artrite Reumatoide/metabolismo , Artrite Reumatoide/microbiologia , Porphyromonas gingivalis/fisiologia , Proteoma/metabolismo , Membrana Sinovial/metabolismo , Animais , Citocinas/metabolismo , Camundongos , Periodontite/microbiologia , Membrana Sinovial/microbiologia
4.
PLoS One ; 15(10): e0237520, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33002030

RESUMO

OBJECTIVES: Gout is the most prevalent inflammatory arthritis. To study the effects of regular physical activity and exercise intensity on inflammation and clinical outcome, we examined inflammatory pathogenesis in an acute model of murine gout and analyzed human gout patient clinical data as a function of physical activity. METHODS: NF-κB-luciferase reporter mice were organized into four groups and exercised at 0 m/min (non-exercise), 8 m/min (low-intensity), 11 m/min (moderate-intensity), and 15 m/min (high-intensity) for two weeks. Mice subsequently received intra-articular monosodium urate (MSU) crystal injections (0.5mg) and the inflammatory response was analyzed 15 hours later. Ankle swelling, NF-κB activity, histopathology, and tissue infiltration by macrophages and neutrophils were measured. Toll-like receptor (TLR)2 was quantified on peripheral monocytes/neutrophils by flow cytometry and both cytokines and chemokines were measured in serum or synovial aspirates. Clinical data and questionnaires accessing overall physical activity levels were collected from gout patients. RESULTS: Injection of MSU crystals produced a robust inflammatory response with increased ankle swelling, NF-κB activity, and synovial infiltration by macrophages and neutrophils. These effects were partially mitigated by low and moderate-intensity exercise. Furthermore, IL-1ß was decreased at the site of MSU crystal injection, TLR2 expression on peripheral neutrophils was downregulated, and expression of CXCL1 in serum was suppressed with low and moderate-intensity exercise. Conversely, the high-intensity exercise group closely resembled the non-exercised control group by nearly all metrics of inflammation measured in this study. Physically active gout patients had significantly less flares/yr, decreased C-reactive protein (CRP) levels, and lower pain scores relative to physically inactive patients. CONCLUSIONS: Regular, moderate physical activity can produce a quantifiable anti-inflammatory effect capable of partially mitigating the pathologic response induced by intra-articular MSU crystals by downregulating TLR2 expression on circulating neutrophils and suppressing systemic CXCL1. Low and moderate-intensity exercise produces this anti-inflammatory effect to varying degrees, while high-intensity exercise provides no significant difference in inflammation compared to non-exercising controls. Consistent with the animal model, gout patients with higher levels of physical activity have more favorable prognostic data. Collectively, these data suggest the need for further research and may be the foundation to a future paradigm-shift in conventional exercise recommendations provided by Rheumatologists to gout patients.


Assuntos
Quimiocina CXCL1/sangue , Gota/terapia , Inflamação/prevenção & controle , Condicionamento Físico Animal , Receptor 2 Toll-Like/sangue , Animais , Modelos Animais de Doenças , Regulação para Baixo , Exercício Físico/fisiologia , Feminino , Gota/sangue , Gota/patologia , Humanos , Inflamação/sangue , Inflamação/patologia , Interleucina-1beta/sangue , Interleucina-1beta/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Neutrófilos/metabolismo , Neutrófilos/patologia , Dor/prevenção & controle , Prognóstico , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia
5.
Yakugaku Zasshi ; 140(9): 1141-1150, 2020.
Artigo em Japonês | MEDLINE | ID: mdl-32879246

RESUMO

Osteoarthritis is the most common joint disorder worldwide and one of the leading causes of disability in the elderly. We have reported that the novel sodium hyaluronate derivative chemically linked with diclofenac (DF), diclofenac etalhyaluronate (SI-613), exerted a potent and long-lasting analgesic effect in experimental arthritis models. In this study, we evaluated the properties of residual SI-613 in the knee joint after an intra-articular (IA) administration. After IA administration of fluorescent labeled SI-613 (FA-SI-613) or fluorescent labeled hyaluronic acid (FA-HA) to rabbits, fluorescence intensities in the synovial membrane and cartilage were higher in the FA-SI-613 group until 7 d after administration than in the FA-HA group. After IA administration of radiolabeled SI-613 (14C-SI-613) to rabbits, the radioactivity remained in the joint cavity and the joint tissues such as synovial membrane and cartilage until 84 d after administration. This residual radioactivity was identified mainly as HA linked with DF, since 14C-SI-613 was labeled at the benzene ring of DF and since more DF-linked HA oligomer was detected on metabolite analysis than free DF in the synovial membrane and synovial lavage fluid up to 28 d after administration. These results suggested that intra-articularly administered SI-613 remained for a longer time in the joint as HA linked with DF than when HA was administered. Therefore, SI-613 was considered to prolong the pharmacological effects of both HA and DF by remaining in the joint as HA linked with DF.


Assuntos
Diclofenaco/administração & dosagem , Diclofenaco/metabolismo , Ácido Hialurônico/análogos & derivados , Articulação do Joelho , Osteoartrite/tratamento farmacológico , Animais , Cartilagem/metabolismo , Modelos Animais de Doenças , Ácido Hialurônico/administração & dosagem , Ácido Hialurônico/metabolismo , Injeções Intra-Articulares , Articulação do Joelho/metabolismo , Masculino , Coelhos , Membrana Sinovial/metabolismo , Fatores de Tempo
6.
Life Sci ; 259: 118250, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32791152

RESUMO

AIMS: Several microbial toll-like receptor (TLR) ligands, bacterial DNA and bacterial cell wall fragments have been identified in the synovium of rheumatoid arthritis (RA) patients, proving bacterial involvement in the pathogenesis of RA. The current study aimed to verify that low dose polymyxin B could prevent the development of chronic inflammatory arthritis. METHODS: Twelve days post adjuvant injection, Sprague-Dawley rats were treated twice weekly with methotrexate (0.5 mg/kg) or daily with polymyxin B (1 mg/kg) or with combination of both for 1 or 2 weeks. Arthritis progression was assessed by hind paw swelling, serum levels of tumor growth factor-1ß (TGF-1ß), tumor necrosis factor-alpha (TNF-α), high sensitivity C-reactive protein (HS-CRP) and nuclear factor kappa B (NF-κB) were measured using ELISA. Cyclooxygenase-1 (Cox-1) and Cox-2 activities, as well as mRNA expression of TLR-2 and TLR-4 were determined. Histopathological examination of the ankle joint was performed as well as immunohistochemistry for anti-TLR-4. Histopathological assessment of toxic effects on the kidney was performed. KEY FINDINGS: Adjuvant arthritis led to a significant swelling of the hind paw and alteration in all serum parameters, TLR-2 and TLR-4 expression, as well as Cox-2 activity. These alterations were associated with histopathological changes of the joints. Polymyxin B reduced significantly all biomarkers of inflammation, showing better effect of the combination in most of the studied parameters, with minimal signs of nephrotoxicity. SIGNIFICANCE: In conclusion, results showed that polymyxin B possesses significant anti-arthritic activity which may be attributed to inhibition of the TLR-4, NF-κB and Cox-2 signaling pathway.


Assuntos
Artrite Experimental/tratamento farmacológico , Polimixina B/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Artrite Experimental/fisiopatologia , Artrite Reumatoide/tratamento farmacológico , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Ciclo-Oxigenase 2/fisiologia , Adjuvante de Freund/farmacologia , Inflamação/tratamento farmacológico , Masculino , NF-kappa B/metabolismo , Polimixina B/metabolismo , Polimixina B/uso terapêutico , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Membrana Sinovial/metabolismo , Receptores Toll-Like/metabolismo , Receptores Toll-Like/fisiologia , Fator de Necrose Tumoral alfa/metabolismo
7.
PLoS One ; 15(6): e0233897, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32484820

RESUMO

OBJECTIVES: miR-155 plays a critical role in the inflammatory process and in diseases such as rheumatoid arthritis (RA). miR155 gene expression is regulated by its gene promoter region CpG island methylation. Previous studies have shown inconsistent changes in circulating levels of mir-155 in RA patients. The aims of our study were to evaluate miR-155 levels in plasma, to investigate its gene methylation level, and to correlate these levels with RA disease activity. METHODS: One hundred and twenty-five patients with RA, and 30 age and sex-matched healthy controls (HC) were enrolled. Whole blood and plasma samples were collected and stored at -80°C until analysis. DAS28 score at the time of the blood draw was used to assess RA disease activity. The methylation status of miR-155 host gene was determined in whole blood by quantitative real-time methylation-specific PCR (qPCR). miR-155 expression levels were evaluated by quantitative reverse transcription PCR. RESULTS: We found significantly lower circulating miR155 levels in RA patients compared to HC. Interestingly, the miR-155 gene methylation level was significantly higher in RA patients than in HC. miR-155 levels did not correlate with ACPA or RF positivity or disease activity. CONCLUSIONS: We show here higher miR-155 methylation in whole blood and lower plasma miR155 expression in RA patients in comparison to HC. The evaluation of miR-155 host gene methylation status or miR155 plasma level might be a potentially useful marker in RA determination.


Assuntos
Artrite Reumatoide/sangue , Biomarcadores/sangue , Metilação de DNA/genética , MicroRNAs/sangue , Artrite Reumatoide/genética , Artrite Reumatoide/patologia , Ilhas de CpG/genética , Feminino , Humanos , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia
8.
Arthritis Rheumatol ; 72(6): 943-956, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32362074

RESUMO

OBJECTIVE: This study was undertaken to uncover the pathophysiologic role of discoidin domain receptor 2 (DDR-2), a putative fibrillar collagen receptor, in inflammation promotion and joint destruction in rheumatoid arthritis (RA). METHODS: In synovial tissue from patients with RA and from mice with collagen antibody-induced arthritis (CAIA) (using Ddr2-/- and DBA/1 mice), gene and protein expression levels of DDR-2, interleukin-15 (IL-15), and Dkk-1 were measured by quantitative reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemistry. Gene knockdown of DDR2 in human RA fibroblast-like synoviocytes (FLS) was conducted via small interfering RNA. Interaction between the long noncoding RNA H19 and microRNA 103a (miR-103a) was assessed in RA FLS using RNA pulldown assays. Cellular localization of H19 was examined using fluorescence in situ hybridization assays. Chromatin immunoprecipitation and dual luciferase reporter assays were applied to verify H19 transcriptional and posttranscriptional regulation by miR-103a. RESULTS: DDR2 messenger RNA (mRNA) expression was significantly associated with the levels of IL-15 and Dkk-1 mRNA in the synovial tissue of RA patients (r2 = 0.2022-0.3293, all P < 0.05; n = 33) and with the serum levels of IL-15 and Dkk-1 in mice with CAIA (P < 0.05). In human RA FLS, activated DDR-2 induced the expression of H19 through c-Myc. Moreover, H19 directly interacted with and promoted the degradation of miR-103a. CONCLUSION: These results indicate a novel role for activated DDR-2 in RA FLS, showing that DDR-2 is responsible for regulating the expression of IL-15 and Dkk-1 in RA FLS and is involved in the promotion of inflammation and joint destruction during pathophysiologic development of RA. Moreover, DDR-2 inhibition, acting through the H19-miR-103a axis, leads to reductions in the inflammatory reaction and severity of joint destruction in mice with CAIA, suggesting that inhibition of DDR-2 may be a potential therapeutic strategy for RA.


Assuntos
Artrite Experimental/metabolismo , Artrite Reumatoide/genética , Receptor com Domínio Discoidina 2/metabolismo , Interleucina-15/metabolismo , Transdução de Sinais/genética , Animais , Regulação da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Inflamação , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Camundongos Endogâmicos DBA , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Membrana Sinovial/metabolismo , Sinoviócitos/metabolismo
9.
Nat Rev Rheumatol ; 16(6): 316-333, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32393826

RESUMO

Rheumatoid arthritis (RA) is a chronic immune-mediated disease that primarily affects the synovium of diarthrodial joints. During the course of RA, the synovium transforms into a hyperplastic invasive tissue that causes destruction of cartilage and bone. Fibroblast-like synoviocytes (FLS), which form the lining of the joint, are epigenetically imprinted with an aggressive phenotype in RA and have an important role in these pathological processes. In addition to producing the extracellular matrix and joint lubricants, FLS in RA produce pathogenic mediators such as cytokines and proteases that contribute to disease pathogenesis and perpetuation. The development of multi-omics integrative analyses have enabled new ways to dissect the mechanisms that imprint FLS, have helped to identify potential FLS subsets with distinct functions and have identified differences in FLS phenotypes between joints in individual patients. This Review provides an overview of advances in understanding of FLS biology and highlights omics approaches and studies that hold promise for identifying future therapeutic targets.


Assuntos
Artrite Reumatoide/imunologia , Reabsorção Óssea/imunologia , Cartilagem Articular/imunologia , Fibroblastos/imunologia , Membrana Sinovial/imunologia , Sinoviócitos/imunologia , Artrite Reumatoide/genética , Artrite Reumatoide/metabolismo , Linfócitos B/imunologia , Reabsorção Óssea/metabolismo , Caderinas/metabolismo , Cartilagem Articular/metabolismo , Metilação de DNA , Células Endoteliais/metabolismo , Epigênese Genética , Fibroblastos/metabolismo , Humanos , Macrófagos/imunologia , Terapia de Alvo Molecular , Monócitos/imunologia , Neovascularização Patológica/imunologia , Neovascularização Patológica/metabolismo , Osteogênese , Proteínas Tirosina Fosfatases/metabolismo , Membrana Sinovial/citologia , Membrana Sinovial/metabolismo , Sinoviócitos/metabolismo , Linfócitos T/imunologia
10.
PLoS One ; 15(4): e0231501, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32330138

RESUMO

Osteoarthritis (OA) is characterized by progressive loss of articular cartilage accompanied by the new bone formation and, often, a synovial proliferation that culminates in pain, loss of joint function, and disability. However, the cellular and molecular mechanisms of OA progression and the relative contributions of cartilage, bone, and synovium remain unclear. We recently found that the extracellular matrix (ECM) protein periostin (Postn, or osteoblast-specific factor, OSF-2) is expressed at high levels in human OA cartilage. Multiple groups have also reported elevated expression of Postn in several rodent models of OA. We have previously reported that in vitro Postn promotes collagen and proteoglycan degradation in human chondrocytes through AKT/ß-catenin signaling and downstream activation of MMP-13 and ADAMTS4 expression. Here we show that Postn induces collagen and proteoglycan degradation in cartilage by signaling through discoidin domain receptor-1 (DDR1), a receptor tyrosine kinase. The genetic deficiency or pharmacological inhibition of DDR1 in mouse chondrocytes blocks Postn-induced MMP-13 expression. These data show that Postn is signaling though DDR1 is mechanistically involved in OA pathophysiology. Specific inhibitors of DDR1 may provide therapeutic opportunities to treat OA.


Assuntos
Doenças das Cartilagens/metabolismo , Cartilagem Articular/metabolismo , Moléculas de Adesão Celular/metabolismo , Receptor com Domínio Discoidina 1/metabolismo , Idoso , Animais , Células Cultivadas , Condrócitos/metabolismo , Feminino , Humanos , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Osteoartrite/metabolismo , Membrana Sinovial/metabolismo
11.
BMC Med Genet ; 21(1): 46, 2020 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-32122327

RESUMO

BACKGROUND: Osteoarthritis (OA) is the most common form of arthritis and a leading cause of disability. This study attempted to investigate the key mRNAs and miRNAs related to OA. PATIENTS AND METHODS: From April 17th, 2018 to May 17th, 2018, five patients with OA and three normal controls were enrolled in this present study. To identify the differentially expressed mRNAs (DEmRNAs) and miRNAs (DEmiRNAs) between patients with OA and normal controls, RNA-sequencing was performed. Then, DEmiRNA-target DEmRNAs analysis and functional annotation of DEmiRNA-target DEmRNAs were performed. To validate the RNA-sequencing results, quantitative real time-PCR (RT-PCR) and western blot analysis were performed as well. RESULTS: A total of 1068 DEmRNAs, 21 DEmiRNAs and 395 DEmiRNA-DEmRNA pairs were identified in synovial tissues of patients with OA. The functional annotation of DEmiRNA-target DEmRNAs revealed that Pathways in cancer and PI3K-Akt signaling pathway were significantly enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. QRT-PCR and western blot results revealed that except for TLR7, the expression level of the others was consistent with the RNA-sequencing results, generally. CONCLUSION: The findings of this present study may provide new clues for the roles of DEmRNAs and DEmiRNAs in the pathogenesis of OA.


Assuntos
MicroRNAs/genética , Osteoartrite/genética , RNA Mensageiro/genética , Membrana Sinovial/metabolismo , Estudos de Casos e Controles , Progressão da Doença , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Humanos , MicroRNAs/análise , Osteoartrite/metabolismo , Osteoartrite/patologia , RNA Mensageiro/análise , Análise de Sequência de RNA/métodos , Membrana Sinovial/química , Membrana Sinovial/patologia , Sequenciamento Completo do Exoma
12.
Nat Commun ; 11(1): 745, 2020 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-32029712

RESUMO

Rheumatoid arthritis affects individuals commonly during the most productive years of adulthood. Poor response rates and high costs associated with treatment mandate the search for new therapies. Here we show that targeting a specific G-protein coupled receptor promotes senescence in synovial fibroblasts, enabling amelioration of joint inflammation. Following activation of the melanocortin type 1 receptor (MC1), synovial fibroblasts acquire a senescence phenotype characterized by arrested proliferation, metabolic re-programming and marked gene alteration resembling the remodeling phase of wound healing, with increased matrix metalloproteinase expression and reduced collagen production. This biological response is attained by selective agonism of MC1, not shared by non-selective ligands, and dependent on downstream ERK1/2 phosphorylation. In vivo, activation of MC1 leads to anti-arthritic effects associated with induction of senescence in the synovial tissue and cartilage protection. Altogether, selective activation of MC1 is a viable strategy to induce cellular senescence, affording a distinct way to control joint inflammation and arthritis.


Assuntos
Artrite Reumatoide/tratamento farmacológico , Receptor Tipo 1 de Melanocortina/agonistas , Animais , Artrite Experimental/tratamento farmacológico , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Variação Genética , Humanos , Imidazóis/farmacologia , Masculino , Redes e Vias Metabólicas/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor Tipo 1 de Melanocortina/deficiência , Receptor Tipo 1 de Melanocortina/genética , Receptores Notch/antagonistas & inibidores , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , alfa-MSH/farmacologia
13.
Arthritis Res Ther ; 22(1): 24, 2020 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-32051018

RESUMO

BACKGROUND: 14-3-3η is an intracellular protein also detected in the serum and synovial fluid of patients with rheumatoid arthritis (RA). It is closely related to disease activity and anti-cyclic citrullinated peptide antibody levels. However, the main source of 14-3-3η and the mechanism of its release into the extracellular space remain unclear. Addressing these two points was the main goal of the current study. METHODS: The source of 14-3-3η was investigated by immunostaining RA synovial tissue. Fibroblast-like synoviocytes, CD4+ cells, and macrophages were selected as candidates among the various cell types in the synovial tissue. Phosphorylation of mixed-lineage kinase domain-like pseudokinase (MLKL) and cell death of macrophages were studied by phalloidin staining and electron microscopy after stimulation with an oxidative stress inducer (diamide) or tumour necrosis factor (TNF)-α. Extracellular 14-3-3η protein levels were examined by western blotting. RESULTS: Macrophages from the synovial tissue from RA, but not osteoarthritis, showed dense and widespread cytoplasmic staining for the 14-3-3η protein, co-localized with peptidylarginine deiminase 4. Swelling and membrane rupture of macrophages were induced by treatment with TNF-α, but not interleukin (IL) 6/soluble IL-6 receptor (sIL-6R). Increased MLKL phosphorylation followed by necroptosis was also induced in TNF-α-stimulated macrophages. Necrostatin-1, a necroptosis inhibitor, antagonized MLKL phosphorylation. High levels of 14-3-3η were detected in the culture supernatants of macrophages stimulated with diamide and TNF-α, but not IL-6/sIL-6R. CONCLUSIONS: Macrophages that highly express 14-3-3η undergo TNF-α-induced necroptosis with damage to the cellular structure, resulting in the secretion of 14-3-3η into the extracellular space. The current study provides a novel mechanism for 14-3-3η level increase in the RA synovial fluid.


Assuntos
Proteínas 14-3-3/metabolismo , Artrite Reumatoide , Macrófagos/metabolismo , Necroptose/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Proteínas 14-3-3/imunologia , Artrite Reumatoide/imunologia , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Humanos , Macrófagos/imunologia , Macrófagos/patologia , Líquido Sinovial/imunologia , Líquido Sinovial/metabolismo , Membrana Sinovial/imunologia , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Fator de Necrose Tumoral alfa/imunologia
14.
Am J Pathol ; 190(5): 1046-1058, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32084364

RESUMO

Cell migration-inducing hyaluronidase 1 (CEMIP), also known as hyaluronan (HA)-binding protein involved in HA depolymerization (HYBID), plays a role in HA degradation. CEMIP2, also known as transmembrane protein 2 (TMEM2), possessing a sequence similarity with HYBID, is reported as a hyaluronidase in mice. However, the expression of these molecules in osteoarthritic synovium and their involvement in HA degradation in synovial fluid (SF) from patients with knee osteoarthritis remain elusive. This study examined their expression in synovial tissue and the relationship with molecular weight of HA in SF in knee osteoarthritis patients. Quantification of mRNA demonstrated that HYBID expression is significantly (5.5-fold) higher in osteoarthritic synovium than in normal control synovium, whereas TMEM2 expression level is similar between the two groups. By immunohistochemistry, HYBID was localized mainly to CD68-negative and fibroblast-specific protein 1-positive synovial lining cells and sublining fibroblasts in osteoarthritic synovium. The mRNA expression levels of HYBID, but not TMEM2, in osteoarthritic synovium positively correlated with distribution of lower-molecular-weight HA with below 1000 kDa in SF. HA-degrading activity in osteoarthritic synovial fibroblasts was abrogated by siRNA-mediated knockdown of HYBID. Among the 12 factors examined, IL-6 significantly up-regulated the HYBID expression and HA-degrading activity in osteoarthritic synovial fibroblasts. These data suggest that HYBID overexpressed by IL-6-stimulated synovial fibroblasts is implicated in HA degradation in osteoarthritic synovium.


Assuntos
Fibroblastos/metabolismo , Ácido Hialurônico/metabolismo , Hialuronoglucosaminidase/metabolismo , Proteínas de Membrana/metabolismo , Osteoartrite do Joelho/metabolismo , Idoso , Feminino , Humanos , Masculino , Osteoartrite do Joelho/patologia , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia
15.
Ann Rheum Dis ; 79(4): 481-489, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32094158

RESUMO

OBJECTIVE: Syndecan-4 (sdc4) is a cell-anchored proteoglycan that consists of a transmembrane core protein and glucosaminoglycan (GAG) side chains. Binding of soluble factors to the GAG chains of sdc4 may result in the dimerisation of sdc4 and the initiation of downstream signalling cascades. However, the question of how sdc4 dimerisation and signalling affects the response of cells to inflammatory stimuli is unknown. METHODS: Sdc4 immunostaining was performed on rheumatoid arthritis (RA) tissue sections. Interleukin (IL)-1 induced extracellular signal-regulated kinases (ERK) phosphorylation and matrix metalloproteinase-3 production was investigated. Il-1 binding to sdc4 was investigated using immunoprecipitation. IL-1 receptor (IL1R1) staining on wild-type, sdc4 and IL1R1 knockout fibroblasts was performed in fluorescence-activated cell sorting analyses. A blocking sdc4 antibody was used to investigate sdc4 dimerisation, IL1R1 expression and the histological paw destruction in the human tumour necrosis factor-alpha transgenic mouse. RESULTS: We show that in fibroblasts, the loss of sdc4 or the antibody-mediated inhibition of sdc4 dimerisation reduces the cell surface expression of the IL-1R and regulates the sensitivity of fibroblasts to IL-1. We demonstrate that IL-1 directly binds to sdc4 and in an IL-1R-independent manner leads to its dimerisation. IL-1-induced dimerisation of sdc4 regulates caveolin vesicle-mediated trafficking of the IL1R1, which in turn determines the responsiveness to IL-1. Administration of antibodies (Ab) against the dimerisation domain of sdc4, thus, strongly reduces the expression IL1R1 on arthritic fibroblasts both in vitro and an animal model of human RA. CONCLUSION: Collectively, our data suggest that Ab that specifically inhibit sdc4 dimerisation may support anti-IL-1 strategies in diseases such as inflammatory arthritis.


Assuntos
Anticorpos Bloqueadores/farmacologia , Artrite Reumatoide/metabolismo , Receptores Tipo I de Interleucina-1/efeitos dos fármacos , Sindecana-4/antagonistas & inibidores , Animais , Artrite Reumatoide/genética , Artrite Reumatoide/patologia , Dimerização , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/metabolismo , Técnicas de Inativação de Genes , Heparitina Sulfato , Membro Posterior , Humanos , Interleucina-1/metabolismo , Interleucina-1beta/metabolismo , Sistema de Sinalização das MAP Quinases , Metaloproteinase 3 da Matriz/metabolismo , Camundongos , Camundongos Transgênicos , Células NIH 3T3 , Osteoartrite/genética , Osteoartrite/metabolismo , Osteoartrite/patologia , Fosforilação/efeitos dos fármacos , Transporte Proteico , Receptores Tipo I de Interleucina-1/metabolismo , Transdução de Sinais , Sindecana-4/genética , Sindecana-4/metabolismo , Membrana Sinovial/metabolismo , Fator de Necrose Tumoral alfa/genética
16.
Chem Biol Interact ; 319: 108984, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-32061742

RESUMO

OBJECTIVES: As one of the main active ingredients of Chinese herbal medicine Andrographis paniculate, andrographolide is used in domestic clinical treatment for respiratory infections and inflammation. This study was designed to investigate the effects of andrographolide as an antioxidant on the level of oxidative stress, neutrophil accumulation and infiltration in joints and synovial tissue of arthritis rats induced by complete freund's adjuvant. METHODS: A rat model of rheumatoid arthritis was induced by subcutaneous injection of complete Freund's adjuvant in the footpad. The model was established 14 days after induction. The treatment was performed from 14th day to 35th day with different doses of andrographolide (25, 50, 100 mg/kg) and positive control methotrexate (3 mg/kg). The effects of andrographolide on oxidative stress, neutrophil accumulation and infiltration were measured by the paw swelling, arthritis score, the hot plate test, biochemical analysis, and histology. RESULTS: The medium and high-dose andrographolide (50, 100 mg/kg) group declined the levels of tumor necrosis factor-α, interleukin-6 and CXC chemokine ligand2, articular elastase and myeloperoxidase, and increased the levels of antioxidant enzymes superoxide dismutase, catalase, and glutathione. The activity of malondialdehyde and nitrite/nitrate in andrographolide (50, 100 mg/kg) group was weakened than the model group. The degree of swelling and arthritis score of andrographolide group was lower than the model group. The results of hot plate test showed that high dose of andrographolide significantly improved the anti-injury ability of rats; Radiological and histological results showed that the joint osteoporosis, inflammatory cell infiltration, synovial hyperplasia and other phenomena in the andrographolide group were significantly improved. CONCLUSIONS: Andrographolide acts as a protective agent for the treatment of complete freund's adjuvant induced rheumatoid arthritis by inhibiting lipid peroxidation and nitrite/nitrate levels in a dose-dependent manner, enhancing antioxidant enzyme activity, reducing levels of chemokines and inflammatory factors, preventing neutrophil accumulation and infiltration.


Assuntos
Artrite Experimental/induzido quimicamente , Artrite Experimental/tratamento farmacológico , Diterpenos/farmacologia , Adjuvante de Freund/farmacologia , Inflamação/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/farmacologia , Artrite Experimental/metabolismo , Artrite Reumatoide/induzido quimicamente , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Catalase/metabolismo , Modelos Animais de Doenças , Edema/induzido quimicamente , Edema/tratamento farmacológico , Edema/metabolismo , Glutationa/metabolismo , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Articulações/efeitos dos fármacos , Articulações/metabolismo , Masculino , Metotrexato/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Ratos , Ratos Sprague-Dawley , Superóxido Dismutase/metabolismo , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
17.
PLoS One ; 15(2): e0229449, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32107493

RESUMO

Anterior cruciate ligament (ACL) transection surgery in the minipig induces post-traumatic osteoarthritis (PTOA) in a pattern similar to that seen in human patients after ACL injury. Prior studies have reported the presence of cartilage matrix-degrading proteases, such as Matrix metalloproteinase-1 (MMP-1) and A disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4), in the synovial fluid of injured or arthritic joints; however, the tissue origin of these proteases is unknown. The objective of this study was to identify transcriptional processes activated in the synovium after surgical induction of PTOA with ACL transection, and to determine if processes associated with proteolysis were enriched in the synovium after ACL transection. Unilateral ACL transection was performed in adolescent Yucatan minipigs and synovium samples were collected at 1, 5, 9, and 14 days post-injury. Transcriptome-wide gene expression levels were determined using bulk RNA-Sequencing in the surgical animals and control animals with healthy knees. The greatest number of transcripts with significant changes was observed 1 day after injury. These changes were primarily associated with cellular proliferation, consistent with measurements of increased cellularity of the synovium at the two-week time point. At five to 14 days, the expression of transcripts relating to proteolysis and cartilage development was significantly enriched. While protease inhibitor-encoding transcripts (TIMP2, TIMP3) represented the largest fraction of protease-associated transcripts in the uninjured synovium, protease-encoding transcripts (including MMP1, MMP2, ADAMTS4) predominated after surgery. Cartilage development-associated transcripts that are typically not expressed by synovial cells, such as ACAN and COMP, were enriched in the synovium following ACL-transection. The upregulation in both catabolic processes (proteolysis) and anabolic processes (cartilage development) suggests that the synovium plays a complex, balancing role in the early response to PTOA induction.


Assuntos
Cartilagem Articular/patologia , Condrogênese/genética , Osteoartrite/genética , Proteólise , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Transcriptoma , Animais , Biomarcadores/metabolismo , Cartilagem Articular/metabolismo , Masculino , Osteoartrite/patologia , Osteoartrite/cirurgia , Suínos , Porco Miniatura
18.
Artigo em Inglês | MEDLINE | ID: mdl-31945707

RESUMO

Some previous studies have demonstrated that Herba Lysimachiae (HL) has a certain protective effect on synovial lesion. But the synovial diseases HL is suitable for treating have remained unclear, as well as the mechanisms involved. To investigate the therapeutic potentials of HL in synovial diseases based on the biolabel-led research pattern. Label-free quantitative proteomics analysis was used to screen the potential biolabels responsible for the intervention of HL on synovium. The effects of HL on the joint swelling and synovial platelet aggregation in osteoarthritis model was applied to confirm the biolabels analysis results. Totally, 140 common proteins were differentially expressed after treatment with HL, out of which 23 were involved in 4 key pathways and considered as the potential biolabels responsible for the interventions of HL on synovium. Biolabels analysis showed that HL increased the levels of the proteins promoting platelet aggregation in physiological situations. The potential biolabels and their related pathways were mainly associated with the pathogenesis of osteoarthritis. In osteoarthritis model, HL inhibited the joint swelling and the overexpression of Itga2b and Itgb3 in synovium to some extent. This study reveals that HL is suitable to treat osteoarthritis. Additionally, HL may produce the dual effects on platelet aggregation in synovium.


Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Primulaceae/química , Proteoma/efeitos dos fármacos , Proteômica/métodos , Membrana Sinovial , Animais , Cromatografia Líquida , Modelos Animais de Doenças , Masculino , Osteoartrite/metabolismo , Ratos , Ratos Sprague-Dawley , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismo
19.
Med Sci Monit ; 26: e918174, 2020 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-31957742

RESUMO

BACKGROUND The aim of this study was to explore the expression of miR-140 and miR-199 in synovia of patients with knee osteoarthritis (KOA) and its correlation with the progression of this disease. We used the Kellgren and Lawrence grading (KLG) system. MATERIAL AND METHODS There were 110 patients with early (KLG <2), middle (KLG=2) and late (KLG >2) stage KOA and 60 healthy individuals (control) included in this study. RESULTS The relative expression levels of miR-140 (1.07±0.091) and miR-199 (1.03±0.110) in synovia of the control group were higher than those of KOA groups (0.511±0.130, 0.298±0.168) and the difference exhibited statistical significance (P<0.01). Expression of miR-140 in the middle and the late stage KOA groups (0.322±0.118 and 0.110±0.088 respectively) were 58.80% and 81.29% lower, respectively, compared to the early stage KOA group (0.588±0.172), which was significant (P<0.05). Expression of miR-199 in the middle and the late stage KOA groups (0.210±0.124 and 0.056±0.068 respectively) were 39.41% (P<0.05) and 83.72% (P<0.01) respectively lower than that in the early KOA group (0.344±0.147). The severity of OA was significantly negatively correlated with the expressions of miR-140 and miR-199 (r=-0.859, P<0.05; r=-0.724, P<0.001 respectively). Matrix metalloproteinase (MMP)-3 levels of the early stage, middle stage and late stage KOA groups were 1.320±0.118, 1.488±0.210, and 1.955±0.023 respectively; and IL-1ß mRNA was 1.401±0.204, 1.522±0.210, and 1.889±0.217 respectively, which were obviously higher than those in the control group (1.020±0.085), (P<0.05). CONCLUSIONS Expression levels of miR-140 and miR-199 in synovia might act as an early diagnostic marker for KOA. These expression levels might also act as indicators of OA progression to some extent.


Assuntos
Progressão da Doença , Regulação da Expressão Gênica , MicroRNAs/genética , Osteoartrite do Joelho/genética , Osteoartrite do Joelho/patologia , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Estudos de Casos e Controles , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Metaloproteinase 3 da Matriz/genética , Metaloproteinase 3 da Matriz/metabolismo , MicroRNAs/metabolismo
20.
Sci Rep ; 10(1): 780, 2020 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-31964950

RESUMO

Synovial fibroblasts (SF) were reported to produce B cell activating factor (BAFF) in response to stimulation with interferon-γ (IFN-γ) or tumor necrosis factor (TNF). However, the influence of these pro-inflammatory cytokines on other receptors/ligands of the TNF superfamily or associated cytokine receptors in SF has not been investigated yet. Here we show the differential regulation of BAFF (CD257), Fn14 (CD266), TACI (CD267), BAFF-R (CD268), BCMA (CD269), CD40 ligand (CD40L, CD154), IFN-γR (CD119), Leptin receptor (ObR, CD295), VCAM-1 (CD106) and membrane TGF-ß in isolated SF and the impact of IFN-γ/TNF co-incubation on proliferation, IL-6 and IL-8 production. In addition, the impact of differentially stimulated SF on B cell survival in co-cultures was assessed. Surface cytokines and cytokine receptors were detected by flow cytometry. Soluble cytokine receptors and cytokines were quantified by ELISA. Proliferation was assessed by cell titer blue. Murine B cell survival in fibroblast/ B cell co-cultures was determined by annexin V/propidium iodide staining and flow cytometry. IFN-γ together with TNF synergistically and significantly increased the cell surface levels of BAFF, Fn14, TACI, BAFF-R, BCMA, CD40L, ObR and IFN-γR in rheumatoid arthritis SF after 72 h incubation. Soluble BAFF was only induced by IFN-γ and inhibited by TNF. Addition of TWEAK had no influence on proliferation or IL-8 production but decreased TNF-induced IL-6 production, whereas APRIL, BAFF and leptin did not modulate TNF or TNF/IFN-γ-induced proliferation or cytokine production. Proliferation was increased by TNF and further enhanced by the addition of IFN-γ. In co-culture experiments, SF stimulated with TNF/IFN but not TNF or IFN-γ alone increased shedding of VCAM-1 and expression of membrane TGFß, which was associated with reduced survival of murine B cells. IFN-γ and TNF regulate the expression of TNF family member cytokines and associated receptors. Ligation of IFN-γR and Fn14 under pro-inflammatory conditions modulated IL-6/IL-8 production and proliferation. In B cell/SF co-cultures, the combination of TNF/IFN reduced B cell survival possibly via enhanced VCAM-1 shedding and/or increased TGF-ß production. IFN-γ is necessary for the observed effects on B cell survival and SF cytokine production and emphasizes its anti-inflammatory role in rheumatoid arthritis.


Assuntos
Artrite Reumatoide/imunologia , Fator Ativador de Células B/metabolismo , Linfócitos B/citologia , Interferon gama/farmacologia , Membrana Sinovial/citologia , Fator de Necrose Tumoral alfa/farmacologia , Idoso , Animais , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Células Cultivadas , Técnicas de Cocultura , Feminino , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Receptores de Citocinas/efeitos dos fármacos , Receptores de Citocinas/metabolismo , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismo
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