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1.
Gene ; 765: 145120, 2021 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-32896590

RESUMO

BACKGROUND: Gout is an inflammatory arthritis resulting from precipitation of monosodium urate (MSU) crystals in joints and surrounding tissues. However, the mechanism underlying high levels of uric acid inducing gouty arthritis has not been clarified. OBJECTIVE: The purpose was to investigate the role of Matrix Metalloproteinase-3 (MMP-3) in the development of gouty arthritis from hyperuricemia. METHOD: MSU crystal-induced gouty arthritis model and chondrocytes were used to evaluate changes of MMP-3 levels. Western blot, qPCR and ELISA were performed to detect MMP-3, Tissue Inhibitors of Metalloproteinase-1 (TIMP-1) and A Disintegrin and Metalloproteinase with Thrombospondin Motifs-4 (ADAMTS-4) expressions in rabbit chondrocytes. Expression of proteoglycan was determined through toluidine blue staining. Concentrations of glycosaminoglycan, Interleukin-6 (IL-6), Interleukin-1ß (IL-1ß) and Tumor Necrosis Factor-α (TNF-α) in chondrocytes were assessed via ELISA kits. Concentration of uric acid in supernate was tested by Automatic Analyzer. RESULTS: MMP-3 was significantly increased in rat serum, synovial fluid, cartilages and chondrocytes treated with high-level uric acid. Increased concentration of glycosaminoglycancould be observed in chondrocytes incubated with MMP-3, as well as the remarkable downregulation of proteoglycan expression. Furthermore, high-level uric acid contributed to the degradation of proteoglycan via the activation of MMP-3. IL-6, IL-1ß and TNF-α concentrations were increased significantly in 35 °C compared to 37 °C with MMP-3 and high-level uric acid. CONCLUSION: Our study showed that MMP-3 was enhanced by high levels of uric acid, which promoted proteoglycan degradation, and induced MSU crystallization in turn. A low temperature environment is an important factor in the development of gout.


Assuntos
Artrite Gotosa/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Proteoglicanas/metabolismo , Animais , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/patologia , Condrócitos/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Hiperuricemia/metabolismo , Interleucina-6/metabolismo , Macrófagos/metabolismo , Masculino , Metaloproteinase 3 da Matriz/fisiologia , Coelhos , Ratos , Ratos Sprague-Dawley , Líquido Sinovial/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Ácido Úrico/metabolismo
2.
PLoS One ; 15(11): e0242614, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33211763

RESUMO

The purpose of this study was to evaluate matrix metalloproteinases (MMP) -2 and MMP-3 in serum, and keratinocyte-derived chemoattractant (KC), interleukin 8 (IL-8) and monocyte chemoattractant 1 (MCP-1) in synovial fluid (SF) as stifle osteoarthritis (OA) biomarkers in dogs. Dogs with naturally occurring cranial cruciate ligament (CrCL) rupture (OA group) and healthy controls were recruited. Stifles with CrCL deficiency were surgically stabilized. Serum, SF, and synovial biopsy samples were collected from the OA group preoperatively, whereas samples were collected once from control dogs. A blinded veterinary pathologist graded synovial biopsies. Serum and SF analyses were performed using xMAP technology. General linear regression was used for statistical comparisons of serum biomarkers, and mixed linear regression for SF biomarkers and temporal concentration changes. The overall discriminative ability was quantified using area under curve (AUC). Spearman's correlation coefficient was used to assess correlations between synovial histology grades and the biomarkers. Samples from 62 dogs in the OA group and 50 controls were included. The MMP-2 and MMP-3 concentrations between the OA and control groups were not significantly different, and both with an AUC indicating a poor discriminative ability. All three SF biomarker concentrations were significantly different between the OA group and controls (P <0.05). The MCP-1 was the only biomarker showing an acceptable discriminative performance with an AUC of 0.91 (95% confidence interval: 0.83-0.98). The sum of the inflammatory infiltrate score was significantly correlated with all three SF biomarkers (P <0.01). Summed synovial stroma, and all scores combined were significantly correlated with IL-8 and MCP-1 concentrations (P <0.003), and the summed synoviocyte scores were significantly correlated with MCP-1 concentrations (P <0.001). Correlations between MCP-1 concentrations and synovial histopathologic grading and its discriminative ability suggest its potential as a synovitis biomarker in canine stifle OA associated with CrCL rupture.


Assuntos
Lesões do Ligamento Cruzado Anterior , Ligamento Cruzado Anterior , Quimiocina CCL2/metabolismo , Quimiocina CXCL1/metabolismo , Doenças do Cão , Interleucina-8/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Osteoartrite , Animais , Ligamento Cruzado Anterior/metabolismo , Ligamento Cruzado Anterior/patologia , Lesões do Ligamento Cruzado Anterior/metabolismo , Lesões do Ligamento Cruzado Anterior/patologia , Lesões do Ligamento Cruzado Anterior/veterinária , Biomarcadores/metabolismo , Doenças do Cão/metabolismo , Doenças do Cão/patologia , Cães , Feminino , Masculino , Osteoartrite/metabolismo , Osteoartrite/patologia , Osteoartrite/veterinária , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia
3.
Sci Rep ; 10(1): 13228, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32764573

RESUMO

Transglutaminase 2 (TG2), also known as tissue transglutaminase, is a calcium-dependent enzyme that has a variety of intracellular and extracellular substrates. TG2 not only increases in osteoarthritis (OA) tissue but also affects the progression of OA. However, it is still unclear how TG2 affects cartilage degradation in OA at the molecular level. Surgically induced OA lead to an increase of TG2 in the articular cartilage and growth plate, and it was dependent on TGFß1 in primary chondrocytes. The inhibition of TG2 enzymatic activity with intra-articular injection of ZDON, the peptide-based specific TG2 inhibitor, ameliorated the severity of surgically induced OA as well as the expression of MMP-3 and MMP-13. ZDON attenuated MMP-3 and MMP-13 expression in TGFß- and calcium ionophore-treated chondrocytes in a Runx2-independent manner. TG2 inhibition with ZDON suppressed canonical Wnt signaling through a reduction of ß-catenin, which was mediated by ubiquitination-dependent proteasomal degradation. In addition, TG2 activation by a calcium ionophore enhanced the phosphorylation of AMPK and FoxO3a and the nuclear translocation of FoxO3a, which was responsible for the increase in MMP-13. In conclusion, TG2 plays an important role in the pathogenesis of OA as a major catabolic mediator that affects the stability of ß-catenin and FoxO3a-mediated MMP-13 production.


Assuntos
Proteína Forkhead Box O3/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Osteoartrite/metabolismo , Transglutaminases/metabolismo , Via de Sinalização Wnt , Animais , Cálcio/metabolismo , Cartilagem Articular/metabolismo , Células Cultivadas , Proteínas de Ligação ao GTP/antagonistas & inibidores , Lâmina de Crescimento/metabolismo , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Osteoartrite/fisiopatologia , Gravidade do Paciente , Fator de Crescimento Transformador beta/metabolismo , Transglutaminases/antagonistas & inibidores , beta Catenina/metabolismo
4.
Biochim Biophys Acta Mol Basis Dis ; 1866(10): 165888, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32599142

RESUMO

Nucleus pulposus (NP) degeneration plays pivotal roles in intervertebral disc degeneration. The effect and mechanism of oxidative stress and epigenetics in NP degeneration is still unclear. We performed this study to evaluate the function of oxidative stress in NP and to explore the potential mechanism of ROS induced expression of matrix metalloproteinases (MMPs). We tested four methyltransferases, KMT2A, KMT2B, KMT2C and KMT2D in human NP samples, only KMT2D was significantly up-regulated in the severe degeneration samples. Knockdown of Kmt2d by siRNA significantly down-regulated the expression levels of catabolic enzymes including Mmp3, Mmp9 and Mmp13. Moreover, an interaction between KMT2D and ubiquitination was confirmed, and the application of H2O2 abrogated this process. Co-IP assay confirmed that H2O2 induced the phosphorylation of KMT2D to block the ubiquitination degradation, which was mainly mediated by phosphorylation of p38/MAPK. Further investigation suggested that ROS induced the alteration in levels of methylation is linked to H3K4me1 and H3K4me2, but not me3. However, usage of OICR-9429 (OICR) also suppressed the expression levels of Mmp3, Mmp9 and Mmp13. In an ex vivo model, application of OICR-9429 (OICR) also attenuated the degeneration of NP according to the H&E and Safranin-O/Fast Green staining assay, and the protein levels of MMP3, MMP9 and MMP13 were down-regulated, as well. In conclusion, we approved that oxidative stress induced ROS production promote the process of NP degeneration by enhancing KMT2D mediated transcriptional regulation of matrix degeneration related genes during NP degeneration.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Degeneração do Disco Intervertebral/patologia , Proteínas de Neoplasias/metabolismo , Núcleo Pulposo/patologia , Animais , Compostos de Bifenilo/farmacologia , Compostos de Bifenilo/uso terapêutico , Células Cultivadas , Metilação de DNA/efeitos dos fármacos , Proteínas de Ligação a DNA/genética , Di-Hidropiridinas/farmacologia , Di-Hidropiridinas/uso terapêutico , Técnicas de Silenciamento de Genes , Histonas/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Degeneração do Disco Intervertebral/tratamento farmacológico , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Proteínas de Neoplasias/genética , Estresse Oxidativo/efeitos dos fármacos , Cultura Primária de Células , Proteólise/efeitos dos fármacos , Ratos , Transdução de Sinais/efeitos dos fármacos , Ativação Transcricional , Ubiquitinação/efeitos dos fármacos , Regulação para Cima
5.
Exp Eye Res ; 196: 108064, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32439396

RESUMO

This work sought to compare aqueous angiographic segmental patterns with bead-based methods which directly visualize segmental trabecular meshwork (TM) tracer trapping. Additionally, segmental protein expression differences between aqueous angiographic-derived low- and high-outflow human TM regions were evaluated. Post-mortem human eyes (One Legacy and San Diego eye banks; n = 15) were perfused with fluorescent tracers (fluorescein [2.5%], indocyanine green [0.4%], and/or fluorescent microspheres). After angiographic imaging (Spectralis HRA+OCT; Heidelberg Engineering), peri-limbal low- and high-angiographic flow regions were marked. Aqueous angiographic segmental outflow patterns were similar to fluorescent microsphere TM trapping segmental patterns. TM was dissected from low- and high-flow areas and processed for immunofluorescence or Western blot and compared. Versican expression was relatively elevated in low-flow regions while MMP3 and collagen VI were relatively elevated in high-flow regions. TGF-ß2, thrombospondin-1, TGF-ß receptor1, and TGF-ß downstream proteins such as α-smooth muscle actin were relatively elevated in low-flow regions. Additionally, fibronectin (FN) levels were unchanged, but the EDA isoform (FN-EDA) that is associated with fibrosis was relatively elevated in low-flow regions. These results show that segmental aqueous angiographic patterns are reflective of underlying TM molecular characteristics and demonstrate increased pro-fibrotic activation in low-flow regions. Thus, we provide evidence that aqueous angiography outflow visualization, the only tracer outflow imaging method available to clinicians, is in part representative of TM biology.


Assuntos
Humor Aquoso/fisiologia , Malha Trabecular/metabolismo , Actinas/metabolismo , Angiografia , Western Blotting , Colágeno Tipo VI/metabolismo , Fibronectinas/metabolismo , Fluoresceína/metabolismo , Humanos , Pressão Intraocular , Metaloproteinase 3 da Matriz/metabolismo , Microscopia Confocal , Microscopia de Fluorescência , Microesferas , Malha Trabecular/diagnóstico por imagem , Fator de Crescimento Transformador beta/metabolismo , Versicanas/metabolismo
6.
Adv Clin Exp Med ; 29(5): 565-572, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32421262

RESUMO

BACKGROUND: Thoracic aortic aneurysm (TAA) formation is accompanied by degradation of extracellular matrix components (EMC). Numerous matrix metalloproteinases (MMPs) have been implicated in the process, but the involvement of MMP-3 remains unclear. Additionally, the changes in proteoglycan (PG) structure can alter the signal transduction pathways in TAA, though the enzymatic systems which originate them are not fully understood. OBJECTIVES: To measure MMP-3 and sulfatase levels in aneurysmal tissue, comparing them with non-aneurysmal vessels, and to investigate possible correlations with patients' serum levels in order to evaluate their potential usefulness in aiding aneurysm detection and monitoring. MATERIAL AND METHODS: The study included 74 patients (TAA: n = 42; control group: n = 32). Sulfatase activity was measured colometrically and MMP-3 levels were measured immunoenzymatically. RESULTS: Sulfatase activities were higher (p = 0.03) and MMP-3 concentrations lower (p = 0.014) in aneurysmal tissue than in normal aortic tissue. Medium-sized dilatations were associated with lower tissue MMP-3 concentrations than small dilatations (p = 0.033). No differences in sulfatase activity or MMP-3 concentration in the serum of TAA patients were observed in comparison with the controls. The serum and tissue levels of MMP-3 were correlated (r = 0.41; p < 0.001). The serum levels of MMP-3 were significantly lower in the female patients than in the male patients (p = 0.006). CONCLUSIONS: Our studies confirmed the lower MMP-3 levels in aneurysmal tissue, but the lack of a statistically confirmed reduction of MMP-3 in the blood serum seems to preclude its usefulness for diagnostic purposes. Our study points to the differences in MMP-3 behavior between TAA and abdominal aortic aneurysms. Significantly higher sulfatase activity in TAA tissue suggests a possible impact of sulfatase on signal transduction pathways involved in aneurysm formation.


Assuntos
Aneurisma da Aorta Torácica/diagnóstico , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Sulfatases/metabolismo , Aorta , Aorta Torácica , Aneurisma da Aorta Torácica/sangue , Estudos de Casos e Controles , Regulação para Baixo/fisiologia , Feminino , Humanos , Masculino , Sulfatases/genética , Regulação para Cima/fisiologia
7.
Int J Mol Sci ; 21(6)2020 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-32245213

RESUMO

Intervertebral disc (IVD) herniation and degeneration is a major source of back pain. In order to regenerate a herniated and degenerated disc, closure of the anulus fibrosus (AF) is of crucial importance. For molecular characterization of AF, genome-wide Affymetrix HG-U133plus2.0 microarrays of native AF and cultured cells were investigated. To evaluate if cells derived from degenerated AF are able to initiate gene expression of a regenerative pattern of extracellular matrix (ECM) molecules, cultivated cells were stimulated with bone morphogenetic protein 2 (BMP2), transforming growth factor ß1 (TGFß1) or tumor necrosis factor-α (TNFα) for 24 h. Comparative microarray analysis of native AF tissues showed 788 genes with a significantly different gene expression with 213 genes more highly expressed in mild and 575 genes in severe degenerated AF tissue. Mild degenerated native AF tissues showed a higher gene expression of common cartilage ECM genes, whereas severe degenerated AF tissues expressed genes known from degenerative processes, including matrix metalloproteinases (MMP) and bone associated genes. During monolayer cultivation, only 164 differentially expressed genes were found. The cells dedifferentiated and altered their gene expression profile. RTD-PCR analyses of BMP2- and TGFß1-stimulated cells from mild and severe degenerated AF tissue after 24 h showed an increased expression of cartilage associated genes. TNFα stimulation increased MMP1, 3, and 13 expression. Cells derived from mild and severe degenerated tissues could be stimulated to a comparable extent. These results give hope that regeneration of mildly but also strongly degenerated disc tissue is possible.


Assuntos
Anel Fibroso/metabolismo , Matriz Extracelular/metabolismo , Regulação da Expressão Gênica/genética , Degeneração do Disco Intervertebral/metabolismo , Deslocamento do Disco Intervertebral/metabolismo , Disco Intervertebral/metabolismo , Anel Fibroso/patologia , Proteína Morfogenética Óssea 2/farmacologia , Células Cultivadas , Matriz Extracelular/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Disco Intervertebral/patologia , Degeneração do Disco Intervertebral/genética , Degeneração do Disco Intervertebral/patologia , Deslocamento do Disco Intervertebral/genética , Deslocamento do Disco Intervertebral/patologia , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/metabolismo , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/genética , Metaloproteinase 3 da Matriz/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Regeneração/efeitos dos fármacos , Regeneração/genética , Fator de Crescimento Transformador beta1/farmacologia , Fator de Necrose Tumoral alfa/farmacologia
8.
Biochem J ; 477(9): 1701-1719, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32296833

RESUMO

To facilitate investigations of protein-protein interactions (PPIs), we developed a novel platform for quantitative mapping of protein binding specificity landscapes, which combines the multi-target screening of a mutagenesis library into high- and low-affinity populations with sophisticated next-generation sequencing analysis. Importantly, this method generates accurate models to predict affinity and specificity values for any mutation within a protein complex, and requires only a few experimental binding affinity measurements using purified proteins for calibration. We demonstrated the utility of the approach by mapping quantitative landscapes for interactions between the N-terminal domain of the tissue inhibitor of metalloproteinase 2 (N-TIMP2) and three matrix metalloproteinases (MMPs) having homologous structures but different affinities (MMP-1, MMP-3, and MMP-14). The binding landscapes for N-TIMP2/MMP-1 and N-TIMP2/MMP-3 showed the PPIs to be almost fully optimized, with most single mutations giving a loss of affinity. In contrast, the non-optimized PPI for N-TIMP2/MMP-14 was reflected in a wide range of binding affinities, where single mutations exhibited a far more attenuated effect on the PPI. Our new platform reliably and comprehensively identified not only hot- and cold-spot residues, but also specificity-switch mutations that shape target affinity and specificity. Thus, our approach provides a methodology giving an unprecedentedly rich quantitative analysis of the binding specificity landscape, which will broaden the understanding of the mechanisms and evolutionary origins of specific PPIs and facilitate the rational design of specific inhibitors for structurally similar target proteins.


Assuntos
Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 1 da Matriz/metabolismo , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas/métodos , Inibidor Tecidual de Metaloproteinase-2/genética , Biologia Computacional/métodos , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Metaloproteinase 3 da Matriz/metabolismo , Mutagênese , Mutação , Engenharia de Proteínas/métodos , Inibidor Tecidual de Metaloproteinase-2/química , Inibidor Tecidual de Metaloproteinase-2/metabolismo
9.
J Enzyme Inhib Med Chem ; 35(1): 672-681, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32156166

RESUMO

Glioblastoma multiforme (GBM) is the deadliest and the most common primary malignant brain tumour. The median survival for patients with GBM is around one year due to the nature of glioma cells to diffusely invade that make the complete surgical resection of tumours difficult. Based upon the connexin43 (Cx43) model of glioma migration we have developed a computational framework to evaluate MMP inhibition in materials relevant to GBM. Using the ilomastat Leu-Trp backbone, we have synthesised novel sulphonamides and monitored the performance of these compounds in conditioned media expressing MMP3. From the results discussed herein we demonstrate the performance of sulfonamide based MMPIs included AP-3, AP-6, and AP-7.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Glioblastoma/tratamento farmacológico , Metaloproteinase 3 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Sulfonamidas/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Inibidores de Metaloproteinases de Matriz/síntese química , Inibidores de Metaloproteinases de Matriz/química , Simulação de Acoplamento Molecular , Estrutura Molecular , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química , Células Tumorais Cultivadas
10.
J Orthop Surg Res ; 15(1): 87, 2020 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-32131874

RESUMO

BACKGROUND AND AIM: The pathophysiology of rheumatoid arthritis (RA) is characterized by excess production of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) by neutrophils and macrophages in synovium. Additionally, these cytokines promote the production of reactive oxygen species (ROS), and increased production of matrix metalloproteinases (MMPs), including MMP-3, in synoviocytes that result in joint destruction. There is limited information on how proteolytic enzymes such as MMP-3 can be regulated. We evaluated the effect of the antioxidant N-acetylcysteine (NAC) on RA and identified the relationship between the c-Jun N terminal kinase (JNK) pathway and MMP-3. We hypothesized that elucidating this relationship would lead to novel therapeutic approaches to RA treatment and management. METHODS: We investigated the effect of administering a low dose (1000 µM or less) of an antioxidant (NAC) to human rheumatoid fibroblast-like synoviocytes (MH7A cells). We also investigated the response of antioxidant genes such as nuclear factor erythroid -derived 2-related factor 2 (Nrf2) and Sequestosome 1 (p62). The influence of MMP-3 expression on the JNK pathway leading to joint destruction and the mechanisms underlying this relationship were investigated through primary dispersion culture cells collected from the synovial membranes of RA patients, consisting of rheumatoid arthritis-fibroblast-like synoviocytes (RA-FLS). RESULTS: Low-dose NAC (1000 µM) increased the expression of Nrf2 and phospho-p62 in MH7A cells, activating antioxidant genes, suppressing the expression of MMP-3, and inhibiting the phosphorylation of JNK. ROS, MMP-3 expression, and IL-6 was suppressed by administering 30 µM of SP600125 (a JNK inhibitor) in MH7A cells. Furthermore, the administration of SP600125 (30 µM) to RA-FLS suppressed MMP-3. CONCLUSIONS: We demonstrated the existence of an MMP-3 suppression mechanism that utilizes the JNK pathway in RA-FLS. We consider that the JNK pathway could be a target for future RA therapies.


Assuntos
Antioxidantes/administração & dosagem , Artrite Reumatoide/enzimologia , Sistemas de Liberação de Medicamentos/métodos , Sistema de Sinalização das MAP Quinases/fisiologia , Metaloproteinase 3 da Matriz/metabolismo , Acetilcisteína/administração & dosagem , Adulto , Idoso de 80 Anos ou mais , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/patologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Feminino , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Pessoa de Meia-Idade , Sinoviócitos/efeitos dos fármacos , Sinoviócitos/enzimologia , Resultado do Tratamento
11.
Aging (Albany NY) ; 12(3): 2246-2260, 2020 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-32023553

RESUMO

The treatment for intervertebral disc degeneration (IDD) has drawn great attention and recent studies have revealed that the p38 MAPK pathway is a potential therapeutic target for delaying the degeneration of intervertebral discs. In this study, we analyzed a nature-derived protein tyrosine kinase inhibitor, Genistein, and its function in delaying IDD in rats both in vitro and in vivo via the p38 MAPK pathway. Nucleus pulposus cells treated with Genistein showed better function compared with untreated cells. Further study revealed that Genistein could play a protective role in IDD by inhibiting phosphorylation of p38, consequently inhibiting the p38 pathway-mediated inflammatory response. The rat IDD model also demonstrated that Genistein could effectively delay the degeneration of intervertebral disc tissue. The current study reveals new biological functions of Genistein, further demonstrates the effects of the p38 MAPK pathway on intervertebral disc degeneration, and deepens our understanding of the treatment and prevention of IDD.


Assuntos
Genisteína/farmacologia , Degeneração do Disco Intervertebral/metabolismo , Núcleo Pulposo/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/efeitos dos fármacos , Agrecanas/efeitos dos fármacos , Agrecanas/genética , Animais , Sobrevivência Celular/efeitos dos fármacos , Colágeno Tipo II/efeitos dos fármacos , Colágeno Tipo II/genética , Colágeno Tipo X/efeitos dos fármacos , Colágeno Tipo X/genética , Inflamação , Interleucina-1beta/efeitos dos fármacos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-1beta/farmacologia , Disco Intervertebral/citologia , Disco Intervertebral/efeitos dos fármacos , Disco Intervertebral/metabolismo , Metaloproteinase 3 da Matriz/efeitos dos fármacos , Metaloproteinase 3 da Matriz/metabolismo , NF-kappa B/efeitos dos fármacos , NF-kappa B/metabolismo , Núcleo Pulposo/citologia , Núcleo Pulposo/metabolismo , Fosforilação , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Ratos , Transdução de Sinais , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
12.
Inflamm Res ; 69(4): 415-421, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32095874

RESUMO

OBJECTIVE: Retinol binding protein 4 (RBP4) is a member of the lipocalin family and a vitamin A carrier in the blood. More recently, RBP4 has been described as an adipokine that is involved in insulin resistance and metabolic syndrome (MetS). As obesity, MetS and some adipokines contribute to the pathogenesis of osteoarthritis (OA), we investigated RBP4 in patients with OA. MATERIALS AND METHODS: Cartilage, synovial fluid and blood samples were collected from 100 OA patients undergoing total knee replacement surgery. Primary chondrocytes and cartilage tissue were cultured to measure the RBP4 expression. The concentrations of RBP4, other adipokines (adipsin, adiponectin, leptin and resistin) and biomarkers of OA (COMP, MMP-1, MMP-3 and YKL-40) were measured by immunoassay, and gene expression was measured by next-generation RNA sequencing. RESULTS: The OA cartilage samples released RBP4 into the culture medium, and the levels correlated positively with the expression of the adipokines adipsin, adiponectin, leptin and resistin. RBP4 was the most prominently expressed of these adipokines in the OA chondrocytes, and the expression of the RBP4 receptors STRA6 (stimulated by retinoic acid gene homologue 6) and TLR4 (Toll-like receptor 4) was also detected. Within the cartilage culture medium, RBP4 showed a positive correlation with MMP-1, MMP-3 and YKL-40. RBP4 was also present in the synovial fluid from the OA patients and correlated positively with the concentrations of RBP4 found in the plasma and the cartilage culture medium. Plasma RBP4 concentrations also showed a positive correlation with MMP-3 and adipsin. CONCLUSIONS: We show here, for the first time, that RBP4 is produced within OA joints and that it is associated with increased levels of adipokines and MMPs. The results suggest a role for RBP4 in the pathogenesis of OA and as a possible target for the disease-modifying drugs for the treatment of OA.


Assuntos
Adipocinas/metabolismo , Osteoartrite/metabolismo , Proteínas Plasmáticas de Ligação ao Retinol/metabolismo , Idoso , Idoso de 80 Anos ou mais , Cartilagem Articular/metabolismo , Condrócitos/metabolismo , Feminino , Humanos , Articulação do Joelho/metabolismo , Masculino , Metaloproteinase 1 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Pessoa de Meia-Idade , Proteínas Plasmáticas de Ligação ao Retinol/genética , Líquido Sinovial/metabolismo
13.
Chem Biol Interact ; 322: 108968, 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32004530

RESUMO

Osteoarthritis (OA) is one of the most prevalent degenerative joint diseases, and the risk of developing OA significantly increases with age as well as with concomitant diseases, such as diabetes. Advanced glycation end products (AGEs) accumulate in the body over time and are associated with increased expression of various molecules involved in the pathophysiology of OA. Prostaglandin E2 (PGE2), along with its precursor cyclooxygenase (COX)-2, plays an integral role in the pathogenesis of OA and is highly upregulated in response to AGEs. The most significant event in OA is excessive degradation of the cartilage extracellular matrix, which is composed primarily of type II collagen and aggrecan. In the present study, we investigated the involvement of the receptor for glucagon-like peptide (GLP)-1 in the response of chondrocytes to insult from AGEs using the selective GLP-1 agonist dulaglutide. Firstly, our results indicate that AGEs reduced the expression of the receptor for GLP-1 (GLP-1R) in human SW1353 chondrocytes. Interestingly, we found that treatment with dulaglutide could ameliorate deterioration of the components of the articular extracellular matrix (ECM), such as type II collagen and aggrecan, induced by AGEs through downregulation of matrix metalloproteinase (MMP)-3 and MMP-13 and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4 and ADAMTS-5. We also found that dulaglutide exerted a potent inhibitory effect against the expression of several proinflammatory cytokines and chemokines closely associated with OA, as well as the production of reactive oxygen species (ROS). Finally, we showed that the effects of dulaglutide were mediated through the nuclear factor kappa-B (NF-κB) pathway. Our findings indicate that dulaglutide displayed a robust protective effect against AGEs-induced damage in chondrocytes, suggesting that it might be a possible therapeutic agent for the treatment of OA.


Assuntos
Agrecanas/metabolismo , Colágeno Tipo II/metabolismo , Peptídeos Semelhantes ao Glucagon/análogos & derivados , Produtos Finais de Glicação Avançada/metabolismo , Fragmentos Fc das Imunoglobulinas/farmacologia , Proteólise/efeitos dos fármacos , Proteínas Recombinantes de Fusão/farmacologia , Linhagem Celular , Quimiocinas/análise , Quimiocinas/metabolismo , Condrócitos/citologia , Condrócitos/metabolismo , Citocinas/análise , Citocinas/metabolismo , Regulação para Baixo/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Peptídeos Semelhantes ao Glucagon/farmacologia , Humanos , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/genética , Metaloproteinase 3 da Matriz/metabolismo , NF-kappa B/metabolismo , Osteoartrite/metabolismo , Osteoartrite/patologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
14.
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
15.
Arch Biochem Biophys ; 684: 108297, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32035098

RESUMO

Although rheumatoid arthritis (RA) has long posed a major threat to global health, the mechanisms driving the development and progression of RA remain incompletely understood. In the present study, we investigated the effects of G protein-coupled receptor 43 (GPR43/FFAR2) in various aspects of the pathogenesis of RA. To our knowledge, this is the first study to demonstrate that GPR43 is expressed on human fibroblast-like synoviocytes (FLS). Furthermore, we show that GPR43 is upregulated in FLS exposed to tumor necrosis factor-α (TNF-α). Importantly, our findings demonstrate that activation of GPR43 using its specific agonist significantly suppressed expression of the following key factors of RA: cytokines, such as interleukin-6 (IL-6), IL-8, high mobility group protein 1 (HMG-1); chemokines, such as monocyte chemoattractant protein 1 (MCP-1), intercellular adhesion molecule 1 (ICAM-1), and vascular cellular adhesion molecule 1 (VCAM-1); markers of oxidative stress, such as production of reactive oxygen species (ROS) and 4-hydroxynoneal (4-HNE); degradative enzymes, such as matrix metalloproteinase-3 (MMP-3) and MMP-13; and activation of the nuclear factor-κB (NF-κB) inflammatory signaling pathway. These results suggest a promising potential role for GPR43 as a specific target in the treatment and prevention of RA.


Assuntos
Receptores de Superfície Celular/metabolismo , Sinoviócitos/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Células A549 , Aldeídos/metabolismo , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Quimiocinas/metabolismo , Humanos , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Receptores de Superfície Celular/agonistas , Transdução de Sinais/efeitos dos fármacos , Tiazóis/farmacologia , Regulação para Cima/efeitos dos fármacos
16.
Proc Natl Acad Sci U S A ; 117(10): 5532-5541, 2020 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-32079724

RESUMO

The role of stromal fibroblasts in chronic inflammation is unfolding. In rheumatoid arthritis, leukocyte-derived cytokines TNF and IL-17A work together, activating fibroblasts to become a dominant source of the hallmark cytokine IL-6. However, IL-17A alone has minimal effect on fibroblasts. To identify key mediators of the synergistic response to TNF and IL-17A in human synovial fibroblasts, we performed time series, dose-response, and gene-silencing transcriptomics experiments. Here we show that in combination with TNF, IL-17A selectively induces a specific set of genes mediated by factors including cut-like homeobox 1 (CUX1) and IκBζ (NFKBIZ). In the promoters of CXCL1, CXCL2, and CXCL3, we found a putative CUX1-NF-κB binding motif not found elsewhere in the genome. CUX1 and NF-κB p65 mediate transcription of these genes independent of LIFR, STAT3, STAT4, and ELF3. Transcription of NFKBIZ, encoding the atypical IκB factor IκBζ, is IL-17A dose-dependent, and IκBζ only mediates the transcriptional response to TNF and IL-17A, but not to TNF alone. In fibroblasts, IL-17A response depends on CUX1 and IκBζ to engage the NF-κB complex to produce chemoattractants for neutrophil and monocyte recruitment.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Artrite Reumatoide/metabolismo , Fibroblastos/metabolismo , Proteínas de Homeodomínio/metabolismo , Inflamação/metabolismo , Interleucina-17/fisiologia , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Transcriptoma/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Artrite Reumatoide/genética , Células Cultivadas , Quimiocina CXCL1/genética , Quimiocina CXCL2/genética , Quimiocinas CXC/genética , Fatores Quimiotáticos/genética , Fibroblastos/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Humanos , Inflamação/genética , Interleucina-17/farmacologia , Interleucina-6/genética , Metaloproteinase 3 da Matriz/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/fisiologia , RNA Interferente Pequeno/genética , Proteínas Repressoras/genética , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Líquido Sinovial , Fator de Transcrição RelA/metabolismo , Fatores de Transcrição/genética , Transcriptoma/efeitos da radiação , Fator de Necrose Tumoral alfa/farmacologia
17.
Eur J Pharmacol ; 875: 172939, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31978425

RESUMO

The mechanisms driving the development and progression of Rheumatoid arthritis (RA) are complex, novel targeted therapies are gaining traction as potential methods to prevent or slow the progression of RA. Nobiletin is a derivative of citrus fruit that has been shown to attenuate the development of osteoarthritis and inhibit the expression of proinflammatory cytokines. However, the exact mechanisms by which nobiletin exerts these chondroprotective effects remain poorly understood. In the present study, we investigated the impact of nobiletin in mediating the effects of interleukin-21 (IL-21) in MH7A fibroblast-like synoviocytes (FLS), the main cell type found in the articular synovium. Firstly, we demonstrate that nobiletin (25 µM and 50 µM) reduced the expression of the IL-21 receptor by 29% and 51%, respectively, in FLS. Additionally, our findings demonstrate that nobiletin potently ameliorated IL-21-induced increased production of reactive oxygen species and 4-hydroxynonenal, increased expression of interleukin 6 (IL-6), tumor necrosis factor-α (TNF-α), and high-mobility group box 1 (HMGB1), and decreased mitochondrial membrane potential. We also demonstrate the ability of nobiletin to attenuate IL-21-induced expression of matrix metalloproteinases 3 and 13 (MMP-3, MMP-13), key degradative enzymes involved in RA-associated cartilage destruction. Finally, we show that the effects of nobiletin are mediated through the JAK1/STAT3 pathway, as nobiletin significantly reduced the phosphorylation of both JAK1 and STAT3. Taken together, our findings indicate that nobiletin may offer a safe and effective treatment against the development and progression of RA induced by the expression of IL-21 and its receptor.


Assuntos
Antioxidantes/farmacologia , Artrite Reumatoide/tratamento farmacológico , Flavonas/farmacologia , Interleucinas/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Antioxidantes/uso terapêutico , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos , Flavonas/uso terapêutico , Humanos , Subunidade alfa de Receptor de Interleucina-21/metabolismo , Interleucinas/metabolismo , Janus Quinase 1/metabolismo , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/imunologia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/imunologia , Membrana Sinovial/citologia , Membrana Sinovial/imunologia , Membrana Sinovial/patologia , Sinoviócitos
18.
Sci Rep ; 10(1): 1386, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31992771

RESUMO

High estrogen concentration leads to an inflammatory reaction in the mammary gland tissue in vivo; however, the detailed mechanism underlying its specific effects on the breast duct has not been fully clarified. We used 3D-cultured MCF-10A acini as a breast duct model and demonstrated various deleterious effects of 17-ß estradiol (E2), including the destruction of the basement membrane surrounding the acini, abnormal adhesion between cells, and cell death via apoptosis and pyroptosis. Moreover, we clarified the mechanism underlying these phenomena: E2 binds to GPER in MCF-10A cells and stimulates matrix metalloproteinase 3 (MMP-3) and interleukin-1ß (IL-1ß) secretion via JNK and p38 MAPK signaling pathways. IL-1ß activates the IL-1R1 signaling pathway and induces continuous MMP-3 and IL-1ß secretion. Collectively, our novel findings reveal an important molecular mechanism underlying the effects of E2 on the integrity of duct-like structures in vitro. Thus, E2 may act as a trigger for ductal carcinoma transition in situ.


Assuntos
Neoplasias da Mama/metabolismo , Carcinoma Intraductal não Infiltrante/metabolismo , Estradiol/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Glândulas Mamárias Humanas/metabolismo , Proteínas de Neoplasias/metabolismo , Receptores Estrogênicos/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Células A549 , Membrana Basal/metabolismo , Membrana Basal/patologia , Neoplasias da Mama/patologia , Carcinoma Intraductal não Infiltrante/patologia , Feminino , Humanos , Interleucina-1beta/metabolismo , Células MCF-7 , Glândulas Mamárias Humanas/patologia , Metaloproteinase 3 da Matriz/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.
Pediatr Rheumatol Online J ; 18(1): 6, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31941549

RESUMO

BACKGROUND: Axial SpA and Enthesitis related arthritis (ERA) patients show strong HLA-B27 association, gut dysbiosis, high toll like receptor (TLR)2 and 4 expression on monocytes, pro-inflammatory cytokine production and elevated levels of TLR4 endogenous ligands [tenascin-c (TNC) and myeloid related protein (MRP)8/14] in serum. Hence, we aimed to understand if these diseases have similar or different monocyte response. METHODS: Fifty adult axial SpA, 52 ERA patients and 25 healthy controls (HC) were enrolled. Cytokine-producing monocyte frequency before and after stimulation with lipopolysaccharide (LPS), peptidoglycan (PG), TNC or MRP8 were measured in whole blood (WB) and synovial fluid mononuclear cells (SFMC) by flow cytometry. Also, IL-6, TNF, MMP3, TNC and MRP8/14 levels were measured in unstimulated and TLR ligand stimulated WB cultures supernatant by ELISA. Finally, the mRNA expression levels of TNF and IL-6 were measured post stimulation with LPS, TNC and MRP8. RESULTS: At baseline, ERA and axial SpA patients showed similar TNF-α producing monocyte frequency which was higher than HC. MRP8 simulation led to increased TNF-α producing monocyte frequency in ERA than axial SpA. TNC and MRP8 stimulation led to similar IL-6 producing monocyte frequency in axial SpA and ERA patients. Baseline TNF and IL-6 producing monocyte frequency also modestly correlated with disease activity scores. TNF and IL-6 producing monocyte frequency increased in response to TLR stimulation in SFMC from both patients. In culture supernatants, axial SpA and ERA patients showed similar TNF production at baseline. MRP8 and TNC stimulation led to higher TNF production from ERA. Baseline IL-6 and MMP3 production was higher in ERA while TLR stimulation led to similar IL-6 and MMP3 production from axial SpA and ERA. TNC stimulation led to higher MMP3 production in ERA. mRNA expression in response to TLR stimulation was observed to be similar in axial SpA and ERA. TNC production was higher in ERA at baseline, while MRP8/14 production was higher in axial SpA than ERA post stimulation. CONCLUSION: ERA patients have similar monocyte response to exogenous and endogenous TLR ligands as patients with axial SpA. This suggests that differences between pediatric and adult-onset SpA are minimal and they may have a common pathogenesis.


Assuntos
Artrite Juvenil/imunologia , Vértebra Cervical Áxis , Monócitos/imunologia , Espondilite Anquilosante/imunologia , Adolescente , Adulto , Criança , Citocinas/metabolismo , Citometria de Fluxo , Antígeno HLA-B27/imunologia , Humanos , Interleucina-6/metabolismo , Masculino , Metaloproteinase 3 da Matriz/metabolismo , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
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