Dataset Information

Inhibition of the Expression of Matrix Metalloproteinases in Articular Chondrocytes by Resveratrol through Affecting Nuclear Factor-Kappa B Signaling Pathway.

ABSTRACT: In the present study, we tried to examine whether resveratrol regulates the expression of matrix metalloproteinases (MMPs) through affecting nuclear factor-kappa B (NF-?B) in articular chondrocytes. Rabbit articular chondrocytes were cultured in a monolayer, and reverse transcription-polymerase chain reaction (RT-PCR) was used to measure interleukin-1? (IL-1?)-induced gene expression of MMP-3, MMP-1, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4), ADAMTS-5 and type II collagen. Effect of resveratrol on IL-1?-induced secretion of MMP-3 was investigated in rabbit articular chondrocytes using western blot analysis. To elucidate the action mechanism of resveratrol, effect of resveratrol on IL-1?-induced NF-?B signaling pathway was investigated in SW1353, a human chondrosarcoma cell line, by western blot analysis. The results were as follows: (1) resveratrol inhibited the gene expression of MMP-3, MMP-1, MMP-13, ADAMTS-4, and ADAMTS-5, but increased the gene expression of type II collagen; (2) resveratrol reduced the secretion of MMP-3; (3) resveratrol inhibited IL-1?-induced activation (phosphorylation) of inhibitory kappa B kinase (IKK), and thus phosphorylation and degradation of inhibitory kappa B? (I?B?); (4) resveratrol inhibited IL-1?-induced phosphorylation and nuclear translocation of NF-?B p65. This, in turn, led to the down-regulation of gene expression of MMPs in SW1353 cells. These results suggest that resveratrol can regulate the expression of MMPs through affecting NF-?B by directly acting on articular chondrocytes.

SUBMITTER: Kang DG

PROVIDER: S-EPMC6254641 | biostudies-other | 2018 Nov

REPOSITORIES: biostudies-other

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Inhibition of the Expression of Matrix Metalloproteinases in Articular Chondrocytes by Resveratrol through Affecting Nuclear Factor-Kappa B Signaling Pathway.

Kang Dong-Geun DG Lee Hyun Jae HJ Lee Choong Jae CJ Park Jin Sung JS

Biomolecules & therapeutics 20181101 6

In the present study, we tried to examine whether resveratrol regulates the expression of matrix metalloproteinases (MMPs) through affecting nuclear factor-kappa B (NF-κB) in articular chondrocytes. Rabbit articular chondrocytes were cultured in a monolayer, and reverse transcription-polymerase chain reaction (RT-PCR) was used to measure interleukin-1β (IL-1β)-induced gene expression of MMP-3, MMP-1, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4), ADAMTS-5 an ...[more]

PMID: 30464073

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Project description:While Lymphotoxin ? (TNF-?), a product of lymphocytes, is known to play a pivotal role in inflammatory joint environment, resveratrol has been shown to possess anti-inflammatory and chondroprotective effects via activation of the histondeacetylase Sirt1. Whether TNF-? induction of inflammatory pathways in primary human chondrocytes (PCH) can be modulated by resveratrol, was investigated. Monolayer and alginate cultures of PCH were treated with TNF-?, anti-TNF-?, nicotinamide (NAM), antisense oligonucleotides against Sirt1 (Sirt1-ASO) and/or resveratrol and co-cultured with T-lymphocytes. We found that resveratrol suppressed, similar to anti-TNF-?, TNF-?-induced increased adhesiveness in an inflammatory microenvironment of T-lymphocytes and PCH. In contrast, knockdown of Sirt1 by mRNA abolished the inhibitory effects of resveratrol on the TNF-?-induced adhesiveness, suggesting the essential role of this enzyme for resveratrol-mediated anti-inflammatory signaling. Similar results were obtained in PCH stimulated with TNF-?. Sirt1-ASO, NAM or TNF-?, similar to T-lymphocytes induced inflammatory microenvironment by down-regulation of cartilage-specific proteins, Sox9, Ki67 and enhanced NF-?B-regulated gene products involved in inflammatory and degradative processes in cartilage (MMP-9/-13, COX-2, caspase-3), NF-?B activation and its translocation to the nucleus. Moreover, resveratrol reversed the TNF-?-, NAM-, T-lymphocytes-induced up-regulation of various NF-?B-regulated gene products. Down-regulation of Sirt1 by mRNA interference abrogated the effect of resveratrol on TNF-?-induced effects. Ultrastructural and cell viability assay investigations revealed that resveratrol revoked TNF-?-induced dose-dependent degradative/apoptotic morphological changes, cell viability and proliferation in PCH. Taken together, suppression of TNF-?-induced inflammatory microenvironment in PCH by resveratrol/Sirt1 might be a novel therapeutic approach for targeting inflammation during rheumatoid arthritis.

Project description:ObjectivesTo investigate the presence and functionality of oestrogen receptor alpha (ERalpha) in interleukin (IL)1beta-treated rabbit articular chondrocytes in culture, and to determine the mechanisms of 17beta oestradiol (E2) effects on IL1beta-induced inducible nitric oxide synthase (iNOS) expression.MethodsThe presence and functionality of ERalpha were investigated by immunocytochemistry and transient expression of an E2-responsive reporter construct. iNOS expression and production were determined by transient expression of a chimeric iNOS promoter-luciferase construct and protein immunoblotting. Nitric oxide (NO) production was determined by the Griess reaction. DNA-binding activities of nuclear factor-kappaB (NF-kappaB) and activated protein 1 were determined by electrophoretic mobility shift assay (EMSA)-ELISA assays. Nuclear translocation of p65 was studied by immunocytochemistry.ResultsERalpha was identified in the nucleus of chondrocytes. ERalpha efficiently transactivated a transiently expressed E2-responsive construct. On IL1beta treatment, ERalpha partially diffused from its nuclear localisation into the cytoplasm and its transactivation ability was impaired. Nevertheless, E2, tamoxifen and raloxifene efficiently inhibited IL1beta-induced NO production (-34%, -31% and -36%, respectively). E2 decreased IL1beta-induced iNOS protein expression (-40%). Transient expression of an iNOS promoter construct strongly suggested that iNOS expression was inhibited at the transcriptional level, and EMSA-ELISA assays showed that E2 reduced (-60%) the IL1beta-induced p65 DNA-binding capacity. Finally, the p65 nuclear translocation induced by IL1beta was also strongly decreased by E2.ConclusionsOur data support a reciprocal antagonism between oestrogens and IL1beta, ultimately resulting in the decrease of cytokine-dependent NO production through transcriptional inhibition of iNOS expression. This effect was associated with selective inhibition of p65 DNA binding and nuclear translocation.

Project description:Autologous chondrocyte implantation (ACI) is a promising approach to repair cartilage defects; however, the cartilage trauma-induced inflammatory environment compromises its clinical outcomes. Cell-derived decellularized extracellular matrix (DECM) has been used as a culture substrate for mesenchymal stem cells (MSCs) to improve the cell proliferation and lineage-specific differentiation. In this study, DECM deposited by synovium-derived MSCs was used as an in vitro expansion system for rabbit articular chondrocytes and the response of DECM-expanded chondrocytes to pro-inflammatory cytokines such as interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) was evaluated. Compared with those grown on tissue culture polystyrene (TCPS), the proliferation rate was significantly improved in DECM-expanded chondrocytes. TCPS- and DECM-expanded chondrocytes were isolated and induced to redifferentiation in a high-density pellet culture. DECM-expanded chondrocytes exerted a stronger resistance to 1 ng/mL of IL-1β than TCPS-expanded cells, but the production of cartilage matrix in both groups was inhibited by 5 ng/mL of IL-1β. When exposed to 1 or 5 ng/mL of TNF-α, DECM-expanded chondrocytes showed higher levels of cartilage matrix synthesis than TCPS-expanded cells. In addition, the gene expression of IL-1β- or TNF-α-induced matrix degrading enzymes (MMP3, MMP9, MMP13, and ADAMTS5) was significantly lower in DECM-expanded chondrocytes than TCPS-expanded cells. Furthermore, we found that SIRT1 inhibition by nicotinamide completely counteracted the protective effect of DECM on chondrocytes in the presence of IL-1β or TNF-α, indicating that the SIRT1 signaling pathway was involved in the DECM-mediated enhancement of anti-inflammatory properties of chondrocytes. Taken together, this work suggests that stem cell-derived DECM is a superior culture substrate for in vitro chondrocyte expansion by improving proliferation and enhancing the anti-inflammatory properties of chondrocytes. DECM-expanded chondrocytes with enhanced anti-inflammatory properties hold great potential in clinically ACI-based cartilage repair.

Dataset Information

Inhibition of the Expression of Matrix Metalloproteinases in Articular Chondrocytes by Resveratrol through Affecting Nuclear Factor-Kappa B Signaling Pathway.

Publications

Inhibition of the Expression of Matrix Metalloproteinases in Articular Chondrocytes by Resveratrol through Affecting Nuclear Factor-Kappa B Signaling Pathway.

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