scholarly journals Lunasin Inhibits Cell Proliferation via Apoptosis and Reduces the Production of Proinflammatory Cytokines in Cultured Rheumatoid Arthritis Synovial Fibroblasts

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Shaohui Jia ◽  
Shufang Zhang ◽  
Hong Yuan ◽  
Ning Chen
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Proinflammatory Cytokines ◽  
Synovial Cell ◽  
Synovial Fibroblasts ◽  
Nutritional Supplement ◽  
Luciferase Reporter ◽  
Drug Candidate ◽  
Matrix Metalloproteinase 3 ◽  
Anticancer Properties

Lunasin, a peptide with 43 amino acid residues and initially isolated and identified in soybean cotyledon, has gained extensive attention due to its anti-inflammatory and anticancer properties. However, its treatment efficacy on rheumatoid arthritis (RA) and corresponding mechanisms have not been reported. Herein, the synovial fibroblasts harvested and isolated from patients with RA were treated with lunasin at various concentrations to examine the proliferation, apoptosis status, and corresponding cell cycle of cultured RA synovial fibroblasts. Meanwhile, the underlying mechanisms of lunasin for RA treatment are explored through Western blot, real-time PCR, ELISA, and luciferase reporter assays. Lunasin significantly inhibited the proliferation and induced the apoptosis of cultured RA synovial fibroblasts. In addition, lunasin reduced the production of interleukin-6 (IL-6), IL-8, and matrix metalloproteinase-3 (MMP-3) and suppressed the activation of NF-κB in cultured RA synovial fibroblasts but did not reveal obvious modulation on the secretion and gene expression of MMP-1. Therefore, lunasin will have promising potential as a novel nutritional supplement or drug candidate for RA due to its potency of suppressing synovial cell proliferation and decreasing the production of proinflammatory cytokines and MMPs in synovial cells.

MiR-544a Regulates Synovial Cell Proliferation, Invasion, Migration and Apoptosis by Targeting E2F2

2019 ◽  
Vol 9 (8) ◽  
pp. 1065-1072
Author(s):  
Lingxiao Pan ◽  
Kaifeng Gan
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Western Blot ◽  
Target Genes ◽  
Synovial Cell ◽  
New Drugs ◽  
Live Cells ◽  
Luciferase Reporter ◽  
Normal Fibroblast ◽  
Fibroblast Like Synoviocytes

In the present study, we aimed to explore the role of MiR-544a in inhibiting the proliferation and promoting apoptosis of rheumatoid arthritis synovial cells. Detected the expression of MiR-544 and E2F2 in Normal Fibroblast-like synoviocytes (NFLs) and Rheumatoid arthritis Fibroblast-like synoviocytes (RA-FLSs) by RT-qPCR and western blot. The target genes of MiR-544a were predicted by miRDB and were verified with the luciferase reporter system and western blot. The E2F2 overexpression plasmid was constructed and transfected into RAFLS. Cell apoptosis were detected by CCK-8. Wound healing and transwell assay were employed to detect cell invasion ability. The expression of E2F2, MMP2 and MMP9 were measured by Western blot. Further, the level of apoptosis and live cells were detected by flow cytometry DAPI staining, respectively. The expression of cell proliferation, apoptosis and migration-related proteins were detected by Western blot. The expression of miR-544 in RA-FLSs is significantly lower than that in NFLs and E2F2 is increased in RA-FLSs. E2F2 is one of the target genes of miR-544a by miRDB detection and luciferase activity assay. Overexpression of miR-544 inhibits cell proliferation, invasion, migration and apoptosis by targeting E2F2 in RA-FLSs. MiR-544a regulates synovial cell proliferation, invasion, migration and apoptosis by targeting E2F2, which is expected to be an attractive target for the development of new drugs for the treatment of rheumatoid arthritis.

scholarly journals Multiomics landscape of synovial fibroblasts in rheumatoid arthritis

2021 ◽  
Vol 41 (1) ◽  
Author(s):  
Haruka Tsuchiya ◽  
Mineto Ota ◽  
Keishi Fujio
Keyword(s):  
Rheumatoid Arthritis ◽  
Genome Analysis ◽  
Association Studies ◽  
Therapeutic Targets ◽  
Synovial Cell ◽  
Synovial Fibroblasts ◽  
Mesenchymal Cells ◽  
Integrated Analysis ◽  
Main Body ◽  
Genome Wide Association Studies

Abstract Background Rheumatoid arthritis (RA) is an autoimmune disease characterized by tumor-like hyperplasia and inflammation of the synovium, which causes synovial cell invasion into the bone and cartilage. In RA pathogenesis, various molecules in effector cells (i.e., immune cells and mesenchymal cells) are dysregulated by genetic and environmental factors. Synovial fibroblasts (SFs), the most abundant resident mesenchymal cells in the synovium, are the major local effectors of the destructive joint inflammation and exert their effects through the pathogenic production of molecules such as interleukin-6. Main body To date, more than 100 RA susceptibility loci have been identified in genome-wide association studies (GWASs), and finding novel therapeutic targets utilizing genome analysis is considered a promising approach because some candidate causal genes identified by GWASs have previously been established as therapeutic targets. For further exploration of RA-responsible cells and cell type-specific therapeutic targets, integrated analysis (or functional genome analysis) of the genome and intermediate traits (e.g., transcriptome and epigenome) is crucial. Conclusion This review builds on the existing knowledge regarding the epigenomic abnormalities in RASFs and discusses the recent advances in single-cell analysis, highlighting the prospects of SFs as targets for safer and more effective therapies against RA.

MicroRNA-16-5p Promoted Fibroblast-Like Synoviocyte Proliferation and Suppressed Apoptosis via Targeting Suppressor of Cytokine Signaling 6 (SOCS6) in Human Rheumatoid Arthritis

2021 ◽  
Vol 11 (9) ◽  
pp. 1744-1751
Author(s):  
Deqian Meng ◽  
Wenyou Pan ◽  
Ju Li
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Cytokine Signaling ◽  
Human Rheumatoid Arthritis ◽  
Reporter Assay ◽  
Functional Roles ◽  
Proliferation And Apoptosis ◽  
Fibroblast Like Synoviocytes

Accumulating evidence have indicated that MicroRNAs (miRNAs) are key regulators in human rheumatoid arthritis (RA). The aim of this study was to explore the functional roles of miR-16-5p in proliferation, inflammation, and apoptosis of fibroblast-like synoviocytes (FLS). The expression of miR-16-5p and SOCS6 in FLA was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation and apoptosis were measured by CCK-8 assay and flow cytometry, respectively. Luciferase reporter assay was used to verify the direct target of miR-16-5p. Western blot analysis was performed to analysis the levels of SOCS6, Bcl-2, Bax and cleaved caspase 3. miR-16-5p expression was significantly upregulated while SOCS6 level was decreased in RA-FLS compared with normal FLS. In addition, luciferase reporter assay confirmed that SOCS6 was the target of miR-16-5p. Silencing of miR-16-5p inhibited cell proliferation, releases of TNF-α, IL-1β, IL-6 and IL-8, and induced the apoptosis. The effects of miR-16-5p silencing on RA-FLS were reversed by downregulation of SOCS6. In summary, knockdown of miR-16-5p could suppress cell proliferation and accelerate the apoptosis of RA-FLS through targeting SOCS6, which may provide a potential therapeutic target for patients with RA.

miR-216a-5p Suppresses the Proliferation, Invasion and Migration of Fibroblast-Like Synoviocyte in Rheumatoid Arthritis by Targeting Zinc Finger and BTB Domain-Containing Protein 2 (ZBTB2)

2021 ◽  
Vol 11 (5) ◽  
pp. 896-902
Author(s):  
Jinwei Zhao ◽  
Ling Li
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Zinc Finger ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Healthy Controls ◽  
Invasion And Migration ◽  
Btb Domain ◽  
And Migration

MicroRNAs have been reported to be associated with the initiation and progression of rheumatoid arthritis (RA). miR-216a-5p, one of the miRNAs, is involved in cancer cell proliferation, invasion and migration. However, the role of miR-216a-5p in RA remains to be explored. The expressions of miR-216a-5p and zinc finger and BTB domain-containing protein 2 (ZBTB2) in fibroblast-like synoviocytes (FLS) of RA or healthy controls were detected by qRT-PCR and western blot analysis. Transfection of overexpressed and silenced miR-216a-5p were performed to explore the functional role of miR-216a-5p in RA-FLS. Cell Counting Kit-8 (CCK-8) assay and transwell assay were employed to assess cell proliferation and cell invasion, respectively. Moreover, luciferase reporter assay was executed to verify the combination of miR-216a-5p and ZBTB2. The results showed that miR-216a-5p expression in RA-FLS was downregulated than healthy controls. Overexpres-sion of miR-216a-5p inhibited RA-FLS cell proliferation, invasion and migration, while miR-216a-5p silencing revealed the opposite results. In addition, ZBTB2 was identified to be a direct target of miR-216a-5p in RA-FLS and its expression was higher than that in healthy controls. Rescue experiments revealed that ZBTB2 overexpression reversed the effects of miR-216a-5p on the proliferation, invasion and migration of RA-FLS. These data indicated the suppressive role of miR-216a-5p in RA-FLS via the regulation of ZBTB2, suggesting that miR-216a-5p and ZBTB2 may be the new targets for the treatment of RA.

scholarly journals Shikonin Inhibits Inflammatory Response in Rheumatoid Arthritis Synovial Fibroblasts via lncRNA-NR024118

2015 ◽  
Vol 2015 ◽  
pp. 1-12 ◽  
Author(s):  
Ke-ya Yang ◽  
Dong-liang Chen
Keyword(s):  
Rheumatoid Arthritis ◽  
Inflammatory Response ◽  
Proinflammatory Cytokines ◽  
Synovial Fibroblasts ◽  
Bone Lesions ◽  
Socs3 Expression ◽  
Major Chemical ◽  
Cellular And Humoral Immunity ◽  
The Impact ◽  
Socs3 Protein

Background. Shikonin is a major chemical component of zicao that possesses anti-inflammatory properties and the ability to mediate cellular and humoral immunity, especially in rheumatoid arthritis (RA). We investigated the impact of shikonin on inflammatory response in RA synovial fibroblasts using the CAIA model.Methods. Severe polyarticular arthritis was induced in Balb/c female mice. Expressions of lncRNA-NR024118, SOCS3, proinflammatory cytokines, and MMPs were evaluated using RT-RCR. Histone acetylation and SOCS3 protein expression were assessed by ChIP assay and western blot, respectively.Results. Mice treated with shikonin showed an abrogation of soft tissue and bone lesions. Shikonin remarkably enhanced the expression of NR024118 and SOCS3 and suppressed the secretion and expression of IL-6, IL-8, and MMPs. Proliferation of cultured RA synovial fibroblasts in the presence of IL-1βwas also significantly inhibited by shikonin. Moreover, shikonin dose-dependently increased acetylation of histone H3 at the promoter of NR024118. Finally, NR024118 overexpression and interference significantly changed SOCS3 expression and NR024118 interference could reverse regulation of shikonin on SOCS3, proinflammatory cytokines, and MMPs expression level in MH7A cells.Conclusion. Our results reveal that, in the CAIA mouse model of RA, shikonin has disease modifying activity that is attributable to the inhibition of inflammatory response via lncRNA-NR024118.

scholarly journals Uridine Diphosphate (UDP) Promotes Rheumatoid Arthritis Through P2Y6 Activation

2020 ◽  
Author(s):  
Hongxing Wang ◽  
Hui Wu ◽  
Kehua Fang ◽  
Xiaotian Chang
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Real Time ◽  
Proinflammatory Cytokines ◽  
Uridine Diphosphate ◽  
Synovial Fluids ◽  
Synovial Tissues ◽  
And Migration ◽  
G Protein Coupled ◽  
Fibroblast Like Synoviocytes

Abstract Background: Uridine diphosphate (UDP) is an extracellular nucleotide signaling molecule implicated in diverse biological processes via specific activation of pyrimidinergic receptor P2Y, G Protein-Coupled, 6 (P2Y6). There is very little knowledge about the function and mechanism of UDP in rheumatoid arthritis (RA).Methods: This study used a quasi-targeted liquid chromatography-mass spectrometry (LC-MS) approach to investigate the unique expression of metabolites in RA synovial fluids (SF) (n = 10) with samples from osteoarthritis (OA) as controls (n = 10). RA fibroblast-like synoviocytes (FLSs) were collected from synovial tissues (n = 5) and cultured with UDP or MRS2578, a P2Y6 antagonist, and FLSs from OA were used as controls (n = 5). Rats with collagen-induced arthritis (CIA) were injected with UDP, MRS2578 or both (n = 9 for each group). P2Y6 expression was examined using real-time PCR, Western blotting and immunohistochemistry. Cell proliferation, apoptosis and migration of RA FLSs were measured using CCK-8 assay, real-time cell analysis, flow cytometry, wound healing assay and Transwell assay, respectively. The UDP levels in the culture medium, synovial fluid (n = 36) and peripheral blood (n = 36) of RA and CIA rats were measured using a Transcreener UDP Assay. Levels of proinflammatory cytokines were measured using a flow assay. Interleukin-6 (IL-6) levels were measured using ELISA and flowResults：LC-MS analysis detected significantly increased UDP levels in RA SF compared with OA SF, and the level was positively correlated with anticyclic citrullinated peptide (anti-CCP) and rheumatoid factor (RF) levels in RA. The increased UDP concentration was verified in the blood and synovial fluids of RA patients compared with samples from OA patients and healthy volunteers, respectively. UDP stimulated cell proliferation, migration and IL-6 secretion in RA FLSs and inhibited their apoptosis in culture, and MRS2578 inhibited these effects of UDP. UDP injection accelerated CIA and stimulated IL-6 production rather than other proinflammatory cytokines in the rat model, but simultaneous injection of MRS2578 suppressed these effects and alleviated CIA. P2Y6 expression was increased in RA and CIA synovial tissues.Conclusion: These results suggest that UDP is highly expressed in RA and stimulates RA pathogenesis by promoting P2Y6 activities to increase IL-6 production.

scholarly journals Lnc RNA ZFAS1 regulates the proliferation, apoptosis, inflammatory response and autophagy of fibroblast-like synoviocytes via miR-2682-5p/ADAMTS9 axis in rheumatoid arthritis

2020 ◽  
Vol 40 (8) ◽  
Author(s):  
Shanshan Yang ◽  
Wei Yin ◽  
Yan Ding ◽  
Fan Liu
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Inflammatory Response ◽  
Large Subunit ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cell Behaviors ◽  
Synovial Tissues ◽  
Related Proteins ◽  
Fibroblast Like Synoviocytes

Abstract Backgrounds: Rheumatoid arthritis (RA) is a frequent autoimmune disease. Emerging evidence indicated that ZNFX1 antisense RNA1 (ZFAS1) participates in the physiological and pathological processes in RA. However, knowledge of ZFAS1 in RA is limited, the potential work pathway of ZFAS1 needs to be further investigated. Methods: Levels of ZFAS1, microRNA (miR)-2682-5p, and ADAM metallopeptidase with thrombospondin type 1 motif 9 (ADAMTS9) were estimated using quantitative real-time polymerase chain reaction (qRT-PCR) assay. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was conducted to explore the ability of cell proliferation in fibroblast-like synoviocytes (FLS-RA). Cell apoptosis was measured via flow cytometry. Also, levels of ADAMTS9, apoptosis-related proteins, cleaved-caspase-3 (active large subunit), and autophagy-related proteins were identified adopting Western blot. Enzyme-linked immunosorbent assay (ELISA) was performed to determine the productions of inflammatory cytokines. Beside, the interrelation between miR-2682-5p and ZFAS1 or ADAMTS9 was verified utilizing dual-luciferase reporter assay. Results: High levels of ZFAS1 and ADAMTS9, and a low level of miR-2682-5p were observed in RA synovial tissues and FLS-RA. Knockdown of ZFAS1 led to the curbs of cell proliferation, inflammation, autophagy, and boost apoptosis in FLS-RA, while these effects were abolished via regaining miR-2682-5p inhibition. Additionally, the influence of miR-2682-5p on cell phenotypes and inflammatory response were eliminated by ADAMTS9 up-regulation in FLS-RA. Mechanically, ZFAS1 exerted its role through miR-2682-5p/ADAMTS9 axis in RA. Conclusion: ZFAS1/miR-2682-5p/ADAMTS9 axis could modulate the cell behaviors, inflammatory response in FLS-RA, might provide a potential therapeutic target for RA treatment.

scholarly journals Uridine Diphosphate Promotes Rheumatoid Arthritis Through P2Y6 Activation

2021 ◽  
Vol 12 ◽  
Author(s):  
Hongxing Wang ◽  
Hui Wu ◽  
Kehua Fang ◽  
Xiaotian Chang
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Real Time ◽  
Proinflammatory Cytokines ◽  
Uridine Diphosphate ◽  
Synovial Fluids ◽  
Synovial Tissues ◽  
And Migration ◽  
G Protein Coupled ◽  
Fibroblast Like Synoviocytes

BACKGROUND: Uridine diphosphate (UDP) is an extracellular nucleotide signaling molecule implicated in diverse biological processes via specific activation of pyrimidinergic receptor P2Y, G Protein-Coupled, 6 (P2Y6). There is very little knowledge about the function and mechanism of UDP in rheumatoid arthritis (RA).METHODS: This study used a quasi-targeted liquid chromatography-mass spectrometry (LC-MS) approach to investigate the unique expression of metabolites in RA synovial fluids (SF) (n = 10) with samples from osteoarthritis (OA) as controls (n = 10). RA fibroblast-like synoviocytes (FLSs) were collected from synovial tissues (n = 5) and cultured with UDP or MRS2578, a P2Y6 antagonist, and FLSs from OA were used as controls (n = 5). Rats with collagen-induced arthritis (CIA) were injected with UDP, MRS2578 or both (n = 9 for each group). P2Y6 expression was examined using real-time PCR, Western blotting and immunohistochemistry. Cell proliferation, apoptosis and migration of RA FLSs were measured using CCK-8 assay, real-time cell analysis, flow cytometry, wound healing assay and Transwell assay, respectively. The UDP levels in the culture medium, synovial fluid (n = 36) and peripheral blood (n = 36) of RA and CIA rats were measured using a Transcreener UDP Assay. Levels of proinflammatory cytokines were measured using a flow assay. Interleukin-6 (IL-6) levels were measured using ELISA and flow.RESULTS: LC-MS analysis detected significantly increased UDP levels in RA SF compared with OA SF, and the level was positively correlated with anticyclic citrullinated peptide (anti-CCP) and rheumatoid factor (RF)levels in RA. The increased UDP concentration was verified in the blood and synovial fluids of RA patients compared with samples from OA patients and healthy volunteers, respectively. UDP stimulated cell proliferation, migration and IL-6 secretion in RA FLSs and inhibited their apoptosis in culture, and MRS2578 inhibited these effects of UDP. UDP injection accelerated CIA and stimulated IL-6 production rather than other proinflammatory cytokines in the rat model, but simultaneous injection of MRS2578 suppressed these effects and alleviated CIA. P2Y6 expression was increased in RA and CIA synovial tissues.CONCLUSION: These results suggest that UDP is highly expressed in RA and stimulates RA pathogenesis by promoting P2Y6 activities to increase IL-6 production.

MiR-1246 Involves in the Proliferation, Apoptosis and Inflammation of Rheumatoid Arthritis Fibroblast-Like Synoviocytes by Targeting Axis Inhibition Protein 2 and Glycogen Synthetase Kinase-3β via Wnt/β-Catenin Pathway in Rheumatoid Arthritis

2021 ◽  
Vol 11 (11) ◽  
pp. 2246-2253
Author(s):  
Siyin Guo ◽  
Shichang Hu ◽  
Guoyuan Zhao ◽  
Weijian Hu ◽  
Yiling Liao ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Bioinformatic Analysis ◽  
Vital Role ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Regulatory Effects ◽  
Apoptosis And Inflammation

Background and Objective: Accumulating evidence supports that fibroblast-like synovial cells (FLS) plays a vital role in the pathogenesis of rheumatoid arthritis (RA). miR-1246 has been reported to be up-regulated in the sera of RA patients. The purpose of the present research was to determine the potential role of miR-1246 in RA and the underlying mechanisms. Methods: miR-1246, GSK3β and AXIN2 levels were determined by RT-qPCR. By exploiting miR-1246 inhibitor, shRNA-GSK3β and shRNA-AXIN2, we detected the effects of miR-1246, GSK3β and AXIN2 on cell proliferation, apoptosis and inflammation in RA-FLSs. Bioinformatics analysis predicted the binding sites of miR-1246 to AXIN2, miR-1246 to GSK3β. Moreover, luciferase reporter assay verified the binding relationship. Results: In comparison with normal human FLSs, higher levels of miR-1246 existed in RA-FLSs. Downregulation of miR-1246 inhibited cell proliferation, inflammation and β-catenin expression, and promoted cell apoptosis. Furthermore, bioinformatic analysis and dual-luciferase reporter assay identified AXIN2 or GSK3β as a target gene of miR-1246. Downregulation of miR-1246 enhanced AXIN2 and GSK3β expression in RA-FLSs. Besides, co-transfection with shRNA-GSK3β or shRNA-AXIN2 partly reversed the regulatory effects of miR-1246 inhibitor in RA-FLSs. Conclusions: Collectively, our in vitro experiments proved that downregulation of miR-1246 might alleviate RA pathogenesis by targeting AXIN2 and GSK3β via Wnt/β-Catenin axis.

Effect of MiR-21 on Regulating the Proliferation of Synovial Fibroblasts in Rheumatoid Arthritis

2020 ◽  
Vol 10 (7) ◽  
pp. 1052-1058
Author(s):  
Zhichao Li ◽  
Jin Wang ◽  
Jing Yang
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Cell Apoptosis ◽  
Mrna Level ◽  
Synovial Fibroblasts ◽  
Control Group ◽  
Spearman Correlation ◽  
Qrt Pcr ◽  
Relationship Of ◽  
The Relationship

Background: This study investigated whether miR-21 regulates the expression of STAT3 and affects FLS cells. Methods: MiR-21 and STAT3 mRNA level was assessed by qRT-PCR and STAT3 and p-STAT3 level was evaluated by Western blot. Spearman correlation was used to analyze the relationship between miR-21 and STAT3 mRNA expression in synovial tissue of RA patients. FLS cells were treated with IL-17A, and the cells without treatment was included as the control group. Under IL-17A treatment, FLS cells were divided into 2 groups: miR-NC group and miR-21 mimic group. MiR-21, STAT3, p-STAT3 expression were detected and compared. EdU staining was used to detect cell proliferation and flow cytometry was used to measure cell apoptosis. Results: There was a target relationship of miR-21 with STAT3 mRNA. IL-17A treatment significantly downregulated miR-21 in FLS cells, upregulated STAT3 and p-STAT3 and enhanced cell proliferation. Transfection of miR21 mimic significantly downregulated STAT3 and p-STAT3 in FLS cells, reduced cell proliferation and increased cell apoptosis. Conclusion: MiR-21 overexpression down-regulates STAT3, inhibits FLS cell proliferation and promotes apoptosis, indicating that it might be a therapeutic target for treating RA.

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