RESUMEN

OBJECTIVE: This study aims to explore the effect of NONHSAT042241 on the function of rheumatoid arthritis -fibroblast-like synoviocyte (RA-FLS) and the underlying mechanisms. METHODS: RA-FLS was treated with NONHSAT042241 overexpression and NONHSAT042241 knockdown lentiviruses. Cell counting kit-8 (CCK-8) assay, colony formation assay, flow cytometry, Transwell assay, western-blot, ELISA, and qRT-PCR were used to measure the changes of cell proliferation, apoptosis, invasion, secretion of inflammatory cytokines and matrix metalloproteinases (MMPs). Fluorescent in situ hybridization (FISH) assay, RNA pull-down assay, mass spectrometry (MS) and RNA immunoprecipitation (RIP) were used to find the target proteins that bond to NONHSAT042241, and western-blot was used to detect the expression of related proteins of Wnt/ß-catenin signaling pathway. RESULTS: Overexpression of NONHSAT042241 inhibited the proliferation of RA-FLS (p < 0.05), invasion, secretion of pro-inflammatory factors (IL-1and IL-6) and MMPs (MMP-1 and MMP-3) (p < 0.05), and elevated the level of pro-apoptotic factors (Bax and cleaved caspase3), while NONHSAT042241 knockdown had the opposite effect. NONHSAT042241 can directly bind to hnRNP D, and down-regulated the expression of ß-catenin (p < 0.05), p-GSK-3ß (p < 0.05), Cyclin D1 (p < 0.05), PCNA (p < 0.05), and thus reduced the cell proliferation. CONCLUSION: NONHSAT042241 may inhibit FLS-mediated rheumatoid synovial proliferation, inflammation and aggression. The underlying mechanisms may be that NONHSAT042241 inhibits the activity of Wnt/ß-catenin signaling.

Asunto(s)

RESUMEN

BACKGROUND: Our previous studies have shown that lipoxin A4 (LXA4) can serve as a potential biomarker for assessing the efficacy of exercise therapy in knee osteoarthritis (KOA), and fibroblast-like synoviocytes (FLSs) may play a crucial role in KOA pain as well as in the progression of the pathology. OBJECTIVE: By analyzing the GSE29746 dataset and collecting synovial samples from patients with different Kellgren-Lawrence (KL) grades for validation, we focused on exploring the potential effect of LXA4 on ferroptosis in FLSs through the ESR2/LPAR3/Nrf2 axis to alleviate pain and pathological advancement in KOA. METHODS: The association between FLSs ferroptosis and chondrocyte matrix degradation was explored by cell co-culture. We overexpressed and knocked down LPAR3 in vitro to explore its potential mechanism in FLSs. A rat model of monosodium iodoacetate (MIA)-induced KOA was constructed and intervened with moderate-intensity treadmill exercise and intraperitoneal injection of PHTPP to investigate the effects of the LXA4 intracellular receptor ESR2 on exercise therapy. RESULTS: ESR2, LPAR3, and GPX4 levels in the synovium decreased with increasing KL grade. After LXA4 intervention in the co-culture system, GPX4, LPAR3, and ESR2 were upregulated in FLSs, collagen II was upregulated in chondrocytes, and MMP3 and ADAM9 were downregulated. LPAR3 overexpression upregulated the expression of GPX4, Nrf2, and SOD1 in FLSs, while downregulating the expression of MMP13 and MMP3; LPAR3 knockdown reversed these changes. Moderate-intensity platform training improved the behavioral manifestations of pain in KOA rats, whereas PHTPP treatment partially reversed the improvement in synovial and cartilage pathologies induced by platform training. CONCLUSION: LXA4 inhibited FLSs ferroptosis by activating the ESR2/LPAR3/Nrf2 axis, thereby alleviating the pain and pathological progression of KOA. This study brings a new target for the treatment of KOA and also leads to a deeper understanding of the potential mechanisms of exercise therapy for KOA.

Asunto(s)

RESUMEN

To explore whether Fengshi Liuhe Decoction (FLD) alleviates rheumatoid arthritis (RA) via the Fzd6/NF-κB signaling axis. We used real-time quantitative PCR (qPCR) and western blotting (WB) to determine the genes of the frizzled (Fzd) protein 1- Fzd protein 10 that are significantly differentially expressed between normal rat fibroblast-like synoviocyte (FLS) and collagen II-induced arthritis (CIA) rat FLS. Next, we used enzyme-linked immunosorbent assay (ELISA) to evaluate the levels of inflammatory factors in cell culture supernatant to determine the ability of FLD to ameliorate RA. Finally, we employed WB to detect the key gene expression in protein levels of the Fzd6/NF-κB signaling axis among normal rat FLS, CIA rat FLS, and FLD-treated CIA rat FLS. Our results showed that Fzd6 expression was significantly higher in CIA rat FLS at both the mRNA and protein levels than in normal rat FLS. FLD was found to downregulate Fzd6 and inflammatory factors, including COX-2, IL-8, and TNF-α, at both the mRNA and protein levels. FLD was also found to downregulate the total protein levels of Fzd6 and the NF-κB signaling pathway key gene phosphorylation of p-p65/p65 and p-IκBα/IκBα. Moreover, FLD inhibited the nuclear translocation of NF-κB p65 in CIA rat FLS. FLD can alleviate inflammation of CIA rat FLS via the Fzd6/NF-κB signaling axis.

Asunto(s)

RESUMEN

The discovery of alternative medicines with fewer adverse effects is urgently needed for rheumatoid arthritis (RA). Sophoridine (SR), the naturally occurring quinolizidine alkaloid isolated from the leguminous sophora species, has been demonstrated to possess a wide range of pharmacological activities. However, the effect of SR on RA remains unknown. In this study, the collagen-induced arthritis (CIA) rat model and tumor necrosis factor alpha (TNFα)-induced fibroblast-like synoviocytes (FLSs) were utilized to investigate the inhibitory effect of SR on RA. The anti-arthritic effect of SR was evaluated using the CIA rat model in vivo and TNFα-stimulated FLSs in vitro. Mechanistically, potential therapeutic targets and pathways of SR in RA were analyzed through drug target databases and disease databases, and validation was carried out through immunofluorescence, immunohistochemistry, and Western blot. The in vivo results revealed that SR treatment effectively ameliorated synovial inflammation and bone erosion in rats with CIA. The in vitro studies showed that SR could significantly suppress the proliferation and migration in TNFα-induced arthritic FLSs. Mechanistically, SR treatment efficiently inhibited the activation of MAPKs (JNK and p38) and NF-κB pathways in TNFα-induced arthritic FLSs. These findings were further substantiated by Immunohistochemistry results in the CIA rat. SR exerts an anti-arthritic effect in CIA rats through inhibition of the pathogenic characteristic of arthritic FLSs via suppressing NF-κB and MAPKs (JNK and p38) signaling pathways. SR may have a great potential for development as a novel therapeutic agent for RA treatment.

Asunto(s)

RESUMEN

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a critical tumor suppressor protein that regulates various biological processes such as cell proliferation, apoptosis, and inflammatory responses by controlling the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PI3K/AKT) signaling pathway. PTEN plays a crucial role in the pathogenesis of rheumatoid arthritis (RA). Loss of PTEN may contribute to survival, proliferation, and pro-inflammatory cytokine release of fibroblast-like synoviocytes (FLS). Also, persistent PI3K signaling increases myeloid cells' osteoclastic potential, enhancing localized bone destruction. Recent studies have shown that the expression of PTEN protein in the synovial lining of RA patients with aggressive FLS is minimal. Experimental upregulation of PTEN protein expression could reduce the damage caused by RA. Nonetheless, a complete comprehension of aberrant PTEN drives RA progression and its interactions with other crucial molecules remains elusive. This review is dedicated to promoting a thorough understanding of the signaling mechanisms of aberrant PTEN in RA and aims to furnish pertinent theoretical support for forthcoming endeavors in both basic and clinical research within this domain.

Asunto(s)

RESUMEN

Rheumatoid arthritis (RA) is a chronic autoimmune disorder marked by persistent synovial inflammation and joint degradation, posing challenges in the development of effective treatments. Nuciferine, an alkaloid found in lotus leaf, has shown promising anti-inflammatory and anti-tumor effects, yet its efficacy in RA treatment remains unexplored. This study investigated the antiproliferative effects of nuciferine on the MH7A cell line, a human RA-derived fibroblast-like synoviocyte, revealing its ability to inhibit cell proliferation, promote apoptosis, induce apoptosis, and cause G1/S phase arrest. Additionally, nuciferine significantly reduced the migration and invasion capabilities of MH7A cells. The therapeutic potential of nuciferine was further evaluated in a collagen-induced arthritis (CIA) rat model, where it markedly alleviated joint swelling, synovial hyperplasia, cartilage injury, and inflammatory infiltration. Nuciferine also improved collagen-induced bone erosion, decreased pro-inflammatory cytokines and serum immunoglobulins (IgG, IgG1, IgG2a), and restored the balance between T helper (Th) 17 and regulatory T cells in the spleen of CIA rats. These results indicate that nuciferine may offer therapeutic advantages for RA by decreasing the proliferation and invasiveness of FLS cells and correcting the Th17/Treg cell imbalance in CIA rats.

Asunto(s)

RESUMEN

Abnormal proliferation of aggressive fibroblast-like synoviocytes (FLS) and perpetuate synovial inflammation can inevitably accelerate the progression of rheumatoid arthritis (RA). Herein, a strategy of simultaneously promoting FLS apoptosis and inhibiting inflammation as mediated by macrophages is proposed to restore synovial homeostasis for effective RA therapy. A hyaluronic acid-based dissolvable microneedle (MN) is fabricated for transdermal delivery of dual human serum albumin (HSA)-contained biomimetic nanocomplexes to regulate RA FLS and macrophages. Upon skin insertion, dual nanocomplexes are released rapidly from the MN and accumulate in RA joint microenvironment through both passive and active targeting as mediated by HSA. Thioketal-crosslinked fluorinated polyethyleneimine 1.8 K (TKPF) was constructed to bind the plasmid encoding pro-apoptotic gene PUMA with HSA coating layer (TKPF/pPUMA@HSA, TPH). TPH nanocomplexes can upregulate PUMA through RA FLS transfection to trigger efficient apoptosis. Also, HSA nanocomplexes encapsulating the classic anti-inflammatory natural product celastrol (Cel@HSA, CH) can inhibit inflammation of macrophages through blocking NF-κB pathway activation. TPH/CH MN can deplete RA FLS and inhibit M1 macrophage activation, suppress synovial hyperplasia as well as reduce bone and cartilage erosion in a collagen-induced arthritis (CIA) mouse model, demonstrating a promising strategy for efficient RA treatment.

RESUMEN

OBJECTIVES: Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory disease that is characterized by persistent morning stiffness, joint pain, and swelling. However, there is a lack of reliable diagnostic markers and therapeutic targets that are both effective and trustworthy. METHODS: In this study, gene expression profiles (GSE89408, GSE55235, GSE55457, and GSE77298) were obtained from the Gene Expression Omnibus database. Differentially expressed necroptosis-related genes were attained from intersection of necroptosis-related gene set, differentially expressed genes, and weighted gene co-expression network analysis. The LASSO, random forest, and SVM-RFE machine learning algorithms were utilized to further screen potential diagnostic genes for RA. Immune cell infiltration was analyzed using the CIBERSORT method. The expressions of diagnostic genes were validated through quantitative real-time PCR, western blotting, immunohistochemistry, and immunofluorescence staining in synovial tissues collected from three trauma controls and three RA patients. RESULTS: Five core necroptosis-related genes (FAS, CYBB, TNFSF10, EIF2AK2, and BIRC2) were identified as potential biomarkers for RA. Two different necroptosis patterns based on these five genes were confirmed to significantly correlated with immune cells (especially macrophages). In vitro experiments showed significantly higher mRNA and protein expression levels of CYBB and EIF2AK2 in RA patients compared to normal controls, consistent with the bioinformatics analysis results. CONCLUSION: Our study identified a novel necroptosis-related subtype and five diagnostic biomarkers of RA, revealed vital roles in the development and occurrence of RA, and offered potential targets for clinical diagnosis and immunotherapy.

Asunto(s)

RESUMEN

INTRODUCTION: Statins have demonstrated chondroprotective effects by reducing inflammation and mitigating extracellular matrix degradation. However, statins are also reported to be cytotoxic to several types of cells. Early-onset osteoarthritis (OA) is characterized by synovial inflammation, which adversely affects hyaluronan (HA) production in fibroblast-like synoviocytes (FLSs). Nevertheless, the precise effects of statins on the synovium remain unclear. METHODS: This study investigated the impact of lovastatin on human FLSs, and HA secretion-related genes, signaling pathways, and production were evaluated. RESULTS: The findings revealed that high doses of lovastatin (20 or 40 µM) decreased FLS viability and increased cell death. FLS proliferation ceased when cultured in a medium containing 5 or 10 µM lovastatin. mRNA expression analysis demonstrated that lovastatin (5 and 10 µM) upregulated the gene level of hyaluronan synthase 1 (HAS1), HAS2, and proteoglycan 4 (PRG4), but not HAS3. While the expression of multidrug resistance-associated protein 5 transporter gene remained unaffected, both inward-rectifying potassium channel and acid-sensing ion channel 3 were upregulated. Western blot further confirmed that lovastatin increased the production of HAS1 and PRG4, and activated the PKC-α, ERK1/2, and p38-MAPK signaling pathways. Additionally, lovastatin elevated intracellular cAMP levels and HA production in FLSs. CONCLUSION: Lovastatin impairs cellular proliferation but enhances HA production in human FLSs.

Asunto(s)

RESUMEN

BACKGROUND: Rheumatoid arthritis (RA) is currently an autoimmune inflammatory disease with an unclear pathogenesis. Fibroblast-like synoviocytes (FLSs) have tumor-like properties, and their activation and secretion of pro-inflammatory factors are important factors in joint destruction. Wogonin (5,7-dihydroxy-8-methoxyflavone), a natural flavonoid isolated from Scutellaria baicalensis root, has been shown to have significant anti-inflammatory, anti-oxidative stress, and anti-tumor effects in a variety of diseases. However, the role of wogonin in RA has not yet been demonstrated. PURPOSE: To investigate the inhibitory effect of wogonin on the invasive behavior of fibroblast-like synoviocytes and to explore the mechanism of action of wogonin in RA. METHODS: CCK-8, EdU, cell migration and invasion, immunofluorescence staining, RT-qPCR, and protein blot analysis were used to study the inhibitory effects of wogonin on migration, invasion, and pro-inflammatory cytokine overexpression in the immortalized rheumatoid synovial cell line MH7A. The therapeutic effects of wogonin were validated in vivo using arthritis scores and histopathological evaluation of collagen-induced arthritis mice. RESULTS: Wogonin inhibited the migration and invasion of MH7A cells, reduced the production of TNF-α, IL-1ß, IL-6, MMP-3 and MMP-9, and increased the expression of IL-10. Moreover, wogonin also inhibited the myofibrillar differentiation of MH7A cells, increased the expression of E-cadherin (E-Cad) and decreased the expression of α-smooth muscle actin (α-SMA). In addition, wogonin treatment effectively ameliorated joint destruction in CIA mice. Further molecular mechanism studies showed that wogonin treatment significantly inhibited the activation of PI3K/AKT/NF-κB signaling pathway in TNF-α-induced arthritic FLSs. CONCLUSION: Wogonin effectively inhibits migration, invasion and pro-inflammatory cytokine production of RA fibroblast-like synoviocytes through the PI3K/AKT/NF-κB pathway, and thus wogonin, as a natural flavonoid, has great potential for treating RA.

RESUMEN

OBJECTIVE: Ferroptosis has been reported to play a role in rheumatoid arthritis (RA). Sulfasalazine, a common clinical treatment for ankylosing spondylitis, also exerts pathological influence on the progression of rheumatoid arthritis including the induced ferroptosis of fibroblast-like synoviocytes (FLSs), which result in the perturbated downstream signaling and the development of RA. The aim of this study was to investigate the underlying mechanism so as to provide novel insight for the treatment of RA. METHODS: CCK-8 and Western blotting were used to assess the effect of sulfasalazine on FLSs. A collagen-induced arthritis mouse model was constructed by the injection of collagen and Freund's adjuvant, and then, mice were treated with sulfasalazine from day 21 after modeling. The synovium was extracted and ferroptosis was assessed by Western blotting and immunofluorescence staining. RESULTS: The results revealed that sulfasalazine promotes ferroptosis. Compared with the control group, the expression levels of ferroptosis-related proteins such as glutathione peroxidase 4, ferritin heavy chain 1, and solute carrier family 7, member 11 (SLC7A11) were lower in the experimental group. Furthermore, deferoxamine inhibited ferroptosis induced by sulfasalazine. Sulfasalazine-promoted ferroptosis was related to a decrease in ERK1/2 and the increase of P53. CONCLUSIONS: Sulfasalazine promoted ferroptosis of FLSs in rheumatoid arthritis, and the PI3K-AKT-ERK1/2 pathway and P53-SLC7A11 pathway play an important role in this process.

Asunto(s)

RESUMEN

OBJECTIVE: Coptisine, a natural bioactive small molecular compound extracted from traditional Chinese herb Coptis chinensis, has been shown to exhibit anti-tumor effect. However, its contribution to autoimmune diseases such as rheumatoid arthritis (RA) is unknown. Here, we evaluate the effect of coptisine in controlling fibroblast-like synoviocytes (FLS)-mediated synovial proliferation and aggression in RA and further explore its underlying mechanism(s). METHODS: FLS were separated from synovial tissues obtained from patients with RA. Protein expression was measured by Western blot or immunohistochemistry. Gene expression was detected by quantitative RT-PCR. The EdU incorporation was used to measure cell proliferation. Migration and invasion were determined by Boyden chamber assay. RNA sequencing analysis was used to seek for the target of coptisine. The in vivo effect of coptisine was evaluated in collagen-induced arthritis (CIA) model. RESULTS: Treatment with coptisine reduced the proliferation, migration, and invasion, but not apoptosis of RA FLS. Mechanistically, we identified PSAT1, an enzyme that catalyzes serine/one-carbon/glycine biosynthesis, as a novel targeting gene of coptisine in RA FLS. PSAT1 expression was increased in FLS and synovial tissues from patients with RA compared to healthy control subjects. Coptisine treatment or PSAT1 knockdown reduced the TNF-α-induced phosphorylation of p38, ERK1/2, and JNK MAPK pathway. Interestingly, coptisine administration improved the severity of arthritis and reduced synovial PSAT1 expression in mice with CIA. CONCLUSIONS: Our data demonstrate that coptisine treatment suppresses aggressive and proliferative actions of RA FLS by targeting PSAT1 and sequential inhibition of phosphorylated p38, ERK1/2, and JNK MAPK pathway. Our findings suggest that coptisine might control FLS-mediated rheumatoid synovial proliferation and aggression, and be a novel potential agent for RA treatment.

Asunto(s)

RESUMEN

OBJECTIVE: Leukocyte Ig-like receptor A3 (LILRA3) is a soluble receptor belongs to the immunoglobulin superfamily. Our previous studies demonstrated that LILRA3 is a common genetic risk for multiple autoimmune diseases, including RA. Functional LILRA3 conferred increased risk of joint destruction in patients with early RA. We undertook this study to further investigate the pathological role of LILRA3 in joint inflammation of RA. METHODS: Soluble LILRA3 was measured by ELISA. LILRA3 plasmids were transfected into human fibroblast-like synoviocytes (FLSs) using electroporation. Activation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) was determined by western blots. Cytokine transcripts were quantified by real-time PCR. Migratory and invasive capacities of FLSs were evaluated using transwell migration and Matrigel invasion assays. FLS apoptosis was analysed using flow cytometry. Colocalization of LILRA3, LILRB1 and HLA-G in RA-FLSs was visualized by immunofluorescence staining. RESULTS: Soluble LILRA3 was specifically expressed in synovial fluid and serum LILRA3 was significantly increased and positively correlated with disease activity/severity in RA patients. LILRA3 induced an increased expression of IL-6, IL-8 and MMP3 in RA-FLSs. In vitro LILRA3 stimulation or overexpression promoted RA-FLS migration and invasion, and enhanced phosphorylation of ERK/JNK. Inhibition of ERK/JNK resulted in suppression of IL-6/IL-8 expression in LILRA3-stimulated RA-FLSs. LILRA3 was co-localized with its homologue LILRB1 and shared ligand HLA-G in RA-FLSs. CONCLUSION: The present study provides the first evidence that soluble LILRA3 is a novel proinflammatory mediator involved in synovial inflammation by promoting RA-FLS activation, migration and invasion, probably through the ERK/JNK signalling pathways.

Asunto(s)

RESUMEN

BACKGROUND: Salidroside (Sal) is a natural product commonly isolated from Rhodiola rosea L., which has been found to have numerous pharmacological activities (e.g., ameliorating apoptosis and inflammation, and acting as an antioxidant) in various diseases, but its concrete function in rheumatoid arthritis (RA) has not been revealed yet. Here, we aimed to explore the specific role and underlying mechanisms of Sal in RA-fibroblast-like synoviocytes (RA-FLSs). METHODS: Cell counting kit 8 (CCK-8) was used to assess the viability of normal-FLSs and RA-FLSs. Cell apoptosis in RA-FLSs was evaluated by flow cytometry. Western blotting was prepared to examine the levels of apoptosis- and signaling-related proteins. Wound-healing and Transwell assays were conducted to examine RA-FLSs migration and invasion. Enzyme-linked immunosorbent assay (ELISA) was used to assess the effect of Sal on tumor necrosis factor-alpha (TNF-α)-induced inflammation in RA-FLSs. RA animal model was established through complete Freund's adjuvant (CFA) induction, and the histopathological changes in synovial tissues of the rat model were analyzed by H&E staining. RESULTS: RA-FLSs were treated with 200 µM Sal for 24 h, and cell viability was significantly suppressed. Sal promoted RA-FLSs apoptosis. The migratory and invasive abilities of RA-FLSs were markedly inhibited by Sal. Sal incubation reduced the levels of inflammatory cytokines interleukin­8 (IL-8), IL-1ß, and IL­6 in RA-FLSs under the stimulation of TNF­α. Subsequently, Sal downregulated phosphorylated phosphatidylinositol­3 kinase (p-PI3K) and protein kinase (p-AKT) expression in RA-FLSs. After the treatment with pathway activator 740Y­P (20 µM) in RA-FLSs, the promotive effect of Sal on cell apoptosis was reversed, and inhibitory effects of it on cell viability, migration, invasion, and inflammatory response were abolished. Sal inhibited RA development in the CFA-induced rat model. CONCLUSION: Sal suppressed cell growth and inflammation in RA-FLSs by inactivating PI3K/AKT-signaling pathways.

Asunto(s)

RESUMEN

OBJECTIVE: Gadd45ß have a regulatory role in cellular inflammation, proliferation and migration. However, the role of Gadd45ß in synovial inflammation in osteoarthritis (OA) remains to be explored. This study aimed to ascertain whether Gadd45ß is involved in OA synovial inflammation. METHODS: The rat model was induced by sodium iodoacetate and the cellular model was constructed with lipopolysaccharide (LPS)-induced fibroblast-like synoviocytes (FLSs). siRNA was applied to interfere with the expression of intracellular Gadd45ß. Real-time quantitative polymerase chain reaction (RT-qPCR) and western blotting were used to detect the expression of Gadd45ß mRNA and protein. The inflammation, proliferation, and migration of OA-FLSs were detected by enzyme-linked immunosorbent assay, cell scratch assay, 5-ethynyl-2'-deoxyuridine assay, etc. The effect of downregulation of Gadd45ß on the nuclear factor-κB (NF-κB) pathway was investigated. RESULTS: Expression of Gadd45ß in OA rat synovial tissues and OA-FLSs was increased, and LPS treatment promoted cell proliferation and enhanced cell migration. Gadd45ß interference inhibited the inflammation, proliferation and migration of cells induced by LPS. LPS promoted P65 expression in the nucleus and activated the NF-κB signaling pathway, whereas si-Gadd45ß reversed this situation. CONCLUSIONS: si-Gadd45ß inhibited the inflammatory response, proliferation and migration of FLSs, and activation of the NF-κB signaling pathway, which could delay the progression of OA. Hence, it may become a potential therapeutic target for OA.

Asunto(s)

RESUMEN

OBJECT: To clarify the involvement of clock genes in the production of inflammatory mediators from RA-FLS, we examined the role of Bmal1, one of the master clock genes. METHODS: RA-FLSs were stimulated with IL-1ß (0, 20 ng/mL), IL-6 (0, 20 ng/mL), IL-17 (0, 20 ng/mL), TNF-α (0, 20 ng/mL) or IFN-γ (0, 20 ng/mL) to examine the expression of Bmal1, MMP-3, CCL2, IL-6, IL-7 and IL-15 by qPCR and immunofluorescence staining. After silencing Bmal1, RA-FLSs were stimulated with IL-1ß (0, 20 ng/mL), TNF-α (0, 20 ng/mL) or IFN-γ (0, 20 ng/mL) to examine the expressions of inflammatory mediators; MMP-3, CCL2, IL-6 and IL-15 by qPCR, ELISA and immunofluorescence staining. RESULTS: Bmal1 expressions were increased by IL-1ß, TNF-α and IFN-γ stimulations. Under stimulations with TNF-α, IL-1ß, and IFN-γ, mRNA and protein expressions of MMP-3, CCL2 and IL-6 were suppressed by siBmal1. CONCLUSION: Results indicate that Bmal1 contributes the production of MMP-3, CCL2, and IL-6 from RA-FLS, implying Bmal1 is involved in the pathogenesis of RA by regulating the inflammation.

Asunto(s)

RESUMEN

Objective @#To investigate ellect of salidroside on the function and activation of rheumatoid arthritis fibroblast-like synoviocyte(HFLS-RA)by regulating the miR-20a-5p/tissue inhibitor of metalloproteinase-2(TIMP2) axis.@*Methods@#HFlS-RA cells were used as the research object. HFlS-RA cells were separated intocontrol group, tumor necrosis factor-a (TNF-a) group, salidroside group, inhibitor NC group, miR-20a-5p inhibitor group, salidroside + mimic NC group, and salidroside + miR-20a-5p mimic group. qRT-PCR was applied to deteet the expression of miR-20a-5p in HFIS-RA cells ; enzyme-linked immunosorbent assay( ELISA) was applied todetect the levels of interleukin-18 ( lL-1β) and IL-6 in the supermatant of HFLS-RA cells: cell counting kit-8(CCK-8) method and 5-ethynyl-2 '-deoxyuridine ( EdU) staining were applied to detect HFLS-RA cell proliferation ; scratch experiment was applied to detect HilS-RA cell migration; Western blot was applied to detect the ex.pression of 'TlMP2, CyclinD1, and matrix metalloproteinase ( MMP ) -9 proteins in HFLS-RA cells; double lucifer.ase was applied to verify the relationship between miR-20a-5p and TIMP2. @*Results@#Compared with the control group, the expression of miR-20a-5p, the levels of lL-1β and IL-6, 0Dso value, EdU positive cell rate, scratchhealing rate, and the expression of CyclinDl and MMP-9 proteins in the TNF-α group increased, the expression of TlMP2 protein decreased ( P <0. 05 ) ; compared with the TNF-α group, the expression of miR-20a-5p, the levelsof lL.-1β and IL-6, OD450 value, EdU positive cell rate, scratch healing rate, and CyclinD1 and MMP-9 proteinsexpression decreased, the expression of TlMP2 protein increased in salidroside group ( P <0. 05 ); compared withthe 'T'NF -a group and inhibitor NC group, the expression of miR-20a-5p, the levels of IL-1 β and IL.-6, OD450 val-ue, EdU positive cell rate, seratch healing rate, and the expression of CyclinDl and MMP-9 proteins in the miR.20a-5p inhibitor group decreased, the expression of TlMP2 protein increased ( P <0. 05 ); compared with the sali.droside group and the salidroside + mimic NC group, the expression of miR-20a-5p, the levels of IL-1 β and IL-6 ,OD.so value, EdU positive cell rate, scratch healing rate, and the expression of CyelinD1 and MMP-9 proteins inthe salidroside + miR-20a-5p mimic group increased, the expression of TIMP2 protein decreased ( P < 0. 05 )There was a targeted regulatory relationship between miR-20a-5p and TIMP2. @*Conclusion@#Salidroside may inhibit TNF-α-induced HFS-RA cell proliferation , migration and infammatory response by regulating miR-20a-5p/TIMP2.

RESUMEN

Rheumatoid arthritis (RA) is a polygenic autoimmune disorder with an uncertain etiology, primarily impacting the joints. Moreover, the disease may manifest beyond articular involvement, leading to extra-articular manifestations. Fibroblast-like synoviocytes (FLS) are cells of mesenchymal origin that possess crucial physiological significance within the synovium, contributing to the synthesis of specific constituents found in the synovial fluid and articular cartilage. Consequently, there has been a growing focus on FLS as a potential therapeutic target in the context of RA. Recent investigations have revealed that non-coding RNAs (ncRNAs) serve as pivotal regulators of FLS function, with their dysregulated expression patterns being detected within FLS populations. NcRNAs, such as microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), assume essential functions as regulators of gene expression at both the post-transcriptional and transcriptional levels, and also serve as guiding molecules for chromatin-modifying complexes. Majority of these ncRNAs contribute to various FLS activities including metastasis, proliferation, and cytokine production. In the current work, we comprehensively review the existing literature on ncRNAs, which play pivotal roles in FLS activity and the pathogenesis of RA. Furthermore, this study provides a comprehensive summary and description of the lncRNA/circRNA-miRNA-mRNA regulatory axes in FLS activity, along with potential implications for the RA development. As well, in the final section, we illustrated that therapeutic agents including herbal medicine, and exosomes by modulating ncRNAs regulate FLS activity.

Asunto(s)

RESUMEN

BACKGROUND: The aggressive phenotype of fibroblast-like synoviocytes (FLS) has been identified as a contributing factor to the exacerbation of rheumatoid arthritis (RA) through the promotion of synovitis and cartilage damage. Regrettably, there is currently no effective therapeutic intervention available to address this issue. Recent research has shed light on the crucial regulatory role of R-spondin-2 (Rspo2) in cellular proliferation, cartilage degradation, and tumorigenesis. However, the specific impact of Rspo2 on RA remains poorly understood. We aim to investigate the function and mechanism of Rspo2 in regulating the aggressive phenotype of FLS and maintaining chondrocyte homeostasis in the context of RA. METHODS: The expression of Rspo2 in knee joint synovium and cartilage were detected in RA mice with antigen-induced arthritis (AIA) and RA patients. Recombinant mouse Rspo2 (rmRspo2), Rspo2 neutralizing antibody (Rspo2-NAb), and recombinant mouse DKK1 (rmDKK1, a potent inhibitor of Wnt signaling pathway) were used to explore the role and mechanism of Rspo2 in the progression of RA, specifically in relation to the aggressive phenotype of FLS and chondrocyte homeostasis, both in vivo and in vitro. RESULTS: We indicated that Rspo2 expression was upregulated both in synovium and articular cartilage as RA progressed in RA mice and RA patients. Increased Rspo2 upregulated the expression of leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), as the ligand for Rspo2, and ß-catenin in FLS and chondrocytes. Subsequent investigations revealed that intra-articular administration of rmRspo2 caused striking progressive synovitis and articular cartilage destruction to exacerbate RA progress in mice. Conversely, neutralization of Rspo2 or inhibition of the Wnt/ß-catenin pathway effectively alleviated experimental RA development. Moreover, Rspo2 facilitated FLS aggressive phenotype and disrupted chondrocyte homeostasis primarily through activating Wnt/ß-catenin pathway, which were effectively alleviated by Rspo2-NAb or rmDKK1. CONCLUSIONS: Our data confirmed a critical role of Rspo2 in enhancing the aggressive phenotype of FLS and disrupting chondrocyte homeostasis through the Wnt/ß-catenin pathway in the context of RA. Furthermore, the results indicated that intra-articular administration of Rspo2 neutralizing antibody or recombinant DKK1 might represent a promising therapeutic strategy for the treatment of RA.

Asunto(s)

RESUMEN

Carboxyamidotriazole (CAI) was initially considered a non-cytotoxic anticancer agent. However, recently, pronounced anti-inflammatory properties of CAI have been reported. Rheumatoid arthritis (RA) is an autoimmune inflammatory disease characterized by aberrant activation of signaling pathways. Therefore, this study explored the therapeutic effects and potential mechanism of action of CAI on RA in the adjuvant arthritis (AA) model. The results showed that CAI reduced the severity of arthritis in AA rats as demonstrated by inhibited hind paw swelling, reduced body weight, and decreased infiltration of joint pathological inflammatory cells. Importantly, pathological scoring of new blood vessels and immunohistochemical assays revealed that CAI inhibited pannus formation. CAI decreased the expression of pro-angiogenic growth factors, such as vascular epidermal growth factor, basic fibroblast growth factor, and metalloproteinases (MMPs), namely, MMP-1 and MMP-3 in the synovium of AA rats. Furthermore, CAI significantly reduced the increased levels of phosphorylated p38, c-Jun N-terminal kinase (JNK)1/2, and extracellular signal-regulated kinase (ERK)1/2 proteins in AA rats. In addition, the proliferation of fibroblast-like synoviocytes (FLS) was downregulated by CAI both in vivo and in vitro. In conclusion, this investigation illustrates the therapeutic effect of CAI on synovitis and erosion of articular cartilage in RA. Furthermore, the mechanism might involve inhibition of aberrantly activated mitogen-activated protein kinase signaling, as well as a decrease in pro-angiogenic factors, MMP expression, and FLS proliferation.