Advances in Noninvasive Molecular Imaging Probes for Liver Fibrosis Diagnosis.

Liver fibrosis is a wound-healing response to chronic liver injury, which may lead to cirrhosis and cancer. Early-stage fibrosis is reversible, and it is difficult to precisely diagnose with conventional imaging modalities such as magnetic resonance imaging, positron emission tomography, single-photon emission computed tomography, and ultrasound imaging. In contrast, probe-assisted molecular imaging offers a promising noninvasive approach to visualize early fibrosis changes in vivo, thus facilitating early diagnosis and staging liver fibrosis, and even monitoring of the treatment response. Here, the most recent progress in molecular imaging technologies for liver fibrosis is updated. We start by illustrating pathogenesis for liver fibrosis, which includes capillarization of liver sinusoidal endothelial cells, cellular and molecular processes involved in inflammation and fibrogenesis, as well as processes of collagen synthesis, oxidation, and cross-linking. Furthermore, the biological targets used in molecular imaging of liver fibrosis are summarized, which are composed of receptors on hepatic stellate cells, macrophages, and even liver collagen. Notably, the focus is on insights into the advances in imaging modalities developed for liver fibrosis diagnosis and the update in the corresponding contrast agents. In addition, challenges and opportunities for future research and clinical translation of the molecular imaging modalities and the contrast agents are pointed out. We hope that this review would serve as a guide for scientists and students who are interested in liver fibrosis imaging and treatment, and as well expedite the translation of molecular imaging technologies from bench to bedside.

A Novel Strain of Bacillus cereus with a Strong Antagonistic Effect Specific to Sclerotinia and Its Genomic and Transcriptomic Analysis.

Sclerotinia, which is caused by Sclerotinia sclerotiorum, is a severe disease of oilseed rape, which is an important oil crop worldwide. In this study, we isolated a novel strain of Bacillus cereus, named B. cereus HF10, from the rhizosphere soil of the reed on the seaside of Yagzhou Bay, Sanya city, Hainan Province, China. HF10 exhibited a significant antagonistic effect on Sclerotinia sclerotiorum, with an inhibition rate of 79%, and to other species in Sclerotinia, but no antagonistic effect was found on various other fungi or bacteria. HF10 had an 82.3% inhibitory effect on the S. sclerotiorum infection of oilseed rape leaves and a 71.7% control effect on Sclerotinia infection in oilseed rape based on in vitro and in vivo experiments, respectively. The genomics and transcriptomics of HF10 and its loss of the antifungal function mutant Y11 were analyzed, and the results provided insight into potential antifungal substances. Our work provides a novel strain, HF10, for developing a promising biological control agent against Sclerotinia, which infects oilseed rape and other plants.

Pathological progress and remission strategies of osteoarthritic lesions caused by long-term joint immobilization.

OBJECTIVE: While joint immobilization is a useful repair method for intra-articular ligament injury and periarticular fracture, prolonged joint immobilization can cause multiple complications. A better understanding how joint immobilization and remobilization impact joint function and homeostasis will help clinicians develop novel strategies to reduce complications. DESIGN: We first determined the effects of long-term immobilization on joint pain and osteophyte formation in patients after an extraarticular fracture or ligament injury. We then developed a mouse model of joint immobilization and harvested the knee joint samples at 2, 4, and 8 weeks. We further determined the effects of remobilization on recovery of the osteoarthritis (OA) lesions induced by immobilization in mice. RESULTS: We found that the long-term (6 weeks) joint immobilization caused significant joint pain and osteophytes in patients. In mice, 2-week immobilization already induced moderate sensory innervation and increased pain sensitivity and infiltration in synovium without inducing marked osteophyte formation and cartilage loss. Long-term immobilization (4 and 8 weeks) induced more severe sensory innervation and inflammatory infiltration in synovium, massive osteophyte formation on both sides of the femoral condyle, and the edge of the tibial plateau and significant loss of the articular cartilage in mice. Remobilization, which ameliorates normal joint load and activity, restored to certain extent some of the OA lesions and joint function in mice. CONCLUSIONS: Joint immobilization caused multiple OA-like lesions in both mice and humans. Joint immobilization induced progressive sensory innervation, synovitis, osteophyte formation, and cartilage loss in mice, which can be partially ameliorated by remobilization.

Piezo1 activation accelerates osteoarthritis progression and the targeted therapy effect of artemisinin.

INTRODUCTION: Osteoarthritis (OA) is a devastating whole-joint disease affecting a large population worldwide with no cure; its mechanism remains poorly defined. Abnormal mechanical stress is the main pathological factor of OA. OBJECTIVES: To investigate the effects of Piezo1 activation on OA development and progression and to explore Piezo1-targeting OA treatment. METHODS: The expression levels of Piezo1 were determined in human OA cartilage and experimental OA mice. Mice with genetic Piezo1 deletion in chondrocytes or intra-articular injection of the Piezo1 activator Yoda1 were utilized to determine the effects on DMM-induced OA progression. Effects of artemisinin (ART), a potent antimalarial drug, on Piezo1 activation, chondrocyte metabolism and OA lesions were determined. RESULTS: Piezo1 expression was elevated in articular chondrocytes in human OA and DMM-induced mouse OA cartilage. Piezo1 deletion in chondrocytes largely attenuates DMM-induced OA-like phenotypes. In contrast, intra-articular injection of Yoda1 aggravates the knee joint OA lesions in mice. PIEZO1 activation increases, while PIEZO1 siRNA knockdown decreases, expression of RUNX2 and catabolic enzymes MMP13 and ADAMTS5 in primary human articular chondrocytes in a PI3K-AKT dependent manner. We have provided strong evidence supporting that ART is a novel and potent inhibitor of Piezo1 activation in primary OA-HACs and all cell lines examined, including human endothelial HUVEC cells, ATDC5 chondrocyte-like cells and MLO-Y4 osteocytes-like cells. Results from in vitro experiments confirmed that ART decreases the Yoda1-induced increases in the levels of OA-related genes and p-PI3K and p-AKT proteins in OA-HACs and alleviates DMM-induced OA lesions in mice. CONCLUSIONS: We establish a critical role of Piezo1 in promoting OA development and progression and define ART as a potential OA treatment.

[Advances in several important antimicrobial lipopeptids from Bacillus spp.]

Bacillus spp. are probiotics and can secrete a variety of natural antimicrobiol active substances, of which lipopeptides are an important class. Up to now, about 90 lipopeptides have been identified, and most of them are cyclic lipopeptides. surfactin, iturin, fengycin, bacillomycin and polymyxins are widely studied, and the first three have huge potential for application due to their properties of surfactants and anti-fungal, anti-bacterial, anti-viral, anti-tumor and anti-inflammatory functions. In this paper, the research progress in the structure, function, synthesis regulation, separation, purification and production of surfactin, iturin and fengycin was reviewed. Synthetic biology is a vital means to increase the yield of lipopeptides, and in the future, lipopeptides can be used in crop cultivation, animal farming, food, medicine and petroleum industries as well as environmental protection. Future research should be strengthened on the discovery of new lipopeptides, synthesis of high-activity lipopeptides, economical production of lipopeptides on a large scale and their safety evaluation.

Repurposing of Pirfenidone (Anti-Pulmonary Fibrosis Drug) for Treatment of Rheumatoid Arthritis.

Clinical studies have shown that pirfenidone (PFD) effectively relieves joint pain in rheumatoid arthritis (RA) patients. However, the detailed mechanisms underlying the anti-RA effects of PFD have not been investigated. This study was undertaken to investigate the repurposing of PFD for the treatment of RA, and explore its anti-rheumatic mechanisms. A collagen-induced arthritis (CIA) rat model was used to observe joint pathological changes following PFD treatment. Based on bioinformatics to predict the mechanism of PFD anti-RA, using EA. hy926 and TNF-α-induced MH7A cells to establish in vitro model to explore its biological mechanism from the perspectives of synovial inflammation and angiogenesis. PFD significantly relieved pathological changes, including joint swelling, synovial hyperplasia, inflammatory cell infiltration and joint destruction. PFD was also associated with reduced expression of MMP-3 and VEGF in articular chondrocytes and synovial cells of CIA rats (p < 0.05). Using bioinformatic methods, we predicted that PFD inhibits cell inflammation and migration by interfering with the JAK2/STAT3 and Akt pathways. These results were verified using in vitro models. In particular, PFD effectively reduced the expression of pro-inflammatory, chondrogenic, and angiogenic cytokines, such as IL-1ß, IL-6, IL-8, MMP-1/3/2/9 and VEGF (p < 0.05), in TNF-α-induced MH7A cells. In addition, PFD significantly reduced the production of MMP-2/9 and VEGF in EA. hy926 cells, thereby weakening migration and inhibiting angiogenesis (p < 0.05). These findings suggest that PFD may alleviate the pathological process in CIA rats, by inhibiting inflammation and angiogenesis through multiple pathways, and serve as a potential therapeutic drug for RA.

Biphasic Effect of Pirfenidone on Angiogenesis.

Pirfenidone (PFD), a synthetic arsenic compound, has been found to inhibit angiogenesis at high concentrations. However, the biphasic effects of different PFD concentrations on angiogenesis have not yet been elucidated, and the present study used an in vitro model to explore the mechanisms underlying this biphasic response. The effect of PFD on the initial angiogenesis of vascular endothelial cells was investigated through a Matrigel tube formation assay, and the impact of PFD on endothelial cell migration was evaluated through scratch and transwell migration experiments. Moreover, the expression of key migration cytokines, matrix metalloproteinase (MMP)-2 and MMP-9, was examined. Finally, the biphasic mechanism of PFD on angiogenesis was explored through cell signaling and apoptosis analyses. The results showed that 10-100 µM PFD has a significant and dose-dependent inhibitory effect on tube formation and migration, while 10 nM-1 µM PFD significantly promoted tube formation and migration, with 100 nM PFD having the strongest effect. Additionally, we found that a high concentration of PFD could significantly inhibit MMP-2 and MMP-9 expression, while low concentrations of PFD significantly promoted their expression. Finally, we found that high concentrations of PFD inhibited EA.hy926 cell tube formation by promoting apoptosis, while low concentrations of PFD promoted tube formation by increasing MMP-2 and MMP-9 protein expression predominantly via the EGFR/p-p38 pathway. Overall, PFD elicits a biphasic effect on angiogenesis through different mechanisms, could be used as a new potential drug for the treatment of vascular diseases.

Amygdalin Attenuates Atherosclerosis and Plays an Anti-Inflammatory Role in ApoE Knock-Out Mice and Bone Marrow-Derived Macrophages.

Amygdalin, the main component of Prunus persica (L.) Stokes, has been used to treat atherosclerosis in mouse model due to its anti-inflammatory role. However, the underlying mechanism remains poorly understood. This study aimed to evidence the influence of amygdalin on high-fat diet-induced atherosclerosis in ApoE knock-out (ApoE-/-) mice, and unravel its anti-inflammatory mechanism. ApoE-/- mice fed with high-fat diet for eight weeks were randomly divided into four groups and injected with amygdalin at the concentration of 0.08 or 0.04 mg/kg for 12 weeks. Additionally, bone marrow-derived macrophages were intervened with oxidized low-density lipoprotein (oxLDL) or lipopolysaccharide plus various concentrations of amygdalin for further exploration. Body weight, serum lipid profiles and inflammatory cytokines were detected by ELISA, gene expression by RT-PCR, plaque sizes by Oil Red O, lymphatic vessels of heart atrium and Tnfα production by immunofluorescence staining. MAPKs, AP-1 and NF-κB p65 pathways were also explored. Amygdalin decreased body weight, serum lipids, plaque size, lymphatic vessels and inflammatory cytokines (Il-6, Tnfα), Nos1 and Nos2, and increased Il-10 expression in ApoE-/- mice. In oxLDL-induced bone marrow-derived macrophages, amygdalin reduced inflammatory cytokines (Il-6, Tnfα), Nos1 and Nos2, and increased Il-10 production. These effects were associated with the decreased phosphorylation of Mapk1, Mapk8, Mapk14, Fos and Jun, and the translocation of NF-κB p65 from nucleus to cytoplasm. The results suggested that amygdalin could attenuate atherosclerosis and play an anti-inflammatory role via MAPKs, AP-1 and NF-κB p65 signaling pathways in ApoE-/- mice and oxLDL-treated bone marrow-derived macrophages.

Corrigendum: Alleviation of Synovial Inflammation of Juanbi-Tang on Collagen-Induced Arthritis and TNF-Tg Mice Model.

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Taoren Honghua Drug Attenuates Atherosclerosis and Plays an Anti-Inflammatory Role in ApoE Knock-Out Mice and RAW264.7 Cells.

Taoren Honghua drug is a traditional Chinese medicinal drug used to treat cardiovascular disease. The aim of the study is to investigate the effects of Taoren Honghua drug on inflammation and atherosclerosis in ApoE knock-out mice and RAW264.7 cells. ApoE knock-out mice fed with high fat diet for 8 weeks were randomly divided into five groups and then continued the high fat diet, or plus Taoren Honghua drug at concentrations of 3.63, 1.815, and 0.9075 g/ml, or plus Simvastatin at 2.57 mg/kg. RAW 264.7 cells were intervened with lipopolysaccharide or lipopolysaccharide plus different concentrations of Taoren Honghua drug. Compared to mice only with high fat diet, mice with high fat diet and Taoren Honghua drug showed lower body weight, triglyceride, cholesterol, IL-6 and TNF-α, smaller plaque sizes, less lymph vessel, and T cell contents of lymph nodes, but higher IL-10 level. In RAW264.7 cells, groups with LPS plus Taoren Honghua drug had lower IL-6 and TNF-α, but higher IL-10 than LPS group, as revealed by PCR or ELISA methods. A decrease of total or phosphorylated ERK1/2, JNK, p38, ERK5, STAT3, and AKT were detected, so was the translocation of NF-κB p65 from nuclear to cytoplasm. These results suggested that Taoren Honghua drug could attenuate atherosclerosis and play an anti-inflammatory role via MAPKs, ERK5/STAT3, and AKT/NF-κB p65 signaling pathways in ApoE knock-out mice and lipopolysaccharide-induced RAW264.7 cells.

Alleviation of Synovial Inflammation of Juanbi-Tang on Collagen-Induced Arthritis and TNF-Tg Mice Model.

Rheumatoid arthritis (RA) is a chronic autoimmune disease that is primarily characterized by synovial inflammation. In this study, we found that a traditional Chinese decoction, Juanbi-Tang (JBT), JBT attenuated the symptoms of collagen-induced arthritis (CIA) mice and in tumor necrosis factor transgenic (TNF-Tg) mice by attenuating the arthritis index and hind paw thickness. According to histopathological staining of ankle sections, JBT significantly decreased the area of inflammation and reduced bone destruction of ankle joints in both these two types of mice. Moreover, decreased tartaric acid phosphatase-positive osteoclasts were observed in the JBT group compared with those found in the control group. We also revealed that JBT suppressed monocytes and T cells as well as the production of CCL2, CCR6, and CXCR3 ligands. We next used high-performance liquid chromatography to investigate the components and pharmacological properties of this classical herbal medicine in traditional Chinese medicine. Based on network pharmacology, we performed computational prediction simulation of the potential targets of JBT, which indicated the NF-kappa B pathway as its target, which was confirmed in vitro. JBT suppressed the production of pro-inflammatory cytokines including interleukin-6 (IL-6) and IL-8, and inhibited the expression of matrix metalloproteinase 1 in fibroblast-like synoviocytes derived from RA patients (MH7A cells). Furthermore, JBT also suppressed the phosphorylation of p38, JNK, and p65 in TNF-α-treated MH7A cells. In summary, this study proved that JBT could inhibit synovial inflammation and bone destruction, possibly by blocking the phosphorylation of NF-kappa B pathway-mediated production of proinflammatory effectors.

Screening strains for microbial biosorption technology of cadmium.

Heavy metals contaminate the environment and provide a threat to public health through drinking water and food chain. Microbial biosorption technology provides a more economical and competitive solution for bioremediation of toxicants such as heavy metals, and microbial genetic modification may modify microbes towards optimal sorption. It is very important to screen suitable strains for this purpose. In this study, three different types of microorganisms Escherichia coli, Bacillus subtilis and Saccharomyces cerevisiae were isolated and identified, from uncontaminated soils, and compared their sorption differences with respect to cadmium (Cd2+). We evaluated the effects of contact time and initial concentration on Cd2+ uptake, and found pseudo-second-order kinetic models were more suitable to describe biosorption processes. Adsorption isotherms were used to reflect their biosorption capacity. The maximum biosorption capacities of three strains calculated by the Langmuir model were 37.764, 56.497, and 22.437 mg Cd/g biomass, respectively. In bacteria, Cd2+ biosorption mainly occurred on cell wall, while the difference in biosorption between yeast inside and outside the cell was not significant. We found that due to the structural differences, the removal rate of E. coli surface decreased at a high concentration, while S. cerevisiae still had a lower biosorption capacity. FTIR spectroscopy reflected the difference in functional groups involved in biosorption by three strains. SEM-EDS analysis showed the binding of Cd2+ to microorganisms mainly relied on ion exchange mechanism. Based on the above results, we suggested that B. subtilis is more suitable to get genetically modified for heavy metal biosorption.

Detection of Metallothionein Proteins by Enzyme-Linked Immunosorbent Assay (ELISA).

Metallothioneins (MTs) are low-molecular-weight, cysteine-rich proteins that bind to heavy metals. MTs play a key role in the homeostasis of metal ions, maintaining intracellular redox equilibria and free radical scavenging. In several studies, under different conditions such as cancer development, drug therapy and heavy metal stress, the unique structural changes and functional effects of MT were studied. Although several assays are available to monitor the content and type of Metallothionein (MT) from environmental samples or in biomedical assays, Enzyme-Linked Immunosorbent Assays (ELISA) became the preferred method of MT detection. ELISA is low in cost, specific, simple, and efficient. This review evaluates the advantages and disadvantages of using different types of ELISA in the detection of metallothioneins from environmental or clinical samples as well as ways of its validation and cross-validation.

Astragalin Suppresses Inflammatory Responses and Bone Destruction in Mice With Collagen-Induced Arthritis and in Human Fibroblast-Like Synoviocytes.

Astragalin, as a bioactive flavonoid with anti-inflammatory, antioxidant, and protective properties, provides a potential agent for rheumatoid arthritis (RA). In this study, its therapeutic efficacy and the underlying mechanisms were explored using DBA/1J mice with collagen-induced arthritis (CIA). It was demonstrated that astragalin could significantly attenuate inflammation of CIA mice. The effects were associated with decreased severity of arthritis (based on the arthritis index), joint swelling and reduced bone erosion and destruction. Furthermore, astragalin treatment suppressed the production of pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6, and IL-8), and inhibited the expression of matrix metalloproteinases (MMP-1, MMP-3, and MMP-13) in chondrocytes and synovial cells of CIA mice. Fibroblast-like synoviocytes derived from RA patients (MH7A cells) were applied to verify these effects. In vitro, astragalin inhibited the expression of matrix metalloproteinases (MMP-1, MMP-3, and MMP-13) dose-dependently in TNF-α-induced MH7A cells, with no apparent cytotoxicity. Furthermore, astragalin suppressed the phosphorylation of p38, JNK, and the activation of c-Jun/AP-1 in TNF-α-induced MH7A cells. In conclusion, it has proven that astragalin could attenuate synovial inflammation and joint destruction in RA at least partially by restraining the phosphorylation of MAPKs and the activating of c-Jun/AP-1. Therefore, astragalin can be a potential therapeutic agent for RA.

Osteogenic magnesium incorporated into PLGA/TCP porous scaffold by 3D printing for repairing challenging bone defect.

Bone defect repair is a challenging clinical problem in musculoskeletal system, especially in orthopaedic disorders such as steroid associated osteonecrosis (SAON). Magnesium (Mg) as a biodegradable metal with properly mechanical properties has been investigating for a long history. In this study, Mg powder, poly (lactide-co-glycolide) (PLGA), ß-tricalcium phosphate (ß-TCP) were the elements to formulate a novel porous PLGA/TCP/Mg (PTM) scaffolds using low temperature rapid prototyping (LT-RP) technology. The physical characterization of PTM scaffold and Mg ions release were analyzed in vitro. The osteogenic and angiogenic properties of PTM scaffolds, as well as the biosafety after implantation were assessed in an established SAON rabbit model. Our results showed that the PTM scaffold possessed well-designed bio-mimic structure and improved mechanical properties. Findings of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and micro-computed tomography (micro CT)-based angiography indicated that PTM scaffold could increase blood perfusion and promote new vessel ingrowth at 4 weeks after surgery, meanwhile, a plenty of newly formed vessels with well-architective structure were observed at 8 weeks. Correspondingly, at 12 weeks after surgery, micro-CT, histological and mechanical properties analysis showed that PTM could significant enhance new bone formation and strengthen newly formed bone mechanical properties. The mean bone volume in PTM group was 56.3% greater than that in PT group. Biosafety assessments from 0 to 12 weeks after implantation did not induce increase in serum Mg ions concentration, and immune response, liver and kidney function parameters were all at normal level. These findings suggested that the PTM scaffold had both osteogenic and angiogenic abilities which were synergistic effect in enhancing new bone formation and strengthen newly formed bone quality in SAON. In summary, PTM scaffolds are promising composite biomaterials for repairing challenging bone defect that would have great potential for its clinical translation.

The efficacy of the traditional Chinese medicine Juanbi pill combined with methotrexate in active rheumatoid arthritis: study protocol for a randomized controlled trial.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by swelling and painful joints, eventually leading to joint destruction. There is still a lack of effective therapy to treat RA. The Juanbi pill is a Chinese medicine that has been widely used to treat active RA in China for hundreds of years, relieving pain and protecting the affected joints from malformation. However, there is no solid evidence to show the effect of the Juanbi pill on the management of active RA. METHODS/DESIGN: We will conduct a multicenter, randomized, double-blind, placebo-controlled clinical trial to determine whether the traditional Chinese medicine Juanbi pill could relieve joint pain in RA and protect the joints. A total of 120 patients with active RA will be enrolled and treated with the Juanbi pill or a placebo for 3 months. The primary outcome measures are as follows: rate of in the American College of Rheumatology (ACR)50, change in the 28-joint Disease Activity Score (DAS28) from baseline at beginning of therapy to 3 months, and a change in the van der Heijde modified Sharp score measured from baseline to 12 months. The secondary outcome measures are as follows: rate of change in ACR20, ACR70, Health Assessment Questionnaire-Disability Index (HAQ-DI), and change in score in the Patient Assessment of Arthritis Pain, Patient Global Assessment of Arthritis, and the Athens Insomnia Scale (AIS) from baseline to 2-week, 1-month, 2-month, 3-month, 6-month, and 12-month follow up. In addition, the rate of change (score) in the ACR50 and DAS28 from the baseline to 2-week, 1-month, 2-month, 6-month, and 12-month follow up are also the secondary outcome measures. DISCUSSION: Although the Juanbi pill has been used in China for many years to treat RA, there is a lack of consensus about its effectiveness. This trial will provide convincing evidence about the effect of Juanbi pill on active RA. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02885597 . Registered on 30 August 2016.

Cucurbitacin E inhibits TNF-α-induced inflammatory cytokine production in human synoviocyte MH7A cells via suppression of PI3K/Akt/NF-κB pathways.

Increasing studies indicated that Cucurbitacin E (CuE), a compound isolated from Cucurbitaceae, has been shown anti-inflammatory effect. However, the effect of CuE on rheumatoid arthritis (RA) inflammatory response and its potential molecular mechanism are still unknown. In this study, we demonstrated that CuE significantly suppressed TNF-α-induced inflammatory cytokines production interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and interleukin-8 (IL-8) mRNA and protein expression in human synoviocyte MH7A cells. Furthermore, we found that CuE also inhibited TNF-α-induced phosphorylation of NF-κBp65, IKKα/ß, and IκBα in a dose-and time-dependent manner as well as NF-κBp65 nuclear translocation. Finally, we showed that CuE blocked the upstream targets of NF-κB pathway RIP1/PI3K/Akt. Interestingly, PI3K inhibitor LY294002 completely blocked the TNF-α-induced activation of p85, Akt and the whole cascade of the NF-κB signaling components and suppressed inflammatory cytokines production in mRNA and protein levels similarly as CuE. Our studies provided the first evidence that CuE inhibited TNF-α-induced inflammatory cytokine production in human synoviocyte MH7A cells via modulation of PI3K/Akt/NF-κB pathway. These findings indicated that CuE is a potential candidate for RA therapy.