Anti-malarial drug artesunate can suppress inflammation and prevent cartilage and bone destruction in collagen-induced arthritis model in rats—suggesting it may be a potent drug for rheumatoid arthritis (RA) therapy. We aimed to investigate its effect on the invasive property of fibroblast-like synoviocytes (FLS) from patients with RA. Synovial tissues were obtained by closed needle biopsy from active RA patients, and FLS were isolated and cultured in vitro. RA-FLS were treated with artesunate at various concentrations, while methotrexate or hydroxychloroquine was employed as comparator drugs. Cell viability, proliferation, cell cycle, apoptosis, migration, invasion, and pseudopodium formation of RA-FLS were assessed by CCK-8 assays, EdU staining, Annexin V-FITC/PI staining, transwell assays, or F-actin staining, respectively. Further, relative changes of expressed proteases were analyzed by Proteome profiler human protease array and verified by quantitative real-time PCR (qPCR), Western blot, and ELISA. The expression of signaling molecules of MAPK, NF-κB, AP-1, and PI3K/Akt pathways were measured by qPCR and Western blot. PDK-1 knockdown by specific inhibitor AR-12 or siRNA transfection was used to verify the pharmacological mechanism of artesunate on RA-FLS. Artesunate significantly inhibited the migration and invasion of RA-FLS in a dose-dependent manner with or without TNF-α stimulation. The effect was mediated through artesunate inhibition of MMP-2 and MMP-9 production, and pre-treatment with exogenous MMP-9 reversed the inhibitory effect of artesunate on RA-FLS invasion. Artesunate had a stronger inhibitory effect on migration and invasion of RA-FLS as well as greater anti-inflammatory effect than those of hydroxychloroquine. Similar inhibitory effect was detected between artesunate and methotrexate, and synergy was observed when combined. Mechanistically, artesunate significantly inhibited PDK-1 expression as well as Akt and RSK2 phosphorylation—in a similar manner to PDK-1-specific inhibitor AR-12 or PDK-1 knockdown by siRNA transfection. This inhibition results in suppression of RA-FLS migration and invasion as well as decreased MMP-2 and MMP-9 expression. Our study demonstrates artesunate is capable of inhibiting migration and invasion of RA-FLS through suppression of PDK1-induced activation of Akt and RSK2 phosphorylation—suggesting that artesunate may be a potential disease-modifying anti-rheumatic drug for RA.

Background: The link between body mass index (BMI) and disease characteristics in rheumatoid arthritis (RA) remains controversial. Body composition (BC) has been more frequently recommended to be used instead of BMI for more accurate assessment. Our study aimed to investigate the characteristics of BC in RA patients and their associations with disease characteristics.Methods: Body composition was assessed in consecutive Chinese RA patients and control subjects by bioelectric impedance analysis. Overfat was defined by body fat percentage (BF%) as ≥25% for men and ≥35% for women. Myopenia was defined by appendicular skeletal muscle mass index (ASMI) ≤7.0 kg/m² in men and ≤5.7 kg/m² in women. BMI and clinical data including disease activity, function, and radiographic assessment were collected. Active disease was defined by disease activity score in 28 joints with four variables including C‐reactive protein (DAS28‐CRP) ≥2.6. Functional limitation was defined as Stanford health assessment questionnaire disability index (HAQ‐DI) >1. Radiographic joint damage (RJD) was defined as the Sharp/van der Heijde modified sharp score (mTSS) >10.Results: There were 457 RA patients (mean age 49.5 ± 13.1 years old with 82.7% women) and 1860 control subjects (mean age 34.3 ± 9.9 years old with 51.2% women) recruited. Comparisons of BMI and BC between RA patients and control subjects in age and gender stratification showed that lower BMI with 17.7% underweight and lower ASMI with 45.1% myopenia are the main characteristics in RA patients. Compared with those without myopenia, RA patients with myopenia had significantly higher DAS28‐CRP (median 3.5 vs. 3.0), higher HAQ‐DI (median 0.38 vs. 0.13) with higher rate of functional limitation (24.8% vs. 7.6%), and higher mTSS (median 22.3 vs. 9.0) with more RJD (71.8% vs. 45.8%) (all P < 0.001). Multivariate logistic regression analysis showed myopenia were positively associated with functional limitation (OR = 2.546, 95% CI: 1.043–6.217) and RJD (OR = 2.660, 95% CI: 1.443–4.904). All RA patients were divided into four BC subgroups according to overfat and myopenia. Those with both overfat and myopenia had the worst disease characteristics. After adjustment for confounding factors, significant additive interactions were observed between overfat and myopenia in active disease (AP = 0.528, 95% CI: 0.086–0.971), functional limitation (AP = 0.647, 95% CI: 0.356–0.937), and RJD (AP = 0.514, 95% CI: 0.139–0.890).Conclusions: Myopenia is very common in RA patients that is associated with functional limitation and joint damage in RA. Further research on the underlying mechanism and the effect of skeletal muscle mass improvement in RA management are worth exploring in the future.

In rheumatoid arthritis (RA), imbalanced T cells subsets play a critical role in sustaining chronic inflammatory responses in the synovium. Naïve T cells in RA patients undergo maldifferentiation, including an increase in the effector Th1/Th17 lineage and a reduction in regulatory T (Treg) cells. Upon stimulation, naïve CD4+CD45RO- T cells from RA patients exhibited insufficient expression of Foxp3, which induced a deficiency in Tregs production and an imbalance of Treg/Th17 differentiation. Further mechanistic study indicated that RA T cells failed to produce sufficient levels of the histone acetyltransferase Tip60, leading to reduced acetylation of Foxp3; this, in turn, decreased Foxp3 expression, impaired Treg commitment, and promoted Th17 production. Moreover, in human synovium chimeric mice, suppression of Tip60 activity in healthy T cells promoted tissue infiltration and arthritogenesis, while reconstitution of Tip60 in RA T cells suppressed synovitis and effector T cell infiltration. Our findings link T cell maldifferentiation and tissue infiltration with Tip60-mediated Foxp3 acetylation and identify Tip60 as a potential therapeutic target for suppression of tissue inflammation and autoimmunogenesis in RA.

Objective: Activation of osteoclastogenesis at the bone site in rheumatoid arthritis (RA) is well established. The mechanisms by which circulating osteoclast precursors contribute are still unclear. Peroxisome proliferator–activated receptor γ coactivator 1β (PGC‐1β) is implicated in transcriptional regulation of osteoclastogenesis in mouse models. This study was undertaken to investigate the contribution of PGC‐1β to circulating osteoclast precursors and its link to bone destruction in RA.Methods: PGC‐1β expression in RA peripheral blood CD14+ monocytes was increased and showed correlation with joint destruction shown on radiographs. Cells from RA patients or healthy controls were transfected with a lentivirus vector for PGC‐1β gene silencing or overexpression and cultured with macrophage colony‐stimulating factor and RANKL. Bone resorption activity, bone‐degrading enzymes, and signaling molecules were measured in these mature osteoclasts.Results: Increased nuclear accumulation of PGC‐1β was observed in RA peripheral blood CD14+ monocytes, and these cells had stronger osteoclastogenesis than in healthy controls. PGC‐1β protein expression was positively correlated with radiographic joint destruction (r = 0.396–0.413; all P < 0.05). PGC‐1β knockdown suppressed (51–82% reduction) the expression of cathepsin K, tartrate‐resistant acid phosphatase (TRAP), and matrix metalloproteinase 9 (MMP‐9), as well as osteoclast differentiation and bone resorption activity. Conversely, PGC‐1β overexpression increased these markers (by 1.5–1.8‐fold) and osteoclastogenesis. VIVIT, an inhibitor of NFATc1 activation, inhibited the effect of overexpressed PGC‐1β by reducing cathepsin K, TRAP, and MMP‐9 expression. Chromatin immunoprecipitation assay and dual‐luciferase reporter gene assay showed PGC‐1β bound to NFATc1 promoter, leading to transcriptional activation.Conclusion: Activation of the PGC‐1β/NFATc1 pathway in circulating osteoclast precursors was associated with bone destruction in RA. This may represent a new treatment target.

Synovium fibroblast-like synoviocytes (FLSs) are important participants in the pathogenesis of synovitis and joint destruction in rheumatoid arthritis (RA). Pyroptosis is a pro-inflammatory and cell lytic programmed cell death mechanism mediated by gasdermin (GSDM) family proteins. In this study, we demonstrated the increased expression of GSDME and increased levels of GSDME-mediated pyroptosis in RA synovial tissues. In vitro, stimulation with TNF-α plus hypoxia mimicking the inflammatory and hypoxic environment in RA synovium induced GSDME-mediated pyroptosis in RA-FLSs in combination with the promotion of migration and invasion abilities and the release of inflammatory cytokines (IL-6, IL-8). Moreover, knockdown of GSDME significantly inhibited the proliferation rate, migration/invasion effects and cytokines released through the reduction of GSDME-mediated pyroptosis. The immunohistochemistry results showed that RA patients with high GSDME N-terminal (GSDME-NT) expression, which is the active form of GSDME, showed higher IL-6 expression in both lining and sublining layer of synovium than that in patients with low GSDME-NT expression, osteoarthritis and non-inflammatory orthopedic arthropathies. Our findings revealed a novel mechanism regulating cell proliferation, migration, invasion and inflammatory cytokines release during the process of GSDME mediated pyroptosis in RA.

Background: Associations between rheumatoid arthritis (RA) and reduced skeletal muscle have been studied, and we firstly reported myopenia independently predict one-year radiographic progression in RA. Myokine myostatin can negatively regulate skeletal muscle mass and promote osteoclast differentiation. However, there is no report about their relationships in RA patients. We firstly explored the relationship of serum myostatin and disease characteristics, as well as aggravated joint destruction during one-year follow-up.Methods: Consecutive RA patients were recruited from a real-world prospective cohort and completed at least one-year follow-up. Baseline serum level of myostatin was measured by enzyme-linked immunosorbent assay. Clinical data in RA patients as well as muscle index in both RA patients and healthy controls were collected. One-year radiographic progression as primary outcome was defined by a change in the total Sharp/van der Heijde modified score ≥0.5 units.Results: Totally 344 RA patients (age 47.9 ± 12.5 years, 84.0% female) and 118 healthy control subjects (age 42.8 ± 11.3 years, 74.6% female) were recruited. Compared with healthy controls, RA patients showed a higher level of serum myostatin at baseline (3.241 ± 1.679 ng/ml vs. 1.717 ± 0.872 ng/ml, P<0.001), although lower appendicular skeletal muscle mass index (ASMI, 6.0 ± 0.9 kg/m²vs. 6.5 ± 1.0 kg/m², P<0.001). In RA patients, those with high myostatin level showed a higher rate of radiographic progression than low myostatin group (45.3% vs. 18.6%, P<0.001). Furtherly, RA patients were stratified into four subgroups according to serum myostatin and myopenia. Compared with other three subgroups, RA patients with high myostatin overlapping myopenia had the highest rate of radiographic progression (67.2% vs. 10.3%-31.4%, P<0.001), as well as the lowest proportion of remission and the highest rate of physical dysfunction during one-year follow-up. After adjustment for confounding factors, high serum myostatin (AOR=3.451, 95%CI: 2.016-5.905) and myopenia (AOR=2.387, 95%CI: 1.416-4.022) at baseline were risk factors for one-year radiographic progression, especially for those with high myostatin overlapping myopenia (AOR=10.425, 95%CI: 3.959-27.450) as the highest-risk individuals among four subgroups. Significant synergistic interaction effect was observed between high myostatin and myopenia on one-year radiographic progression (AP=66.3%, 95%CI: 43.2%-89.3%).Conclusion: Myostatin is a novel predictor of aggravated joint destruction in RA patients which has synergistic interaction with myopenia for predicting value.