Development of Spondyloarthritis After COVID-19 in HLAB27-Positive Monozygotic Twins: Case Reports With Single Cell Transcriptome Profiling

A subset of spondyloarthritis (SpA) called ‘reactive arthritis’ is triggered by causal pathogens, usually bacteria related to venereal disease or gastrointestinal infection. During the outbreak of coronavirus disease 2019 (COVID-19), there have been case reports about SpA after COVID-19, but the causality is still elusive. We described cases of 23-year-old monozygotic twins both diagnosed with SpA after COVID-19. The probable linkage between SpA and COVID-19 was elaborated with our cases as well as literature reviews. Of note, shared genetic traits by monozygotic twins, particularly HLA-B27 positivity, might have contributed to their susceptibility to COVID-19-induced SpA. Moreover, single-cell transcriptome analysis revealed that the transcriptomic profile of peripheral compartment of SpA after COVID-19 was distinctive from that of typical radiographic axial SpA as shown by differential expression of ribosomal protein S26 (RPS26) and small nucleolar RNA host gene 5 (SNHG5) in nearly all subsets of peripheral blood mononuclear cells.

Lupus Heart Disease Modeling with Combination of Induced Pluripotent Stem Cell-Derived Cardiomyocytes and Lupus Patient Serum

Background and Objectives@#Systemic lupus erythematosus (SLE) is a chronic autoimmune disease mainly affecting young women of childbearing age. SLE affects the skin, joints, muscles, kidneys, lungs, and heart. Cardiovascular complications are common causes of death in patients with SLE. However, the complexity of the cardiovascular system and the rarity of SLE make it difficult to investigate these morbidities. Patient-derived induced pluripotent stem cells (iPSCs) serve as a novel tool for drug screening and pathophysiological studies in the absence of patient samples. @*Methods@#and Results: We differentiated CMs from HC- and SLE-iPSCs using 2D culture platforms. SLE-CMs showed decreased proliferation and increased levels of fibrosis and hypertrophy marker expression; however, HC-and SLE-monolayer CMs reacted differently to SLE serum treatment. HC-iPSCs were also differentiated into CMs using 3D spheroid culture and anti-Ro autoantibody was treated along with SLE serum. 3D-HC-CMs generated more mature CMs compared to the CMs generated using 2D culture. The treatment of anti-Ro autoantibody rapidly increased the gene expression of fibrosis, hypertrophy, and apoptosis markers, and altered the calcium signaling in the CMs. @*Conclusions@#iPSC derived cardiomyocytes with patient-derived serum, and anti-Ro antibody treatment could serve in effective autoimmune disease modeling including SLE. We believe that the present study might briefly provide possibilities on the application of a combination of patient-derived materials and iPSCs in disease modeling of autoimmune diseases.

Janus kinase inhibitors for the treatment of rheumatoid arthritis

Rheumatoid arthritis is a chronic inflammatory destructive disorder that affects the joints, muscles, and tendons accompanying various extra-articular manifestations. Traditional disease-modifying anti-rheumatic drugs (DMARDs) represent the basic treatment for rheumatoid arthritis. Over the last 20 years, biologic DMARDs (tumor necrosis factor inhibitors, interleukin-1 inhibitors, interleukin-6 inhibitors, T cell inhibitors, and B cell inhibitors) have been widely used as a novel class of DMARDs that have efficacy and efficiency. Discovery of the underlying pathogenesis of autoimmune disease enables us to develop new target therapies such as a Janus kinase (JAK) inhibitor. Activated JAK is known to activate signal transducers as well as activators of transcription (STAT) signaling. A JAK inhibitor is a type of medication that functions by inhibiting the JAK-STAT signaling pathway. In addition, it is easy to take a JAK inhibitor orally. In Korea, several JAK inhibitors have been approved. This review describes the types of JAK inhibitors, recommended doses, side effects, and updated European Alliance of Associations for Rheumatology guidelines. Clinicians should more often consider JAK inhibitors in the treatment of refractory rheumatoid arthritis in current rheumatology clinics

Human kidney organoids model the tacrolimus nephrotoxicity and elucidate the role of autophagy

Background/Aims@#Tacrolimus has been used as an immunosuppressive agent in organ transplantation. Despite the therapeutic benefits, tacrolimus’s use is limited due to its nephrotoxicity. To reduce tacrolimus nephrotoxicity, effective humanized experimental models may be helpful. Here, we modeled tacrolimus nephrotoxicity using kidney organoids derived from human inducible pluripotent stem cells (iPSCs) in vitro. @*Methods@#Kidney organoids were differentiated from the CMC11 iPSC cell line, re-seeded in 96-well plates, and treated with tacrolimus at doses of 0, 30, or 60 μM for 24 hours. This in vitro model was compared to a mouse model of tacrolimus nephrotoxicity and the associated mechanisms were investigated. @*Results@#The size of the kidney organoids and cell viability decreased in dose-dependent manners after treatment with tacrolimus. The number of tubular cells decreased with a loss of polarity, similar to the effects seen in mouse tacrolimus nephrotoxicity. Ultrastructural analysis showed numerous vacuoles in the proximal tubular cells of the kidney organoids treated with tacrolimus. Tacrolimus treatment induced oxidative stress and mitochondrial dysfunction, and autophagic activity was enhanced in the kidney organoids. Rapamycin, an autophagy inducer, accelerated cell death in the kidney organoid model of tacrolimus nephrotoxicity, which was attenuated by treatment with 3-methyladenine, an autophagy inhibitor. These findings indicate that the augmentation of autophagy by rapamycin treatment accelerated tacrolimus nephrotoxicity. @*Conclusions@#Our data suggest that human kidney organoids are an effective in vitro model of tacrolimus nephrotoxicity and that autophagy plays a critical role in tacrolimus nephrotoxicity.

Echocardiographic Evaluation of Axial Spondyloarthritis in Korea: Data From the Catholic Axial Spondyloarthritis Cohort

OBJECTIVE: Axial spondyloarthritis (axSpA) is often accompanied by cardiac manifestations, such as valvular heart disease. In this prospective cohort study, we evaluated the incidence of cardiac abnormalities in Korean axSpA patients by echocardiography.METHODS: AxSpA patients were prospectively recruited from a single tertiary hospital. Baseline demographic, clinical, radiographic, and echocardiographic data were collected at the time of enrollment. Echocardiography evaluations were performed with a focus on valvular heart disease and systolic and diastolic function. Logistic regression analyses were used to identify factors associated with diastolic dysfunction in axSpA.RESULTS: A total of 357 axSpA patients were included in the analyses, of whom 78 (21.8%) exhibited diastolic dysfunction, with no reports of systolic dysfunction. Thirteen patients (3.6%) had valvular heart disease, and aortic valve regurgitation (n=5) and mitral valve regurgitation (n=6) were most common. Multivariable logistic regression analyses indicated that older age and higher body mass index (BMI) were positively associated with diastolic dysfunction, whereas human leukocyte antigen (HLA)-B27 positivity was negatively associated with diastolic dysfunction.CONCLUSION: Valvular heart disease is infrequent in Korean axSpA patients. However, diastolic dysfunction is common in axSpA patients, and is significantly associated with older age, higher BMI, and HLA-B27.

Current Stem Cell Therapy for Osteoarthritis / 대한내과학회지

Osteoarthritis is a musculoskeletal disease representative of an aging society. As medical conditions are usually complicated in an aging population, osteoarthritis becomes more frequently encountered in the physician's office. There is a growing need, therefore, for physicians to pay attention to this common orthopedic condition. Cartilage degeneration, arthritic pain, and joint dysfunction are major manifestations of osteoarthritis, and degenerated cartilage is difficult to repair with conventional treatment modalities. Scientists and physicians have developed various therapeutic strategies, including the use of stem cells. Here, we discuss previous and current progress in cartilage regenerative therapy against osteoarthritis.

Erratum: Effects of Individual Education Using a Treating-to-target Strategy in Patients with Rheumatoid Arthritis

The original version of this article contained an error of the ORCID identifier of corresponding author (Ji Hyeon Ju).

Insulin resistance is an independent predictor of erectile dysfunction in patients with gout

BACKGROUND/AIMS: Gout is associated with metabolic disorders that are important risk factors for cardiovascular disease and erectile dysfunction (ED). We aimed to identify independent predictors of ED in patients with gout. METHODS: From August 2014 to August 2015, male outpatients who were being treated for gout in our rheumatology clinic and healthy males without any history of inflammatory disease (control group) were studied. ED was assessed in participants using the five-item version of the International Index of Erectile Function questionnaire. Insulin resistance (IR) was estimated using the homeostatic model assessment (HOMA-IR). Logistic regression analysis was performed to determine the effect of variables on ED risk in all of the study subjects and in patients with gout. RESULTS: We analyzed 80 patients with gout and 70 healthy controls. The median age of patients with gout was 52 years and median disease duration was 120 months. Gout patients were more likely to have ED than controls (55.3% vs. 41.4%, p < 0.047). After adjustment for confounding factors, only HOMA-IR was significantly associated with ED (odds ratio [OR], 1.82; 95% confidence interval [CI], 1.05 to 3.15). Gout patients with ED were more likely to be older (p < 0.001), have higher HOMA-IR (p = 0.048), and have lower glomerular filtration rate (p = 0.038) than those without ED. Multivariate logistic regression analysis showed that HOMA-IR was an independent predictor for ED (OR, 1.62; 95% CI, 1.03 to 2.82) in gout patients. CONCLUSIONS: IR is an independent predictor of ED in patients with gout.

Clinical outcomes and pathological characteristics of immunoglobulin G4-related ophthalmic disease versus orbital inflammatory pseudotumor

BACKGROUND/AIMS: This study investigated the clinical and pathological features of immunoglobulin G4 (IgG4)-related ophthalmic disease. To clarify the features, we compared IgG4-related ophthalmic disease and orbital inflammatory pseudotumor. METHODS: We retrospectively reviewed the medical records of 103 patients who were initially diagnosed with orbital inflammatory pseudotumor, and identified 16 cases in which the diagnosis was based on surgical biopsy and for which data in medical records were sufficient for analysis. Immunohistochemical staining of pathological specimens for IgG and IgG4 was performed. Finally, six of IgG4-related ophthalmic disease patient and 10 of orbital inf lammatory pseudotumor patient were analyzed. RESULTS: The IgG4-related ophthalmic disease group had more IgG4-positive plasma cells and a higher IgG4/IgG plasma cell ratio than the orbital inflammatory pseudotumor group. Collagenous fibrosis and lacrimal gland involvement were significantly more frequent in the IgG4-related ophthalmic disease group. Dense lymphocyte infiltration, obliterative phlebitis, and bilateral lesions were more frequent in IgG4-related ophthalmic disease, but the differences were not significant. The recurrence-free period was shorter in the IgG4-related ophthalmic disease group (p = 0.035). CONCLUSIONS: The location of the lesion (lacrimal gland), count and ratio of IgG4-positive plasma cells, and collagenous fibrosis aid the diagnosis of IgG4-related ophthalmic disease in patients with idiopathic orbital mass-like lesions. In addition, maintenance therapy should be considered in patients with IgG4-related ophthalmic disease to prevent recurrence.

Sodium Chloride Aggravates Arthritis via Th17 Polarization

PURPOSE: Sodium chloride (NaCl) has been proposed as a driving factor in autoimmune diseases through the induction of pathogenic CD4+ T helper cells that produce interleukin-17 (Th17 cells). This study investigated the effects of NaCl on inflammatory arthritis in mice and humans. MATERIALS AND METHODS: Collagen-induced arthritis (CIA) mice were fed a normal or high-salt diet ad libitum, and clinical and histologic features of arthritis were evaluated. The proportion of Th17 cells in the spleens of CIA mice fed a normal or high-salt diet was evaluated by flow cytometry, and the expression of IL-17 in joints and intestines was determined by immunohistochemical staining. We also analyzed the effect of NaCl on Th17 differentiation from peripheral blood monocytes of patients with rheumatoid arthritis (RA) and osteoarthritis (OA) and evaluated the contents of sodium and IL-17 in the synovial fluid of RA and OA patients. RESULTS: NaCl increased murine and human Th17 cell differentiation in a dose-dependent manner. Clinical and histological arthritis was more severe in the high-salt-fed CIA mice, compared to control CIA mice. The proportion of Th17 cells among splenocytes was higher in CIA mice fed a high-salt diet. Expression of synovial and intestinal IL-17 was also higher in high-salt-fed CIA mice. Comparison of synovial fluid between RA patients and OA patients revealed that Na+ and IL-17 were more abundant in RA synovial fluid. CONCLUSION: This study suggests that NaCl can aggravate arthritis by affecting Th17 differentiation. Accordingly, limiting salt intake may be helpful for treating inflammatory arthritis, such as RA.

Rheumatoid arthritis is associated with early tooth loss: results from Korea National Health and Nutrition Examination Survey V to VI

BACKGROUND/AIMS@#To examine the association between rheumatoid arthritis (RA) and periodontitis or tooth loss.@*METHODS@#The study used data from the fifth and sixth Korea National Health and Nutrition Examination Surveys conducted from 2010 to 2015. RA was defined as participant-reported physician-diagnosed RA that was being treated. Periodontitis and the number of natural teeth were determined by dental examination. Periodontitis was defined according to the community periodontal index (periodontal probing depth ≥ 4 mm). The association between RA and periodontitis or tooth loss was examined after controlling for confounding variables (e.g., age, smoking status, socioeconomic status, dental caries, frequency of toothbrushing, body mass index, alcohol consumption, and diabetes) in men and women. Subgroup analyses stratified by age were also performed.@*RESULTS@#The study enrolled 20,297 participants aged ≥ 19 years (157 RA patients and 20,140 non-RA controls). There was no association between RA and periodontitis or tooth loss in men and women. Subgroup analyses in those aged < 60 years revealed a non-significant association between RA and periodontitis (adjusted odds ratio, 1.53; p = 0.162), but they revealed a significant association between RA and tooth loss (adjusted β, 0.20; p = 0.042).@*CONCLUSIONS@#RA was not associated with periodontitis, but was associated with tooth loss in younger adults. Younger RA patients are more likely to suffer tooth loss than general younger population; thus dental management is required.

Risk of malignancy in patients with rheumatoid arthritis after anti-tumor necrosis factor therapy: results from Korean National Health Insurance claims data

BACKGROUND/AIMS@#Inhibition of tumor necrosis factor (TNF) is an effective treatment for rheumatoid arthritis (RA), but safety concerns about malignancy remain. The aim of this study was to evaluate cancer risk in RA patients treated with TNF inhibitors (TNFi), based on Korean Nationwide Health Insurance claims data.@*METHODS@#Patients with seropositive RA were selected from the health insurance database containing all citizens' medical information, based on both RA diagnosis codes and medications. Between 2010 and 2014, RA patients treated with conventional synthetic disease-modifying anti-rheumatic drugs (csDMARDs) and TNFi were enrolled and followed up. We compared the cancer incidence between patients treated with TNFi and csDMARDs using incidence rate ratios (IRRs) after adjustment for age, gender, and observational periods.@*RESULTS@#Of 45,423 selected patients with seropositive RA, 2,337 were treated with TNFi and 43,086 were treated with csDMARDs. The TNFi group was younger and was followed-up for a longer duration. During the observational period, 1,732 and 49 cases of cancer were detected in patients treated with csDMARDs and TNFi, respectively. Old age and male sex were associated with cancer occurrence. Adjusted IRRs for all cancers and common cancers demonstrated that cancer incidence did not differ significantly between the TNFi group and csDMARDs group (IRR = 0.913 for all cancers, p = 0.546).@*CONCLUSIONS@#This study revealed that cancer incidence was similar in RA patients treated with TNFi and csDMARDs. Anti-TNF therapy may be a safe therapeutic option for RA treatment, in terms of malignancy.

Current Stem Cell Therapy for Osteoarthritis / 대한내과학회지

Osteoarthritis is a musculoskeletal disease representative of an aging society. As medical conditions are usually complicated in an aging population, osteoarthritis becomes more frequently encountered in the physician's office. There is a growing need, therefore, for physicians to pay attention to this common orthopedic condition. Cartilage degeneration, arthritic pain, and joint dysfunction are major manifestations of osteoarthritis, and degenerated cartilage is difficult to repair with conventional treatment modalities. Scientists and physicians have developed various therapeutic strategies, including the use of stem cells. Here, we discuss previous and current progress in cartilage regenerative therapy against osteoarthritis.

Korean Guideline for the Prevention and Treatment of Glucocorticoid-induced Osteoporosis

BACKGROUND: To develop guidelines and recommendations to prevent and treat glucocorticoid (GC)-induced osteoporosis (GIOP) in Korea. METHODS: The Korean Society for Bone and Mineral Research and the Korean College of Rheumatology have developed this guideline based on Guidance for the Development of Clinical Practice Guidelines ver. 1.0 established by the National Evidence-Based Healthcare Collaborating Agency. This guideline was developed by adapting previously published guidelines, and a systematic review and quality assessment were performed. RESULTS: This guideline applies to adults aged ≥19 years who are using or plan to use GCs. It does not include children and adolescents. An initial assessment of fracture risk should be performed within 6 months of initial GC use. Fracture risk should be estimated using the fracture-risk assessment tool (FRAX) after adjustments for GC dose, history of osteoporotic fractures, and bone mineral density (BMD) results. All patients administered with prednisolone or an equivalent medication at a dose ≥2.5 mg/day for ≥3 months are recommended to use adequate calcium and vitamin D during treatment. Patients showing a moderate-to-high fracture risk should be treated with additional medication for osteoporosis. All patients continuing GC therapy should undergo annual BMD testing, vertebral X-ray, and fracture risk assessment using FRAX. When treatment failure is suspected, switching to another drug should be considered. CONCLUSIONS: This guideline is intended to guide clinicians in the prevention and treatment of GIOP.

Korean Guideline for the Prevention and Treatment of Glucocorticoid-induced Osteoporosis

OBJECTIVE: To develop guidelines and recommendations to prevent and treat glucocorticoid-induced osteoporosis (GIOP) in Korea. METHODS: The Korean Society for Bone and Mineral Research and the Korean College of Rheumatology developed this guideline based on Guidance for the Development of Clinical Practice Guidelines version 1.0 established by the National Evidence-Based Healthcare Collaborating Agency. This guideline was developed by adapting previously-published guidelines, and a systematic review and quality assessment were conducted. RESULTS: This guideline applies to adults aged 19 years or older who are using or plan to use glucocorticoids (GCs), but does not include children and adolescents. An initial assessment of fracture risk should be performed within 6 months of initial GC use. Fracture risk should be estimated using FRAX (Fracture Risk Assessment Tool) with adjustments for GC dose, previous osteoporotic fracture history, and bone mineral density (BMD) results. All patients taking more than 2.5 mg/day prednisolone or equivalent for more than 3 months are recommended to take adequate calcium and vitamin D. Patients at moderate to high fracture risk should be treated with additional osteoporosis medication. All patients continuing GC therapy should receive an annual BMD measurement, vertebral X-ray, and fracture risk assessment using FRAX. When a treatment failure is suspected, switching to another drug should be considered. CONCLUSION: This guideline is intended to provide guidance for clinicians in prevention and treatment of GIOP.

Effects of Individual Education Using a Treating-to-target Strategy in Patients with Rheumatoid Arthritis

OBJECTIVE: To examine effects of an individual education program using the treating rheumatoid arthritis to target (RA T2T) strategy in patients with moderate-severe rheumatoid arthritis. METHODS: Patients were assigned randomly to an educational intervention (n=33) or conventional care group (n=33). The intervention was a nurse-delivered 9-month educational program consisting of 3 monthly sessions and monthly telephone counseling. The assessments occurred at the baseline and every 3 months in both groups, but only the intervention group completed the 9-month education follow-up. The outcome variables included the disease activity (DAS28), functional disability (KHAQ), fatigue (FACIT-Fatigue), and quality of life (SF-36). Repeated measures ANOVA and a Bonferroni multiple comparison were used to evaluate the outcome variables comparing the groups and follow-up times. RESULTS: Significant interactions were observed between the groups and follow-up times in the disease activity (p=0.041), fatigue (p=0.042), and physical (p=0.006) and mental (p=0.031) health-related quality of life, but there was no significant interaction in the functional disability (p=0.110). Significant differences were noted between the groups at the 9-month period (p=0.048) in disease activity and fatigue, and at the 6-month (p=0.023) and 9-month periods (p=0.027) in the physical health-related quality of life. CONCLUSION: This education program using the RA T2T strategy had significant benefits on the disease activity, fatigue, and quality of life in patients with moderate to severe rheumatoid arthritis, and the results suggested that this contributed to positive clinical outcomes as a good practical nursing intervention.

Interrupting oral infection of Porphyromonas gingivalis with anti-FimA antibody attenuates bacterial dissemination to the arthritic joint and improves experimental arthritis

Rheumatoid arthritis (RA) is a chronic autoimmune disease that typically results in strong inflammation and bone destruction in the joints. It is generally known that the pathogenesis of RA is linked to cardiovascular and periodontal diseases. Though rheumatoid arthritis and periodontitis share many pathologic features such as a perpetual inflammation and bone destruction, the precise mechanism underlying a link between these two diseases has not been fully elucidated. Collagen-induced arthritis (CIA) mice were orally infected with Porphyromonas gingivalis (Pg) or Pg preincubated with an anti-FimA antibody (FimA Ab) specific for fimbriae that are flexible appendages on the cell surface. Pg-infected CIA mice showed oral microbiota disruption and increased alveolar bone loss and had synovitis and joint bone destruction. However, preincubation with FimA Ab led to a significant reduction in the severity of both oral disease and arthritis. Moreover, FimA Ab attenuated bacterial attachment and aggregation on human gingival and rheumatoid arthritis synovial fibroblasts. In addition, we discovered bacteria may utilize dendritic cells, macrophages and neutrophils to migrate into the joints of CIA mice. These results suggest that disrupting Pg fimbriae function by FimA Ab ameliorates RA.

CRISPR-Cas9: a promising tool for gene editing on induced pluripotent stem cells

Recent advances in genome editing with programmable nucleases have opened up new avenues for multiple applications, from basic research to clinical therapy. The ease of use of the technology—and particularly clustered regularly interspaced short palindromic repeats (CRISPR)—will allow us to improve our understanding of genomic variation in disease processes via cellular and animal models. Here, we highlight the progress made in correcting gene mutations in monogenic hereditary disorders and discuss various CRISPR-associated applications, such as cancer research, synthetic biology, and gene therapy using induced pluripotent stem cells. The challenges, ethical issues, and future prospects of CRISPR-based systems for human research are also discussed.

Platelet Indices Are Associated with Disease Activity Scores and the Severity of Sacroiliitis on Magnetic Resonance Imaging in Axial Spondyloarthritis Patients

OBJECTIVE: To investigate the associations among platelet indices, disease activity scores, and inflammatory markers in axial spondyloarthritis, and to determine the relation between platelet indices and inflammation measured on magnetic resonance imaging (MRI). METHODS: The study included 161 patients who fulfilled Assessment of Spondyloarthritis International Society criteria. Platelet indices such as mean platelet volume (MPV), plateletcrit (PCT), platelet large cell ratio (PLCR), and platelet distribution width (PDW) were measured. Ninety patients underwent sacroiliac (SI) MRI at baseline. Bone marrow edema (BME) and erosion on MRI were scored using the SPondyloArthritis Research Consortium of Canada (SPARCC) method. Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI), Ankylosing Spondylitis Disease Activity Score (ASDAS) and spinal radiologic progression were also assessed. The associations among platelet indices and disease activity scores and inflammatory markers were evaluated. RESULTS: Of the 161 patients, 130 (81%) were male. MPV, PLCR, and PDW were negatively associated with ASDAS and inflammatory marker expression, whereas PCT was positively associated with these parameters. MPV, PLCR, and PDW were negatively associated with BME and erosion scores on SI MRI. However, platelet indices were not associated with the BASDAI and BASFI. The mean erythrocyte sedimentation rate, C-reactive protein, and BME and erosion scores were significantly higher in patients with low MPV. Changes in MPV, PCT, and PDW at baseline and after one year were associated with changes in ASDAS and inflammatory marker expression. CONCLUSION: Platelet indices are associated with ASDAS, inflammatory marker levels, and severity of BME and erosion measured on MRI.

Eupatilin Ameliorates Collagen Induced Arthritis

Eupatilin is the main active component of DA-9601, an extract from Artemisia. Recently, eupatilin was reported to have anti-inflammatory properties. We investigated the anti-arthritic effect of eupatilin in a murine arthritis model and human rheumatoid synoviocytes. DA-9601 was injected into collagen-induced arthritis (CIA) mice. Arthritis score was regularly evaluated. Mouse monocytes were differentiated into osteoclasts when eupatilin was added simultaneously. Osteoclasts were stained with tartrate-resistant acid phosphatase and then manually counted. Rheumatoid synoviocytes were stimulated with TNF-alpha and then treated with eupatilin, and the levels of IL-6 and IL-1beta mRNA expression in synoviocytes were measured by RT-PCR. Intraperitoneal injection of DA-9601 reduced arthritis scores in CIA mice. TNF-alpha treatment of synoviocytes increased the expression of IL-6 and IL-1beta mRNAs, which was inhibited by eupatilin. Eupatilin decreased the number of osteoclasts in a concentration dependent manner. These findings, showing that eupatilin and DA-9601 inhibited the expression of inflammatory cytokines and the differentiation of osteoclasts, suggest that eupatilin and DA-9601 is a candidate anti-inflammatory agent.