Fig. 3

EC-18 combined with methotrexate attenuated the severity of collagen-induced arthritis. The vehicle only or EC-18 (125 mg/kg) was administered to mice once daily from day 21 after the first immunization, and methotrexate (1.5 mg/kg) was administered three times a week by oral feeding (n = 10/group). A A graphic scheme of CIA induction and drug administration. B Arthritis development was assessed based on the arthritis score (upper) and incidence (lower). *P < 0.05, **P < 0.01, ***P < 0.001 vs. vehicle-treated control group by two-way ANOVA (Bonferroni posttests). C At 70 days after the first immunization with CII, sections of ankle joint tissues of subjects with the average arthritis score of each group (n = 5/group) were stained with H&E (original magnification ×40) and safranin O (original magnification ×200). Representative histological features are shown. Two independent, blinded observers assessed the cartilage damage, inflammation, and bone damage was assessed on a scale of 0–4 according to the criteria. D At 70 days after the first immunization, serum level of CII-specific IgG2a were measured via enzyme-linked immunosorbent assay (ELISA) (n = 8/group). *P < 0.05, **P < 0.01, ***P < 0.001 vs. vehicle-treated control group