Department of Pharmacy, The Affiliated Drum Tower Hospital of Nanjing University Medical School, 321 Zhong Shan Road, Nanjing 210008, Jiang Su, China.
Rheumatol Int. 2013 Jul;33(7):1845-53. doi: 10.1007/s00296-013-2674-7. Epub 2013 Jan 19.
The objectives of this study were to determine the effect of combination of methotrexate (MTX) and leflunomide (LEF) on type II collagen-induced arthritis rats and its mechanism. Curative effect was confirmed on CIA rats, which were randomized and divided into model, MTX, LEF and MTX + LEF group. Weights and joint swelling scores of rats were recorded. Interleukin (IL)-17, receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) concentration in serum were determined by ELISA. H&E dyeing of joint was used to estimate the inflammation and osteoclasia extent. The mechanism was investigated through fibroblast-like synoviocytes isolated from RA patients. The effect of MTX and LEF on cell viability, and RANKL and OPG expression were indicated through MTT and RT-PCR analysis, respectively. Combination therapy would be effective in treating CIA rats. Joint swelling scores and IL-17 and RANKL level in serum were decreased obviously (P < 0.05), while OPG level was elevated (P < 0.05). Anti-inflammatory and anti-osteoclasia effect would be indicated by H&E dyeing results. Moreover, FLS cell viability was inhibited by combination treatment in vitro (P < 0.05), and expression of osteoclasia-related genes (RANKL and OPG) was modified (P < 0.05). Combination therapy would relive the synovium hypertrophy through depressing cell viability and osteoclasia through decreasing RANKL and increasing OPG expression. Otherwise, combination was superior to monotherapy.
本研究旨在探讨甲氨蝶呤（MTX）和来氟米特（LEF）联合治疗对Ⅱ型胶原诱导性关节炎（CIA）大鼠的作用及其机制。采用随机分组方法，将 CIA 大鼠分为模型组、MTX 组、LEF 组和 MTX+LEF 组，观察各组大鼠的疗效，记录体质量和关节肿胀评分，采用 ELISA 法检测血清中白细胞介素（IL）-17、核因子-κB 受体活化因子配体（RANKL）和骨保护素（OPG）的浓度，采用 H&E 染色观察关节炎症和破骨细胞增生程度。分离 RA 患者的成纤维样滑膜细胞（FLS），通过 MTT 法和 RT-PCR 法检测 MTX 和 LEF 对细胞活力和 RANKL、OPG 表达的影响。结果表明，MTX 和 LEF 联合治疗可显著降低 CIA 大鼠的关节肿胀评分和血清中 IL-17 和 RANKL 水平（P<0.05），提高 OPG 水平（P<0.05），减轻关节炎症和破骨细胞增生程度；体外实验结果显示，联合治疗可抑制 FLS 细胞活力（P<0.05），并调节破骨细胞相关基因（RANKL 和 OPG）的表达（P<0.05）。综上，MTX 和 LEF 联合治疗可能通过抑制滑膜细胞增殖、减少 RANKL 表达和增加 OPG 表达，从而缓解 CIA 大鼠的滑膜增生，减轻关节炎症状。
