祛痹方对胶原诱导性关节炎大鼠氧自由基代谢及低氧诱导因子-1α水平的影响

目的观察祛痹方对Ⅱ型胶原诱导的免疫性关节炎模型大鼠氧自由基代谢物超氧化物歧化酶（superoxide dismutase, SOD）、丙二醛（malondialdehyde, MDA）、谷胱甘肽过氧化物酶（glutathione peroxidase, GSH Px）及低氧诱导因子（hypoxia inducible factor, HIF）-1α的影响，探讨祛痹方对类风湿关节炎（rheumatoid arthritis, RA）的治疗作用及其机制。方法选用SPF级雄性SD大鼠72只，随机选取12只作为空白对照组，其余60只采用尾根部皮下注射Ⅱ型胶原与不完全弗氏佐剂的方法诱导制备关节炎模型，免疫15 天后，将造模大鼠随机分为胶原诱导性关节炎（collagen induced arthritis, CIA）模型组、雷公藤多苷治疗组、祛痹方高剂量及低剂量组，每组15只，予以灌胃给药，祛痹方高剂量组：6.66 g/（kg· d）；祛痹方低剂量组：3.33 g/（kg· d）；雷公藤多苷组：9.68 mg/（kg· d）；空白对照组和CIA模型组大鼠灌服等体积纯净水。每日1次，连续4周。饲养期间测量大鼠踝关节肿胀度。治疗4周后处死动物，采用比色法测定血浆中SOD、MDA 和GSH Px 活性；采用免疫组化法检测大鼠膝关节HIF-1α的表达。结果（1）与CIA模型组比较，祛痹方高剂量组与雷公藤多苷组大鼠关节肿胀度均显著减轻（P<0.01，P<0.05），且祛痹方高剂量组关节肿胀度明显轻于雷公藤多苷组（P<0.05）；（2）与CIA模型组比较，雷公藤多苷组及祛痹方高、低剂量组大鼠血浆GSH-Px活性明显升高，MDA活性降低，雷公藤多苷和祛痹方高剂量组大鼠血浆SOD活性明显升高（均P<0.05）；（3）与CIA模型组比较，祛痹方高剂量组及雷公藤多苷组大鼠膝关节HIF-1α表达水平明显降低（P<0.05），祛痹方低剂量组有下降趋势，但差异无统计学意义（P>0.05）。结论袪痹方可显著减轻CIA模型大鼠踝关节肿胀程度，其疗效优于雷公藤多苷。其作用机制可能是通过下调关节HIF-1α表达及调节抗氧化水平，从而达到治疗CIA的作用。

OBJECTIVE: To observe the effect of Qubi Recipe (QR) on the expression of superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH-Px), and hypoxia-inducible factor (HIF)-1alpha in rats with type II collagen-I induced arthritis (CIA), and to explore its therapeutic roles and mechanism.

METHODS: Totally 72 male SD rats of SPF grade were recruited. Twelve were randomly selected as the blank control group. The CIA model was established in the rest 60 rats by subcutaneously injecting type II collagen of bovine emulsion from the tail root and induction of incomplete Freund's adjuvant. On day 15 after primary immunization rats were randomly divided into four groups, i.e., the CIA model group, the Tripterygium Glycosides (TG) group (at the daily dose of 9.68 mg/kg body weight), the high dose QR group (at the daily dose of 6.66 g/kg body weight), and the low dose QR group (at the daily dose of 3.33 g/kg body weight), 15 in each group. Corresponding medication was given to rats in all groups by gastrogavage once daily for 4 successive weeks. An equal volume of pure water was given to rats in the blank control group and the CIA model group by gastrogavage, once daily for 4 successive weeks. The swelling degree of the joints was measured. Rats were sacrificed after 4-week treatment. Plasma levels of SOD, MDA, and GSH-Px were measured with colorimetric method. The expression of HIF-1alpha was detected by immunohistochemistry.

RESULTS: (1) Compared with the CIA model group, the swelling degree of the joints was significantly alleviated in the TG group and the high dose QR group (P < 0.01, P < 0.05), and it was obviously milder in the high dose QR group than in the TG group (P < 0.05). (2) Compared with the CIA model group, the activities of GSH-Px could be obviously elevated and activities of MDA lowered in the TG group, the high dose QR group, and the low dose QR group (P < 0.05). Plasma activities of SOD could be obviously elevated in the high dose QR group and the TG group (P < 0.05). (3) Compared with the CIA model group, the expression of HIF-1alpha obviously decreased in the TG group and the high dose QR group (P < 0.05), and it showed a decreasing tendency in the low dose QR group with no statistical difference (P > 0.05).

CONCLUSIONS: QR could markedly alleviate the swelling degree of ankle joints in CIA model rats. Its therapeutic efficacy was superior to that of TG. Its mechanism might be achieved through down-regulating expression of HIF-1alpha in the joint, and regulating activities of SOD, MDA and GSH-Px in the plasma.