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目的观察温化蠲痹方对胶原诱导性关节炎(collagen-inducing arthritis,CIA)大鼠外周血单核细胞(peripheral blood mononuclear cells,PBMCs)DNA甲基化转移酶(DNA methyltrsansferases,DNMTs)表达的影响,探讨温化蠲痹方治疗CIA的作用机制。方法将90只Wistar大鼠随机分为造模组(80只)及正常对照组(10只,简称正常组)。造模组在大鼠尾根部注射牛Ⅱ型胶原乳剂制备CIA模型。将造模成功的50只大鼠随机分为模型组、甲氨喋呤组(MTX)、温化蠲痹方低、中、高剂量组(简称中药低、中、高剂量组),每组10只。模型组给予生理盐水灌胃;中药低、中、高剂量组分别给予温化蠲痹方22.9、45.8、68.7 g/(kg·d)灌胃,均每日1次;MTX组按0.78 mg/kg剂量灌胃MTX混悬液,每周1次,连续30日。采用容积法(排水体积)评价足趾肿胀度。给药干预后提取各组大鼠PBMCs,采用实时定量PCR法检测PBMCs DNMTs(DNMT1、DNMT3a及DNMT3b)mRNA表达水平。结果与正常组比较,模型组大鼠足趾明显肿胀(P<0.01);与模型组比较,中药低、中、高剂量组及MTX组大鼠足趾肿胀明显减轻(P<0.01)。与本组治疗前比较,中药各剂量组及MTX组大鼠足趾肿胀减轻(P<0.01)。与正常组比较,模型组大鼠PBMCs DNMT1、DNMT3a及DNMT3b表达明显降低(P<0.01);与模型组比较,中药各剂量组及MTX组DNMT1、DNMT3a及DNMT3b表达水平明显升高(均P<0.01);中药各剂量组DNMT1、DNMT3a及DNMT3b表达水平比较,差异无统计学意义(P>0.05)。结论 CIA大鼠PBMCs DNMTs表达降低。温化蠲痹方上调CIA大鼠PBMCs DNMTs表达无明显剂量依赖性。通过上调DNMTs表达,调节CIA大鼠甲基化状态可能是其治疗CIA作用机制之一。
Objective To observe the effect of Wenhua Pianbi recipe on the expression of DNA methyltransferases (DNMTs) in peripheral blood mononuclear cells (PBMCs) induced by collagen-inducing arthritis (CIA) Influence, explore the mechanism of Wenhua Weibi treatment of CIA. Methods Ninety Wistar rats were randomly divided into model group (n = 80) and normal control group (n = 10). The model group was made by injecting cow Ⅱ collagen emulsion into the tail of rats. The 50 successful rats were randomly divided into model group, methotrexate group (MTX), Wenliaobi Bi Fang low, middle and high dose group (referred to as low, medium and high dose of traditional Chinese medicine group), each group 10 only The rats in the model group were given saline intragastrically. The rats in low, medium and high dose groups were given gavage of 22.9, 45.8 and 68.7 g / (kg · d) kg dose gavage MTX suspension once a week for 30 days. Volume method (drainage volume) evaluation of toe swelling. PBMCs from each group were extracted after administration of the drug, and the mRNA expression levels of DNMTs (DNMT1, DNMT3a and DNMT3b) in PBMCs were detected by real-time quantitative PCR. Results Compared with the normal group, the toe of the model group was significantly swollen (P <0.01). Compared with the model group, the toe swelling of the low, medium and high dose groups and the MTX group were significantly reduced (P <0.01). Compared with the group before treatment, the rats in each dose group and MTX group had less toe swelling (P <0.01). Compared with normal group, the expressions of DNMT1, DNMT3a and DNMT3b in model group were significantly decreased (P <0.01). Compared with model group, the expressions of DNMT1, DNMT3a and DNMT3b in each dose group and MTX group were significantly increased (all P < 0.01). There was no significant difference in the expression levels of DNMT1, DNMT3a and DNMT3b in each dose group (P> 0.05). Conclusion The expression of DNMTs in PBMCs of CIA rats decreased. No significant dose-dependent effect of Wenhua Baipi on DNMTs expression in PBMCs of CIA rats. Regulating the methylation status of CIA rats may be one of its mechanisms in the treatment of CIA by up-regulating the expression of DNMTs.