彝药痹Ⅱ号介导KLF2调控PI3K/Akt/mTOR通路对类风湿性关节炎小鼠模型表达及生物学行为特性影响的机制研究

The Mechanism of YiyaobiⅡMediating KLF2 Regulation of PI3K/Akt/mTOR Pathway on the Expression and Biological Behavior of Rheumatoid Arthritis Mouse Model

目的:探讨彝药痹Ⅱ号介导KLF2调控PI3K/Akt/mTOR通路对类风湿性关节炎小鼠模型表达及生物学行为特性影响的机制。方法:80只昆明种小鼠分为对照组、模型组、彝药痹Ⅱ号组低、高剂量组;除对照组外,其余各组通过左后足趾皮内注射弗氏完全佐剂建立小鼠RA模型;彝药痹Ⅱ号组低、高剂量组于造模成功第1天开始给予相应剂量药物灌胃,持续给予3个月,对照组和模型组给予等体积生理盐水。试验结束后测定小鼠PEL、关节炎症积分、足趾容积;RT-PCR法、蛋白印迹法、免疫组化法测定小鼠踝关节软骨KLF2、PI3K、Akt、mTOR水平。结果:模型组小鼠PEL低于对照组,关节炎症积分、足趾容积高于对照组(P<0.05);彝药痹Ⅱ号低、高剂量组小鼠PEL高于对照组,关节炎症积分、足趾容积低于对照组(P<0.05);且随着彝药痹Ⅱ号剂量的逐渐升高,各剂量组小鼠PEL逐渐升高,关节炎症积分、足趾容积逐渐降低(P<0.05)。模型组小鼠踝关节KLF2、PI3K、Akt、mTOR mRNA和蛋白表达水平高于对照组(P<0.05);与模型组比较,彝药痹Ⅱ号低、高剂量组小鼠踝关节KLF2、PI3K、Akt、mTOR mRNA和蛋白表达水平低于对照组(P<0.05);且随着彝药痹Ⅱ号剂量的逐渐升高,各剂量组小鼠踝关节KLF2、PI3K、Akt、mTOR mRNA和蛋白表达水平逐渐降低(P<0.05)。结论:彝药痹Ⅱ号对小鼠类风湿性关节炎具有明显治疗作用,能明显抑制小鼠踝关节软骨滑膜炎症反应;其机制与彝药痹Ⅱ号抑制类风湿性关节炎小鼠踝关节软骨炎症KLF2-PI3K-Akt-mTOR通路的激活有关。

Objective:To investigate the effect of YiyaobiⅡon the expression and biological behavior of mouse model of rheumatoid arthritis by mediating KLF2 regulation of PI3K/Akt/mTOR pathway.Methods:A total of 80 Kunming mice were divided into control group,model group,YiyaobiⅡgroup,low dose group and high dose group.Except the control group,RA model was established by intradermal injection of Freund's complete adjuvant in the left posterior toe of the other groups.YiyaobiⅡgroup,low,and high dose groups were given corresponding doses of drugs by intragastric administration on the first day of successful modeling for 3 months,and control group and model group were given equal volume of normal saline.After the experiment,PEL,joint inflammation score and toe volume were measured.The levels of KLF2,PI3K,Akt and mTOR were determined by RT-PCR,Western blot and immunohistochemistry.Results:The PEL of the model group was lower than that of the control group,while the arthritis inflammation score and paw volume were higher than those of the control group(P<0.05).In the low-and high-dose YiyaobiⅡgroups,the PEL was higher,and the arthritis inflammation score and paw volume were lower compared to the model group(P<0.05).With increasing doses of YiyaobiⅡ,PEL increased,while the arthritis inflammation score and paw volume decreased(P<0.05).The expression of KLF2,PI3K,Akt,and mTOR mRNA and proteins in the ankle cartilage of the model group was higher than in the control group(P<0.05).In the low-and high-dose YiyaobiⅡgroups,the expression of these markers was lower than in the model group(P<0.05),with expression levels decreasing as the dose increased(P<0.05).Conclusion:YiyaobiⅡhas a significant therapeutic effect on rheumatoid arthritis in mice,notably inhibiting synovitis and inflammation in the ankle cartilage.The mechanism is associated with suppressing the activation of the KLF2-PI3K-Akt-mTOR pathway in rheumatoid arthritis.