摘要
目的观察五味子木脂素(SCL)对佐剂性关节炎(AA)大鼠的抗炎作用,并探讨其自噬相关抗炎作用机制。方法大鼠右后足足跖皮下注射弗氏完全佐剂0.2 mL,足肿胀度≥0.8 mL为造模成功。第1天(D1)开始,大鼠ig给予地塞米松(Dex)0.25 mg·kg^-1或SCL 25,50和100 mg·kg^-1,持续28 d。测量D0和D28大鼠体质量,观察D1,D21和D28足跖形态。排水法检测足跖肿胀度;检测胸腺和脾指数;HE染色观察大鼠关节组织病理形态并分别半定量检测炎症细胞、滑膜组织增生和OOCHAS评分;RT-PCR法检测足跖滑膜组织中肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6、IL-17、环氧合酶1(COX-1)和COX-2 mRNA表达水平;RT-PCR法和Western印迹法检测足跖滑膜组织中腺苷酸激活蛋白激酶(AMPK)、哺乳动物雷帕霉素靶蛋白(mTOR)、Beclin1、微管相关蛋白轻链3-Ⅱ(LC3-Ⅱ)、自噬相关基因5(ATG5)、ATG7和ATG12基因及蛋白的表达。结果与正常对照组相比,模型组体质量明显下降(P<0.01);模型组炎症细胞、滑膜组织增生和OOCHAS评分显著升高(P<0.01);COX-1,COX-2,TNF-α,IL-1β,IL-6和IL-17 mRNA表达显著升高(P<0.05,P<0.01),AMPK,Beclin1和ATG12 mRNA和蛋白表达明显降低(P<0.05,P<0.01),mTOR mRNA和蛋白表达显著升高(P<0.01),LC3-Ⅱ,ATG5和ATG7 mRNA表达显著降低(P<0.05,P<0.01)。与模型组相比,模型+SCL 50和100 mg·kg^-1组体质量显著增加(P<0.01)。D1大鼠足跖红肿明显,D21继发性炎症达到高峰。模型+SCL 50和100 mg·kg^-1组大鼠滑膜组织炎症细胞明显减少(P<0.05,P<0.01),滑膜组织增生显著减轻(P<0.05,P<0.01),OOCHAS评分显著降低(P<0.01);模型+SCL 25,50和100 mg·kg^-1组胸腺指数明显降低(P<0.01);模型+SCL 50和100 mg·kg^-1组脾指数明显降低(P<0.05,P<0.01);模型+SCL 50和100 mg·kg^-1组关节软骨损伤明显改善(P<0.01);模型+SCL 25,50和100 mg·kg^-1组COX-1 mRNA表达无显著差异,模型+SCL 100 mg·kg^-1组COX-2,TNF-α和IL-17 mRNA表达明显降低(P<0.05),模型+SCL 50,100 mg·kg^-1组IL-1β和IL-6 mRNA表达明显降低(P<0.05,P<0.01);模型+Dex组COX-1 mRNA表达无显著差异,COX-2,TNF-α,IL-1β,IL-6和IL-17 mRNA表达显著降低(P<0.05,P<0.01);模型+SCL 50和100 mg·kg^-1组AMPK,Beclin1,LC3-Ⅱ,ATG5和ATG7 mRNA表达显著升高(P<0.05,P<0.01),模型+SCL 100 mg·kg^-1组mTOR mRNA表达显著降低(P<0.05),ATG12 mRNA表达水平显著升高(P<0.01);模型+SCL 100 mg·kg^-1组AMPK,Beclin1,LC3-Ⅱ,ATG5和ATG7蛋白表达显著升高(P<0.05,P<0.01),模型+SCL 25,50和100 mg·kg^-1组mTOR蛋白表达显著降低(P<0.05,P<0.01),ATG12表达明显升高(P<0.05,P<0.01)。结论SCL对佐剂性关节炎大鼠炎症反应有抑制作用,其机制可能与促进关节滑膜细胞自噬相关。
OBJECTIVE To observe the effect of Schisandra chinensis lignan(SCL)on adjuvant arthritis(AA)of rats,and to explore its anti-inflammatory mechanism related to autophagy.METHODS Except for the normal control group,rats were injected subcutaneously with Freund's complete adjuvant(CFA)0.2 mL in the right hind foot plantar.After 8 h,the swelling of the inflammed feet of the rats was more than 0.8 mL,which was a successful model.On the first day(D1),the rats of normal control group while the model group were given ig saline,which the others were given dexamethasone 0.25 mg·kg^-1,SCL 25,50,and 100 mg·kg^-1 once a day for 28 d.The body mass of rats on D0 and D28 was recorded.The morphological changes in the rats'inflammed feet and plantar on D1 and D28 were detected.Drainage method was used to detect the plantar swelling of the rats.The thymus and spleen indexes were detected.The pathological changes in the rat joints were observed by HE staining.The mRNA expressions of tumor necrosis factorα(TNF-α),interleukin-1β(IL-1β),IL-6,IL-17,cyclooxygenase-1(COX-1)and COX-2 in the foot and plantar synovial tissue of rats were detected by RT-PCR.The mRNA and protein expressions of adenosine 5'-monophosphate-activated protein kinase(AMPK),mammalian target of Rapamyoin(mTOR),Beclin1,microtubule-associated protein light chain 3-Ⅱ(LC3-Ⅱ),autophagy-related gene-5(ATG5),ATG7 and ATG12 in the plantar synovial tissue of rats were detected by RT-PCR and Western blotting,respectively.RESULTS Compared with the normal control group,the body mass of the model group decreased significantly(P<0.01).Compared with the model group,the body mass of the model+SCL 50 and 100 mg·kg^-1 groups increased significantly(P<0.01).On D24,the redness and swelling of the model rats were obvious,while those of the model+SCL 25,50 and 100 mg·kg^-1 groups were significantly improved.Beginning on D8 of administration,compared with the model group,the swelling of the model+SCL 100 mg·kg^-1 group was significantly reduced(P<0.05,P<0.01).On D24,foot swelling in the model+SCL 50 mg·kg^-1 group was significantly reduced(P<0.05).Compared with the normal control group,the thymus index and spleen index of the model group were significantly increased(P<0.01).Compared with the model group,the thymus index of the model+SCL 25,50 and 100 mg·kg^-1 groups was significantly reduced(P<0.01),so were the spleen indexes of model+SCL 50 and 100 mg·kg^-1 groups(P<0.05,P<0.01).Compared with the normal control group,inflammatory cell infiltration,hyperplasia and OOCHHAS score of the model group increased obviously(P<0.01).Compared with the normal control group,the mRNA expressions of TNF-α,IL-1β,IL-6,IL-17,COX-1 and COX-2 significantly increased(P<0.05,P<0.01).Compared with the model group,those of the model+SCL 100 mg·kg^-1 groups were significantly reduced(P<0.05).Compared with the normal control group,the mRNA expressions of AMPK,Beclin1,LC3-Ⅱ,ATG5,ATG7 and ATG12 in the model group were significantly decreased(P<0.05,P<0.01),so was mTOR mRNA(P<0.01).The protein expressions of AMPK,Beclin1 and ATG12 were decreased significantly(P<0.05,P<0.01);while the mTOR,LC3-Ⅱ,ATG5 and ATG7 proteins increased significantly(P<0.05,P<0.01).Compared with the model group,mTOR mRNA and protein levels were significantly reduced(P<0.05),while ATG12 mRNA expression significantly increased(P<0.05)in model+SCL 100 mg·kg^-1 group.AMPK,Beclin1,LC3-Ⅱ,ATG5 and ATG7 mRNA and protein expressions were significantly increased in model+SCL 50 and 100 mg·kg^-1 groups.CONCLUSION SCL has inhibitory effect on the inflammatory response in adjuvant arthritis rats,and the mechanism may be related to the promotion of autophagy in joint synoviocytes.
作者
范紫微
古虹
邹天琪
柏琳
孙靖辉
张成义
FAN Zi-wei;GU Hong;ZOU Tian-qi;BAI Lin;SUN Jing-hui;ZHANG Cheng-yi(College of Pharmacy,Beihua University,Jilin 132013,China)
出处
《中国药理学与毒理学杂志》
CAS
北大核心
2020年第6期442-450,共9页
Chinese Journal of Pharmacology and Toxicology
基金
吉林省卫生健康委员会(2019J046)
吉林省科学技术厅科技攻关项目(20190304048YY)
吉林市科技计划项目(201820847)
吉林省中医药科技项目(2018111)
北华大学研究生创新计划项(北华研创合字[2019]059)。
关键词
五味子木脂素
细胞自噬
炎症
类风湿性关节炎
Schisandra chinensis lignans
autophagy
inflammation
rheumatoid arthritis
