芍药苷调节RhoA/ROCK信号通路对实验性自身免疫性葡萄膜炎小鼠Th17/Treg免疫平衡的影响  

Effect of Paeoniflorin on Th17/Treg Immune Balance in Experimental Autoimmune Uveitis Mice by Regulating RhoA/ROCK Signaling Pathway

作　　者：庞彬彬 陈震[1] 邢怡桥[1] PANG Binbin;CHEN Zhen;XING Yiqiao(Department of Ophthalmology,Renmin Hospital of Wuhan University,Wuhan 430060 Hubei,China)

机构地区：[1]武汉大学人民医院眼科中心,湖北武汉430060

出　　处：《中药新药与临床药理》2024年第4期506-512,共7页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：湖北省卫生健康委员会科研项目(WJ2021M156)。

摘　　要：目的探讨芍药苷调节Ras同源基因家族蛋白A(RhoA)/Rho相关卷曲螺旋形成蛋白激酶(ROCK)信号通路对实验性自身免疫性葡萄膜炎(experimental autoimmune uveitis,EAU)小鼠辅助性T淋巴细胞17(Th17)/调节性T细胞(Treg)免疫平衡的影响。方法随机选出10只小鼠作为正常对照组,其余小鼠构建EAU模型,将造模成功小鼠随机平分为模型组、芍药苷组(10 mg·kg^(-1)芍药苷)、RhoA/ROCK信号通路激活剂溶血磷脂酸(LPA)组(25μmol·L^(-1)LPA)、芍药苷+LPA组(10 mg·kg^(-1)芍药苷+25μmol·L^(-1)LPA),每组10只。模型组和正常对照组给予等量生理盐水,给药每天1次,持续14 d。眼前节临床表现评分评估炎症严重程度;HE染色观察眼球组织病理形态;ELISA法检测血清中炎性因子转化生长因子-β(trarsforming growth factor beta,TGF-β)、白细胞介素10(IL-10)、白细胞介素17(IL-17)、白细胞介素23(IL-23)水平;流式细胞术检测脾脏Th17/Treg细胞水平;Western Blot法检测维甲酸相关核孤儿受体γt(RORγt)、叉头状转录因子3(Foxp3)以及RhoA/ROCK信号通路蛋白表达。结果正常对照组小鼠无虹膜、结膜血管舒张充血,与正常对照组比,模型组小鼠出现虹膜血管舒张充血,前房、虹膜、睫状体和视网膜内均有大量炎性细胞膨胀,眼前节临床表现评分、IL-17和IL-23含量、Th17细胞水平、Th17/Treg、RORγt、RhoA、ROCK1蛋白水平明显升高(P<0.05),IL-10和TGF-β含量、Treg细胞水平、Foxp3水平明显下降(P<0.05)。与模型组比,芍药苷组小鼠虹膜血管充血症状明显缓解,眼前节临床表现评分、IL-17和IL-23含量、Th17细胞水平、Th17/Treg、RORγt、RhoA、ROCK1蛋白水平明显降低(P<0.05),IL-10和TGF-β含量、Treg细胞水平、Foxp3水平明显升高(P<0.05);而LPA加重了眼部损伤(P<0.05),逆转了芍药苷对EAU小鼠的改善效果(P<0.05)。结论芍药苷可通过下调RhoA/ROCK信号通路调节EAU小鼠Th17/Treg免疫平衡减轻EAU小鼠葡萄膜炎。To investigate the effect of paeoniflorin on the immune balance of T-helper 17 lymphocytes(Th17)/regulatory T cells(Treg)in experimental autoimmune uveitis(EAU)mice by regulating the Ras homolog gene family member A(RhoA)/Rho-associated coiled-coil containing kinase(ROCK)signaling pathway.Methods Ten mice were randomly selected as the control group,and the remaining mice were used to construct an EAU model.The successful modeling mice were randomly divided into model group,paeoniflorin group(10 mg·kg^(-1)paeoniflorin),LPA group(25μmol·L^(-1)LPA,activator of RhoA/ROCK signaling pathway),and paeoniflorin+LPA group(10 mg·kg^(-1)paeoniflorin+25μmol·L^(-1)LPA),with 10 mice in each group.The model and control groups were given equal amounts of physiological saline,once a day for 14 days.The clinical minifestation score of the anteior segment of the eye score was applied to assess the severity of inflammation.HE staining was applied to observe the pathological morphology of eyeball tissue.ELISA method was applied to detect the levels of inflammatory factors trarsfornning growth factor beta(TGF-β),interleukin 10(IL-10)and interleukin 17(IL-17)and interleukin-23(IL-23)in serum.Flow cytometry was applied to detect the level of splenic Th17/Treg cells.Western Blot was applied to detect the expression of retinoid-related orphan nuclear receptorγt(RORγt),forkhead box P3(Foxp3),and RhoA/ROCK signaling pathway proteins.Results The mice in the control group had no iris and conjunctival vasodilation and congestion.When compared with the control group,the mice in the model group had iris vasodilation and congestion,and a number of inflammatory cells in the anterior chamber,iris,ciliary body,and retina were found.The clinical minifestation score of the anteior segment of the eye score,the content of IL-17 and IL-23,the level of Th17 cells,Th17/Treg and the protein levels of RORγt,RhoA,and ROCK1 were obviously increased(P<0.05).The content of IL-10 and TGF-β,the levels of Treg cells and Foxp3 were obviously decreased(P<0.

关 键 词：芍药苷 RhoA/ROCK信号通路 自身免疫性葡萄膜炎 TH17/TREG 小鼠 

分 类 号：R285.5[医药卫生—中药学]