益气解毒复方含药血清对胸腺增生型重症肌无力mTEC和Treg细胞增殖的影响  

Effect of Yiqi Jiedu Prescription-containing Serum on Proliferation of mTEC and Regulatory T Cells in Myasthenia Gravis Patients with Thymus Hyperplasia

作　　者：陈森林 况时祥 刘建飞 赵芝兰 郑雄 邹进 刘爽[3] CHEN Sen-lin;KUANG Shi-xiang;LIU Jian-fei;ZHAO Zhi-lan;ZHENG Xiong;ZOU Jin;LIU Shuang(Guizhou University of Traditional Chinese Medicine(TCM),Guiyang 550025,China;The Second Affiliated Hospital of Guizhou University of TCM,Guiyang 550003,China;Capital Medical University,Beijing 100069,China)

机构地区：[1]贵州中医药大学,贵阳550025 [2]贵州中医药大学第二附属医院,贵阳550003 [3]首都医科大学,北京100069

出　　处：《中国实验方剂学杂志》2022年第4期68-75,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81760815);贵州省科技厅项目(黔科合平台人才[2018]5605);贵州省教育厅青年科技人才成长项目(黔科合KY[2017]174);贵州省中医药管理局中医药、民族医药科学技术研究课题(QZYY-2015-055)。

摘　　要：目的:探讨益气解毒复方含药血清对胸腺增生型重症肌无力(MG)患者胸腺髓质上皮细胞(mTEC)和调节性T细胞(Treg)增殖的影响。方法:采用血清药理学方法,将35只SD大鼠适应性喂养1周后随机分为益气解毒复方低、中、高剂量组,空白组,强的松组,每组7只。大鼠连续灌胃1周,制备上述各组含药血清。利用细胞增殖与活性检测(CCK-8)法观察不同浓度益气解毒复方含药血清对mTEC和Treg细胞增殖的作用,并通过共同培养方式观察mTEC及益气解毒复方含药血清对Treg细胞增殖的作用。结果:胸腺细胞经培育,使用mTEC特征性标记荆豆凝集素Ⅰ(UEAI)做流式检测阳性率均值为92.54%。采用磁珠分选Treg细胞,多次磁珠分选后的Treg细胞纯度高达92%,mTEC和Treg细胞均呈较高阳性率表达,细胞为mTEC和Treg细胞,细胞纯度符合后续实验的要求。益气解毒复方含药血清体积分数2.5%~15%时,对mTEC和Treg细胞均有抑制作用,益气解毒复方含药血清浓度≥20%时对细胞的增殖有促进作用,且随作用的时间延长,含药血清对细胞的增殖的促进作用越为明显。培养48 h后的结果显示,与空白组比较,强的松组和益气解毒复方低剂量组差异无统计学意义,中、高剂量组细胞增殖抑制率显著降低(P<0.01)。与mTEC共同培养的Treg细胞,与空白组比较,强的松组、益气解毒复方低剂量组Treg细胞增殖抑制率差异无统计学意义,中、高剂量组细胞增殖抑制率显著降低(P<0.01)。结论:益气解毒复方含药血清能够影响胸腺增生型MG患者mTEC和Treg细胞的增殖,独立培养MG患者的Treg细胞相比,与mTEC共同培养的MG患者Treg细胞增殖具有促进作用,提示mTEC能够调控Treg细胞的增殖,且这种作用在予以益气解毒复方含药血清干预之后更为明显,说明益气解毒复方对Treg细胞的干预,可以通过对mTEC的治疗作用产生,这可能是益气解毒复方治疗MG的作用机制之一。Objective:To investigate the effect of Yiqi Jiedu prescription-containing serum on the proliferation of medullary thymic epithelial cells(mTEC)and regulatory T(Treg)cells in myasthenia gravis(MG)patients with thymus hyperplasia.Method:According to serological methods,35 SD rats were adaptively fed for one week and randomized into the low-,medium-,and high-dose Yiqi Jiedu prescription groups,control group,and prednisone group,with seven rats in each group,which were then gavaged with the corresponding drugs for one week for preparing the drug-containing serum.The effect of Yiqi Jiedu prescription-containing serum at different concentrations on the proliferation of mTEC and Treg cells were determined by cell counting kit-8(CCK-8)assay.Besides,the effect of mTEC and Yiqi Jiedu prescription-containing serum on Treg cell proliferation were observed through co-culture.Result:Thymocytes were cultured for a period of time.Their mean positive rate revealed by flow cytometry using mTEC characteristic marker Ulex europaeus agglutininⅠ(UEAI)was 92.54%.Treg cells were sorted by magnetic beads.The purity of Treg cells after repeated magnetic bead sorting was as high as 92%.mTEC and Treg cells showed high positive expression rates,and their cell purity met the requirements of subsequent experiments.When the concentration of Yiqi Jiedu prescriptioncontaining serum was 2.5%-15%,it exhibited an inhibitory effect against mTEC and Treg cells.When the concentration was equal to or greater than 20%,it promoted cell proliferation,which was further enhanced with the extension of action time.The results after 48 h of culture showed that compared with the control group,prednisone and low-dose Yiqi Jiedu prescription had no significant effect on the proliferation of these two kinds of cells,but the medium-and high-dose Yiqi Jiedu prescription remarkably reduced their proliferation inhibition rate(P<0.01).After co-culture with mTEC,the control group was not significantly different from the prednisone group and the low-dose Yiqi Jiedu pres

关 键 词：益气解毒复方 重症肌无力 含药血清 胸腺髓质上皮细胞 调节性T细胞 增殖 

分 类 号：R22[医药卫生—中医基础理论] R242[医药卫生—中医学]