氮网球花定碱盐酸盐对活化T细胞增殖的抑制作用  

Inhibition of N-methylhemeanthidine hydrochloride on activated T cell proliferation

作　　者：邓弘仙 吴利红 张云惠 范诗逸 黄玲 蒋海静 赖翼[3] 杨淑霞 刘阳 罗兴燕 DENG Hong-xian;WU Li-hong;ZHANG Yun-hui;FAN Shi-yi;HUANG Ling;JIANG Hai-jing;LAI Yi;YANG Shu-xia;LIU Yang;LUO Xing-yan(School of Pharmacy,Chengdu Medical College,Chengdu 610500,China;School of Clinical Medicine,Chengdu Medical College,Chengdu 610500,China;School of Laboratory Medicine,Chengdu Medical College,Chengdu 610500,China;Research Center,Chengdu Medical College,Chengdu 610500,China)

机构地区：[1]成都医学院药学院,四川成都610500 [2]成都医学院临床医学院,四川成都610500 [3]成都医学院检验医学院,四川成都610500 [4]成都医学院科研实验中心,四川成都610500

出　　处：《中国药理学与毒理学杂志》2021年第1期43-50,共8页Chinese Journal of Pharmacology and Toxicology

基　　金：大学生创新创业训练计划项目(S201913705068);植物化学与西部植物资源持续利用国家重点实验室开放项目(P2018-KF03);四川省教育厅重点项目(18ZA0143)。

摘　　要：目的探究氮网球花定碱盐酸盐(NMHC)抑制活化T细胞增殖的作用机制。方法Ficoll法提取分离健康人外周血单个核细胞(PBMC),免疫磁珠法纯化出PBMC中的T细胞,抗人CD3/CD28抗体活化纯化后的静息T细胞。NMHC 0.125,0.5,2和8μmol·L^(-1)与静息T细胞作用,用流式细胞术检测细胞毒性(96 h)、活化T细胞增殖(96 h)、T细胞表面活化标志CD25表达(24 h)和活化T细胞细胞周期(72 h)。NMHC 0.125,0.5,2和8 pmol·L^(-1)与活化T细胞作用24或48 h,用ELISA测定培养上清中白细胞介素2(IL-2)、IL-6、IL-17A和干扰素γ(IFN-γ)水平。结果NMHC 0.125,0.5,2和8μmol·L^(-1)抑制抗CD3/CD28抗体活化的T细胞增殖,IC_(50)为(0.28±0.04)μmol·L^(-1),且与静息T细胞作用96 h无显著细胞毒性。NMHC显著抑制CD25表达,具有浓度效应关系(r=-0.980,P<0.05)。NMHC 2和8μmol·L^(-1)作用72 h可调节细胞周期,G_(0)/G_(1)期细胞增加(P<0.05),S期细胞减少(P<0.05),具有浓度-效应关系(r=0.920,P<0.05);细胞上清液中IL-2,IL-6,IL-17A和IFN-γ水平显著降低(P<0.05)。结论NMHC可抑制T细胞活化标志分子CD25的表达和IL-2的分泌,阻滞细胞周期于G_(0)/G_(1)期,同时抑制炎性细胞因子IL-6,IL-17A和IFN-γ的分泌,这可能是其抑制T细胞增殖的作用机制之一。OBJECTIVE To investigate the mechanism by which N-methylhemeanthine hydrochloride(NMHC)inhibits the proliferation of activated T cells.METHODS Healthy peripheral blood mononuclear cells(PBMCs)were extracted and isolated by Ficoll,human peripheral blood T cells were purified by immunomagnetic microbeads from PBMCs,and purified naive T cells were activated by anti-human CD3/CD28 mAbs.Flow cytometry analysis was used to detect the cytotoxicity of naive T cells at 96 h,the proliferation of activated cells at 96 h,the expression of T cell surface activation markers and the cell cycle of activated cells at 72 h with NMHC 0.125,0.5,2 and 8μmol L^(-1).The cytokine levels of interleukin-2(IL-2),IL-6,IL-17 A and interferon-γ(IFN-γ)in the cultured supernatant of activated T cells at24 or 48 h were determined by ELISA.RESULTS NMHC obviously inhibited the proliferation of human T cells activated by anti-human-CD3/CD28 mAbs.IC50 was(0.28±0.04)μmol·L^(-1),and there was no significant cytotoxicity in naive T cells at 8μmol·L^(-1) after 96 h.NMHC significantly inhibited CD25 expression with a concentration-effect relationship(r=-0.980,P<0.05).NMHC 2 and 8μmol·L^(-1) arrested cell cycle in G0/G1 phase(P<0.05)for 72 h,and reduced the percentage of S phase cells(P<0.05)also in a concentration-dependent manner(r=0.920,P<0.05).The levels of IL-2,IL-6,IL-17 A and IFN-γsignificantly decreased in the supernatant(P<0.05).CONCLUSION NMHC inhibits the expression of activated T cell marker molecular CD25 and the secretion of IL-2,arrests cell cycle in G0/G1 phase,and inhibits secretion of IL-6,IL-17 A and IFN-y inflammatory cytokines,which may be one of the mechanisms of its inhibition of cell proliferation.

关 键 词：氮网球花定碱盐酸盐 T细胞 细胞增殖 免疫抑制 

分 类 号：R285.5[医药卫生—中药学] R979.5[医药卫生—中医学]