三七总皂苷调控mTORC1-HIF1α通路干预Th17/Treg分化平衡的作用研究  

Study on effect of panax notoginseng saponins treatment in Th 17/Treg cells differentiation balance through regulating mTORC 1-HIF 1αpathway

作　　者：包宇杰 申美玉 狄昱希 王福荣 周玲玲[1] BAO Yujie;SHEN Meiyu;DI Yuxi;WANG Furong;ZHOU Lingling(School of Pharmacy,Nanjing University of Chinese Medicine,Nanjing 210023,China)

机构地区：[1]南京中医药大学药学院,南京210023

出　　处：《中国免疫学杂志》2024年第11期2310-2315,共6页Chinese Journal of Immunology

基　　金：国家自然科学基金项目(82174306);江苏省高等学校基础科学(自然科学)重大项目(21KJA360009)。

摘　　要：目的:分析三七总皂苷(PNS)对mTORC1-HIF1α信号通路的作用,探讨其对CD4^(+)T细胞中Th17/Treg细胞分化平衡的影响机制。方法:磁珠分选C57BL/6小鼠脾脏CD4^(+)T细胞进行体外培养,采用CCK-8筛选PNS最佳给药浓度,之后分别设置对照组、PNS给药组(5、10、20μg/ml),各组给予相应药物处理48 h后,采用流式细胞术检测Th17/Treg细胞分化,实时荧光定量PCR检测RORγt、Foxp3、mTOR、Raptor、HIF1α mRNA的表达,ELISA检测细胞培养上清中IL-17A和IL-10的水平,Western blot检测4EBP1、S6K、HIF1α蛋白表达及磷酸化水平。结果:5、10、20μg/ml PNS显著抑制Th17细胞分化,促进Treg细胞分化;5、10、20μg/ml PNS显著下调CD4^(+)T细胞中RORγt mRNA表达,降低IL-17A水平;20μg/ml PNS显著上调Foxp3 mRNA表达,提高IL-10水平;10、20μg/ml PNS显著降低4EBP1、S6K磷酸化水平;5、10、20μg/ml PNS显著下调HIF1α mRNA表达,显著抑制HIF1α蛋白表达。结论:一定浓度的PNS可以抑制CD4^(+)T细胞中Th17细胞分化,促进Treg细胞分化,这一效应与影响mTORC1-HIF1α信号通路有关。Objective:To analyze the effect of panax notoginseng saponins(PNS)on mTORC1-HIF1αsignaling pathway,and to explore its effect and mechanisms on the differentiation balance of Th17/Treg cells in CD4^(+)T cells.Methods:Isolate the spleens of C57BL/6 mice,then select CD4^(+)T cells by magnetic beads and cultured in vitro.The optimal concentration of PNS was screened by the CCK-8,and then these cells were divided into control group and PNS treatment group(5,10 and 20μg/ml),each gives correspond-ing drug treatment after 48 h.Afterwards,flow cytometry was used to detect differentiation of Th17/Treg cells.Real-time quantitative fluorescent PCR was used to detect the expressions of RORγt,Foxp3,mTOR,Raptor,HIF1αmRNA.ELISA was used to detect the levels of IL-17A and IL-10 in the supernatant of cell culture.Western blot was used to detect the expressions and phosphorylation levels of 4EBP1,S6K and HIF1αproteins.Results:5,10,20μg/ml PNS could significantly inhibit Th17 cells differentiation and promote Treg cells differentiation;5,10,20μg/ml PNS could significantly reduce the expression of RORγt mRNA,and then reduce the level of IL-17A;20μg/ml PNS could significantly promote the expression of Foxp3 mRNA and increase the level of IL-10;10,20μg/ml PNS could significantly decrease the phosphorylation of 4EBP1 and S6K;5,10,20μg/ml PNS could significantly reduce the expression of HIF1αmRNA and inhibit the expression of HIF1αprotein.Conclusion:Certain concentrations of PNS can inhibit the differentiation of Th17 cells in CD4^(+)T cells,and promote the differentiation of Treg cells,which is related with modulating mTORC1-HIF1αsignaling pathway.

关 键 词：三七总皂苷 TH17 TREG mTORC1 HIF1Α 

分 类 号：R285.5[医药卫生—中药学]