摘要
目的观察三七总皂苷(panax notoginseng saponins,PNS)调控HIF-1α/PDHK1通路介导的有氧氧化,探讨其对促进Naive CD4^(+)T细胞向Treg细胞分化的机制。方法磁珠分选C57BL/6小鼠脾脏Naive CD4^(+)T细胞,诱导其向Treg细胞分化并进行体外培养,将Naive CD4^(+)T细胞分为PNS处理组(5、10、20μg·mL^(-1))、PNS联合HIF-1α抑制剂(PX-478)组,并设置对照组,采用流式细胞术检测Treg细胞分化比例,Western blotting检测HIF-1α、PDHK1蛋白表达,实时荧光定量PCR检测HIF-1α、PDHK1、FOXP3 mRNA的表达,酶联免疫吸附实验检测细胞培养上清液中IL-10的水平。结果PNS可显著增加Treg细胞比例,并增加其IL-10的分泌水平,使细胞中FOXP3 mRNA的表达增加;同时抑制HIF-1α、PDHK1蛋白及mRNA的表达。10μmol·L^(-1)的PX-478处理细胞后再以10μg·mL^(-1)的PNS干预细胞,其对PDHK1、FOXP3的表达及Treg细胞的分化比例的调控作用与单独使用10μmol·L^(-1)的PX-478处理细胞无明显差异。结论PNS通过HIF-1α抑制PDHK1的表达而增强Naive CD4^(+)T细胞的有氧氧化,促进其向Treg细胞分化。
OBJECTIVE To observe the regulation of aerobic oxidation mediated by HIF-1α/PDHK1 pathway by PNS,and to explore its mechanism of promoting the differentiation of Naive CD4^(+)T cells into Treg cells.METHODS Naive CD4^(+)T cells were isolated from the spleen of C57BL/6 mice by magnetic beads and induced to differentiate into Treg cells for in vitro culture.Naive CD4^(+)T cells were divided into PNS treatment group(5,10,20μg·mL^(-1)),PNS combined with HIF-1αinhibitor(PX-478)group,and the control group was set up.The proportion of Treg cells differentiation was detected by flow cytometry.The expression of HIF-1αand PDHK1 protein was detected by Western blotting.The expression of HIF-1α,PDHK1 and FOXP3 mRNA was detected by real-time fluorescence quantitative PCR.The level of IL-10 in cell culture supernatant was detected by enzyme-linked immunosorbent assay.RESULTS PNS could significantly increase the proportion of Treg cells and the secretion level of IL-10,and increase the expression of FOXP3 mRNA in cells.At the same time,the expression of HIF-1αand PDHK1 protein and mRNA was inhibited.When the cells were treated with 10μmol·L^(-1)PX-478 and then treated with 10μg·mL^(-1)PNS,the expression of PDHK1 and FOXP3 and the differentiation ratio of Treg cells were not significantly different from those treated with 10μmol·L^(-1)PX-478 alone.CONCLUSION PNS can reduce the expression level of PDHK1 by HIF-1αto enhance the aerobic oxidation of Naive CD4^(+)T cells and promote their differentiation into Treg cells.
作者
王福荣
颜星宇
葛玉
包宇杰
周玲玲
WANG Furong;YAN Xingyu;GE Yu;BAO Yujie;ZHOU Lingling(Nanjing University of Chinese Medicine,Nanjing 210023,China)
出处
《中国现代应用药学》
CAS
CSCD
北大核心
2024年第8期1015-1020,共6页
Chinese Journal of Modern Applied Pharmacy
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)。
