知母皂苷元对H2O2损伤SH-SY5Y细胞中脑源性神经营养因子的调控及机制研究  

Regulation on brain-derived neurotrophic factor and relevant mechanism of anemarrhena saponin in H2O2-induced SH-SY5Y cells

作　　者：杨双双 高天行[2] 何萱 张瑞[1] 张永芳[1,3,4] YANG Shuang-shuang;GAO Tian-xing;HE Xuan;ZHANG Rui;ZHANG Yong-fang(Shanghai Jiao Tong University College of Basic Medical Sciences,Shanghai 200025,China;Department of Pharmacy,Xinhua Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200092,China)

机构地区：[1]上海交通大学基础医学院,上海200025 [2]上海交通大学医学院附属新华医院药剂科,上海200092 [3]上海市核学会 [4]上海市中西医结合学会青年委员会

出　　处：《上海交通大学学报（医学版）》2019年第6期578-585,578,共8页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金(81573401);上海市高水平地方高校建设项目(XD18015)~~

摘　　要：目的·观察知母皂苷元(anemarrhena saponin,ZMS)对H2O2损伤人神经母细胞瘤细胞SH-SY5Y中脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)的调控作用,并初步探索其分子机制。方法·用H2O2损伤SH-SY5Y细胞建立氧化应激细胞模型。通过CCK-8试剂盒检测经H2O2损伤后的SH-SY5Y细胞活力。采用实时荧光定量PCR(quantitative real-time PCR,qPCR)检测BDNF及其重要转录子的mRNA水平。利用组蛋白脱乙酰酶(histone deacetylases,HDACs)活性荧光分析试剂盒检测ZMS对HDACs活性的影响。采用蛋白质印迹(Western blotting)检测乙酰化组蛋白H3、H4,特定乙酰化位点相关蛋白及HDAC1/2/3蛋白的表达水平。结果·qPCR检测显示,ZMS可以上调受损SH-SY5Y细胞中BDNF及转录子Ⅳ的mRNA水平。Western blotting检测显示,ZMS预处理后可增加乙酰化组蛋白H3、H4、H3K14的蛋白水平,而对HDAC1/2/3蛋白的表达无显著性影响。HDACs活性荧光分析试剂盒检测结果显示,ZMS可抑制HDACs的活性。结论·ZMS可增加氧化应激细胞模型中BDNF及转录子Ⅳ的mRNA水平,其作用机制可能与调节组蛋白乙酰化水平相关。Objective·To investigate the effect of anemarrhena saponin (ZMS) on mRNA level of brain-derived neurotrophic factor (BDNF) and relevant mechanism in oxidative stress damage of SH-SY5Y cells.Methods·SH-SY5Y cells treated with H2O2 were chosen as cell models of oxidative stress.Cell viability was determined using cell counting kit-8 (CCK-8).The mRNA levels of BDNF and its important transcripts were detected by quantitative real-time PCR (qPCR).The histone deacetylases (HDACs) activity fluorescence quantification assay kit was used to measure the effect of ZMS on HDACs activity.Western blotting was used to detect the protein expression levels of acetylated histone H3,acetylated histone H4,specific acetylation site-related proteins,and HDAC1/2/3.Results·qPCR showed that ZMS could increase the mRNA levels of BDNF and its transcript Ⅳ in the cell models.Western blotting showed that ZMS pretreatment could increase the protein levels of acetylated histone H3,acetylated histone H4 and acetylated histone H3K14,and there was no significant effect on protein levels of HDAC1/2/3.In addition,HDACs activity fluorescence quantification assay kit showed that ZMS could inhibit HDACs activity significantly.Conclusion·ZMS can increase the mRNA levels of BDNF and its transcript Ⅳ in oxidative stress damage cell models,which may be related to the regulation of histone acetylation level.

关 键 词：知母皂苷元 氧化应激 脑源性神经营养因子 组蛋白乙酰化 

分 类 号：R285.5[医药卫生—中药学]