分子营养 Molecular nutrition

硒蛋白x基因沉默和过氧化氢处理对人正常肝脏细胞中硒蛋白基因表达和抗氧化酶活性的影响

  • 汤加勇 ,
  • 赵华 ,
  • 贾刚 ,
  • 刘光芒 ,
  • 陈小玲 ,
  • 蔡景义 ,
  • 吴彩梅
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  • 四川农业大学动物营养研究所, 成都 611130

收稿日期: 2016-02-28

  网络出版日期: 2016-08-17

基金资助

国家自然科学基金面上项目(31072043,31272468);四川隆达畜牧科技有限公司项目(2015SCLD001)

Effects of Selenoprotein x Knockdown and Hydrogen Peroxide Challenge on Selenoprotein Genes Expression and Antioxidant Enzyme Activities in Human Normal Hepatocyte

  • TANG Jiayong ,
  • ZHAO Hua ,
  • JIA Gang ,
  • LIU Guangmang ,
  • CHEN Xiaoling ,
  • CAI Jingyi ,
  • WU Caimei
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  • Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, China

Received date: 2016-02-28

  Online published: 2016-08-17

摘要

本试验旨在探讨硒蛋白x(Selx)基因沉默和过氧化氢(H2O2)处理对人正常肝脏细胞(LO2)中硒蛋白基因表达和抗氧化酶活性的影响,为探索Selx与其他硒蛋白之间的调控关系及Selx的功能奠定基础。对本实验室保存的Selx基因稳定沉默LO2细胞株(siSelx)和对照细胞株(CK),利用噻唑兰(MTT)法比较其生长速率;siSelx和CK用200 μmol/L H2O2处理2.5 h后,采用荧光定量PCR技术考察25个硒蛋白基因的表达量;同时比较Selx基因沉默以及H2O2处理对LO2细胞株中谷胱甘肽过氧化物酶(GSH-Px)、超氧化物歧化酶(SOD)活性及总抗氧化能力(T-AOC)的影响。结果显示:1)Selx基因沉默显著降低了LO2细胞株的生长速率(P<0.05);2)Selx基因沉默显著上调了Gpx6Txnrd2Seli基因的表达量(P<0.05),显著下调了Dio2Dio3SelmSepw1基因的表达量(P<0.05);H2O2处理显著上调了Dio1基因的表达量(P<0.05),显著下调了Gpx2基因的表达量(P<0.05);Selx基因沉默后H2O2处理显著上调了Gpx6Txnrd2Dio1基因的表达量(P<0.05),显著下调了Txnrd3Dio2SelmSepw1Selv基因的表达量(P<0.05);3)Selx基因沉默显著提高了LO2细胞株GSH-Px活性(P<0.05),显著提高了SOD的活性(P<0.05);Selx基因沉默后H2O2处理显著提高了GSH-Px活性(P<0.05)。由此可见,Selx基因沉默降低了LO2细胞株的生长速率,Selx基因沉默后H2O2处理影响部分硒蛋白基因在细胞中的表达量,同时也影响了GSH-Px、SOD活性。

关键词: Selx基因; 沉默; H2O2; 表达

本文引用格式

汤加勇 , 赵华 , 贾刚 , 刘光芒 , 陈小玲 , 蔡景义 , 吴彩梅 . 硒蛋白x基因沉默和过氧化氢处理对人正常肝脏细胞中硒蛋白基因表达和抗氧化酶活性的影响[J]. 动物营养学报, 2016 , 28(8) : 2431 -2438 . DOI: 10.3969/j.issn.1006-267x.2016.08.014

Abstract

The objective of this experiment was to explore the effects of selenoprotein x (Selx) gene knockdown and hydrogen peroxide (H2O2) challenge on selenoprotein genes expression and antioxidant enzyme activities in human normal hepatocyte (LO2), for further study its roles and regulatory relationship with other selenoprotein. The LO2 cells with Selx silenced (siSelx) and the control cells (CK) kept in our lab were used to compare the cell growth rates by methylthiazolyldiphenyl-tetrazolium bromide (MTT) methods. The siSelx and CK were further exposed to H2O2 for 2.5 h, and the mRNA levels of selenoprotein genes and activities of GSH-Px, SOD and T-AOC in cells were detected. The results showed as follows:1) Selx knockdown significantly decreased cell growth rates in LO2 cells (P<0.05). 2) Selx knockdown significantly up-regulated Gpx6, Txnrd2, Seli genes expression and significantly down-regulated Dio2, Dio3, Selm and Sepw1 genes expression (P<0.05); H2O2 challenge significantly up-regulated Dio1 gene expression and down-regulated Gpx2 gene expression (P<0.05); Selx knockdown followed H2O2 challenge significantly up-regulated Gpx6, Txnrd2, Dio1 genes expression (P<0.05), and significantly down-regulated Txnrd3, Dio2, Selm, Sepw1 and Selv genes expression (P<0.05). 3) Selx knockdown significantly improved the activities of GSH-Px and SOD in LO2 cells (P<0.05). Selx knockdown followed H2O2 challenge further significantly improved GSH-Px activity (P<0.05). In conclusion, Selx knockdown decreases the growth rate of LO2 cells and affects the expression of other selenoprotein genes with H2O2 challenge, also affects the activities of GSH-Px and SOD.

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