X盒结合蛋白1调控胶质瘤细胞氧化应激的研究

Role of XBP1 in the Regulation of Sensitivity of Glioma to Oxidative Stress

背景与目的:活性氧(ROS)可以通过诱导细胞氧化应激而诱导凋亡,肿瘤包括胶质瘤在体内的生长均处于高活性氧的状态,提高胶质瘤细胞对活性氧-氧化应激的敏感性可抑制肿瘤在体内的生长侵袭,而且还可以增强一些诱导活性氧的化疗药物的疗效。本研究通过干预XBP1基因的表达,观察XBP1是否可调控胶质瘤细胞对氧化应激的敏感性,并探讨其机制。方法:用siRNA转染技术抑制U251MG细胞XBP1基因的表达后对比细胞对H2O2诱导的细胞死亡、线粒体膜电位下降、活性氧堆积、以及P38磷酸化,以此来反映细胞内氧化应激的水平;并比较细胞内抗氧化分子的表达情况;将XBP1基因过表达后观察能否起到相反的效果;最后应用启动子突变技术分析XBP1对Catalase表达的影响。结果:U251MG细胞XBP1表达抑制后可以显著提高H2O2诱导的细胞死亡、线粒体膜电位下降、活性氧堆积以及延长P38的磷酸化,并降低细胞内一些抗氧化分子包括过氧化氢酶表达,XBP1过表达可以增强Catalase表达并且降低ROS堆积;启动子序列突变分析结果显示XBP1增强Catalase表达的效应完全依赖于启动子序列的CCAAT框。结论:XBP1对氧化应激引起的细胞损伤具有保护作用;其可能机制至少是通过上调Catalase的表达实现。抑制XBP1的表达可能作为一种分子靶向,对胶质瘤治疗有积极意义。

BACKGROUND OBJECTIVE：High level of reactive oxygen species（ROS） can activate oxidative stress,which may lead to apoptosis.Cancer cells,including glioma cells,have to survive under conditions of high ROS level in vivo.Tumor invasion or development may be suppressed and the anti-cancer effects of some ROS-inducing agents may be augmented by increased sensitivity to oxidative stress in cancer cells.In this study,we investigated whether XBP1 has an antioxidant role in glioma cells and what is the mechanism of this role.METHOD：Glioma cell line U251 was used in this experiment.XBP1 expression level in glioma cells was knocked down by SiRNA transfection.After exposure to hydrogen peroxide（H2O2）,cell death,MMP loss,ROS level and P38 phosphorylation were examined to show the oxidative stress level and expression of a variety of antioxidant molecules was compared in those cells.Overexpression of XBP1 was performed to observe the converse effects.Promoter mutation analysis was taken to explore the mechanism of varied catalase expression level by XBP1.RESULT：After exposure to H2O2,more extensive loss of mitochondrial membrane potential（MMP） and subsequent cell death,ROS generation and prolonged P38 phosphorylation were observed in XBP1-KO cells.Lower expression of several antioxidant molecule s including catalase was also found.In addition,similar results were confirmed by XBP1 SiRNA transfection in glioma cells.Overexpression of XBP1 recovered catalase expression and diminished ROS generation after H2O2 exposure.Mutation analysis of the catalase promoter region demonstrated a pivotal role of CCAAT boxes for the XBP1-mediated enhancing effect.CONCLUSION：These results indicate a protective role of XBP1 against oxidative stress,which is at least partially due to XBP1＇s positive regulation on catalase expression.XBP1 may be good candidate as a molecular target for glioma treatment.