谷胱甘肽S转移酶mu5对肿瘤坏死因子α诱导人支气管上皮细胞氧化损伤的调节作用研究  

The Regulation of GSTM5 in Oxidative Stress Induced by TNF-α in Human Bronchus Epithelial Cells

作　　者：赵艳雪[1,2] 李理[1] 袁伟锋[1] 郭振辉[3] 黄文杰[1] 

机构地区：[1]广州军区广州总医院呼吸内科,广东广州510010 [2]广州医科大学研究生学院,广东广州510182 [3]广州军区广州总医院MICU科,广东广州510010

出　　处：《中国呼吸与危重监护杂志》2016年第3期284-288,共5页Chinese Journal of Respiratory and Critical Care Medicine

基　　金：国家自然科学基金面上项目(编号:81370173);国家自然科学基金青年科学基金(编号:81200002);广东省老年感染与器官功能支持重点实验室;广州市老年感染与脏器功能支持重点实验室资金支持

摘　　要：目的建立肿瘤坏死因子α(TNF-α)诱导支气管上皮(16HBE)细胞炎症模型,探讨谷胱甘肽S转移酶mu5(GSTM5)在炎症诱导的氧化应激损伤中的作用。方法以TNF-α(10 ng/m L)刺激16HBE细胞,运用真核细胞表达载体技术,增强GSTM5基因表达,比色法检测细胞内丙二醛(MDA)及总抗氧化能力(T-AOC);MTT法检测细胞存活率;RT-PCR法检测NADPH氧化酶(NOX)家族NOX1、NOX2、NOX3、NOX4、NOX5、双重氧化酶1(DUOX1)、DUOX2相关基因转录水平;蛋白质印迹法检测NOX1和NOX2蛋白表达水平。结果 TNF-α刺激16HBE细胞后,细胞内MDA水平增高,T-AOC能力减低,细胞存活率明显降低。预转染GSTM5质粒可有效降低TNF-α诱导的细胞内MDA的增高(P<0.05),显著增强T-AOC能力(P<0.05),同时增加16HBE细胞存活率(P<0.05)。GSTM5质粒组NOX1、NOX2基因转录强度以及蛋白表达水平与TNF-α组及阴性对照组相比显著降低(P均<0.05),而NOX3、NOX4、NOX5、DUOX1、DUOX2基因表达强度比较,差异无统计学意义(P均>0.05)。结论增强GSTM5表达对炎症诱发的人支气管上皮细胞氧化应激损伤有保护性调节作用,其机制与下调NOX1、NOX2基因表达密切相关。Objective To establish a cell inflammation model induced by tumor necrosis factor-α（ TNF-α） in human bronchus epithelial cells,and investigate the effects of glutathione S-transferase mu 5（ GSTM5） on the inflammation and oxidative stress. Methods 16 HBE cells were treated with TNF-α（ 10 ng/m L,24 h） in the absence or presence of the constructed GSTM5 eukaryotic expression vector（ 1 μg/m L）. The concentration of malondialdehyde（ MDA） and total antioxidation capacity（ T-AOC） were detected by colorimetric method. The survival rate of cells was assessed by the methyl thiazolyl tetrazolium（ MTT） assay. The transcription level of NADPH oxidase-1（ NOX1）,NOX2,NOX3,NOX4,NOX5,dual oxidase-1（ DUOX1） and DUOX2 were evaluated by RT-PCR. Western blot was performed to investigate the protein levels of NOX1 and NOX2. Results TNF-α simulation significantly increased the level of MDA in cells,and decreased the level of T-AOC and survival rate of 16 HBE. When transfected with the GSTM5 eukaryotic expression vector,the concentration of MDA significantly decreased（ P〈 0. 05）,and the activation of T-AOC increased dramatically（ P〈 0. 05）. Consequently,the survival rate of 16 HBE in the GSTM5 group improved（ P〈 0. 05）. The 16 HBE cells transfected with the constructed GSTM5 eukaryotic expression vector had a lower transcription and protein levels of NOX1 and NOX2（ all P〈 0. 01）. There were no significant changes in the mRNA expressions of NOX3,NOX4,NOX5,DUOX1 or DUOX2.Conclusion GSTM5 may down-regulate the transcription level of NOX1 and NOX2 to reduce the inflammation and oxidative stress induced by TNF-α.

关 键 词：谷胱甘肽S转移酶mu 5 真核细胞表达载体 氧化应激 炎症 

分 类 号：R563.8[医药卫生—呼吸系统]