Nrf2基因过表达对百草枯诱导的骨髓间充质干细胞损伤的影响  

Effect of Nrf2 overexpression damaged bone marrow mesenchymal stem cells induced by Paraquat

作　　者：蔡晓霞[1] 陈燕珍[1] 周挺[1] 赵光举[1] 邱俏檬[1] 洪广亮[1] 卢阳[1] 卢中秋[1] 

机构地区：[1]温州医科大学附属第一医院急诊医学中心,浙江省温州325000

出　　处：《中华急诊医学杂志》2015年第8期851-856,共6页Chinese Journal of Emergency Medicine

基　　金：浙江省医药卫生科技计划项目（2012-ZD-A033）;浙江省“十二五”重点学科建设计划资助项目;浙江省医学创新学科计划项目（11-CX26）;浙江省中医药重点学科计划项目（2012-XK-A28）

摘　　要：目的观察核因子E2相关因子2转录因子（nuclear factor erythroid 2-related factor 2，Nri2）基因过表达对百草枯（paraquat，PQ）诱导的骨髓间充质干细胞（bone marrow mesenchymal stem ceils，BMSCs）损伤的影响。方法BMSCs随机（随机数字法）分为6组：正常BMSCs组、BMSCs-mCherry（红色荧光蛋白）组、BMSCs-Nrf2组、BMSCs＋PQ组、BMSCs．mCherry＋PQ组、BMSCs-Nrf2＋PQ组。正常BMSCs组、BMSCs—mCherry组、BMSCs-Nrf2组：正常培养基培养；BMSCs＋PQ组、BMSCs-mCherry＋PQ组、BMSCs—Nrf2＋PQ组：终浓度1．0mmol／L的PQ培养。24h之后收集细胞及上清液，蛋白免疫印迹法（westernblot）检测胞核内Nrf2蛋白量，CCK-8法检测细胞存活率，流式细胞仪检测细胞凋亡情况，化学比色法检测上清液脂质过氧化产物丙二醛（MDA）的含量及过氧化氢酶（CAT）的活性，酶联免疫吸附试验（ELISA）检测上清液白细胞介素_6（IL-6）、IL-10及肿瘤坏死因子-α（TNF-α）的含量。数据通过SPSS20．0进行统计学分析，组间比较采用单因素方差分析。结果BMSCs—mCherry＋PQ组与BMSCs＋PQ组比较，各测定数据差异无统计学意义（P〉0．05）。与BMSCs组比较，BMSCs＋PQ组细胞核Nrf2蛋白表达量明显上升（P=0．008）；与BMSCs＋PQ组比较，BMSCs—Nrf2＋PQ组Nrf2蛋白表达量升高（P=0．031）。与BMSCs组比较，BMSCs＋PQ组细胞存活率明显下降，细胞凋亡率明显升高（P=0．000）；与BMSCs＋PQ组比较，BMSCs-Nri2＋PQ组细胞存活率明显升高[（53．27±2．40）％掷．（75．20±2．92）％，P=0．000]，凋亡数量显著降低[（26．43±2．21）％郴．（16．30±1．73）％，P=0．000]。与BMSCs组比较，BMSCs＋PQ组MDA、IL-6、TNF．0L水平明显上升，CAT、IL-10水平明显降低（P=0．000）；与BMSCs＋PQ组比较，BMSCs—Nrf2＋PQ组MDA含量明显降低[（43．88±0．89）nmol／L138．（37．29±1．88）nmol／L，P=0．000]，上清液CAObjective To observe the effects of nuclear factor erythroid 2-related factor 2 （Nrf2） overexpression on damaged bone marrow mesenchymal stem cells （BMSCs） induced by paraquat （PQ）. Methods BMSCs were randomly （random number） divided into six groups： normal BMSCs group, BMSCs-mCherry （Red flurescent protein） group, BMSCs-Nrf2 group, BMSCs ± PQ group, BMSCs- mCherry ＋ PQ group and BMSCs-Nrf2 ＋PQ group. The BMSCs group, BMSCs-mCherry group and BMSCs- Nrf2 group were cultured by medium. The BMSCs ＋ PQ group, BMSCs-mCherry ＋ PQ group and BMSCs- Nrf2 ＋ PQ group were cultured by 1.0 mmol/L final concentration PQ. All cells and supernatant were collected at 24 h after culture in medium or in PQ. The nucleus protein level of Nrf2 was measured by Western blot; the cell viability was measured by CCK-8 assay; apoptosis of cells was detected by flow cytometry; the levels of malondialdehyde （MDA）, catalase （CAT） in supernatant were determined by chemical colorimetry; the levels of interleukin （IL） -6, IL-10 and tumor necrosis factor （TNF） -α in supernatants were detected by Enzyme-linked immunosorbent assays （ELISA） . One-way analysis of variance （AVOVA） was employed for statistical analysis by using SPSS version 20. 0 to compare values among all groups. Results There were no significant differences in all biomarker variables between BMSCs ＋ PQ group and BMSCs-mCherry ＋ PQ group （ P 〉 0. 05 ）. Compared with BMSCs group, the nucleus protein level of Nrl2 in BMSCs ＋ PQ group was higher markedly （P = 0. 008） ; compared with BMSCs ＋ PQ group, the nucleus protein level of Nrf2 in BMSCs-Nrf2 ＋ PQ group was also higher （ P = 0. 031 ）. The cell viability was much lower in BMSCs ＋ PQ group than that in BMSCs group, while the cell apoptosis was much higher in BMSCs ± PQ group （ P = 0. 000 ） ; compared with BMSCs ± PQ group, the cell viability in BMSCs-Nrf2 ＋ PQ group increased remarkably [ （75.20 ± 2. 92） % vs. （ 53.27 ± 2.40） % （ P

关 键 词：NRF2 百草枯 骨髓间充质干细胞 氧化应激 

分 类 号：R595.4[医药卫生—内科学]