钙结合蛋白S100A8在γ射线损伤小鼠巨噬细胞RAW264.7中的作用

为探索钙结合蛋白S100A8在γ射线对造血免疫系统损伤中的作用,构建S100A8真核表达载体,转染小鼠巨噬细胞RAW264.7,研究S100A8在γ射线诱导RAW264.7细胞周期紊乱和胞内活性氧改变中的作用。运用RT-PCR和细胞荧光免疫方法鉴定增强表达S100A8的RAW264.7稳定细胞克隆。流式技术检测10Gyγ射线照射6h后细胞周期及H2O2刺激后胞内活性氧的变化。结果显示,与转染空质粒的对照细胞相比,增强表达S100A8细胞克隆G0/G1期细胞比例增高,S期细胞减少,不发生G2/M阻滞;并且H2O2作用后细胞中活性氧水平低于转染空白质粒的细胞。因此,增强表达S100A8的RAW264.7细胞与转染空白质粒细胞相比对辐射更具抗性,其可能原因是与增强表达的S100A8分子具有抗氧化功能有关。

Effect of calcium-binding protein S100A8 in macrophage cell line RAW264.7 of mouse injured by irradiation

In order to establish the stable clones of RAW264.7 cells with S100A8 gene overexpression, and directly investigate the role of S100A8 in γ-rays induced hematoiesis/immune system injury, RT-PCR method and im-muno-fluorescence staining analysis were performed to measure the expression of S100A8 mRNA and protein.The cell cycle was detected by flow cytometry at 6 h after irradiation with 10 Gy γ-rays.The level of reactive oxygen intermediates （ROIs） was detected by flow cytometry at 6h after H2O2 treatment.The cell cycle analysis showed that the percentages of RAW264.7/S100A8 cells had an increase in G0/G1 phase and a decrease in S phase, and G2/M arrest didn＇t occur in experimental group while the level of ROIs was lower than that of the negative control cells.These results indicate that radiation resistant of RAW264.7/S100A8 cells may be related to the antioxidant role of over-expressed S100A8 in comparison to RAW264.7/pcDNA3.1＋ cells.