过氧化氢诱导人子宫内膜癌Ishikawa细胞氧化应激模型的建立

Establishment of hydrogen peroxide -induced oxidative stress model in Ishikawa cells

目的探讨不同浓度过氧化氢（H2O2）处理不同时间对子宫内膜癌Ishikawa细胞氧化应激水平的影响，确定构建体外子宫内膜癌氧化应激模型的条件。方法不同浓度（50、100、200、400μmol／L）H2O2处理Ishika-wa细胞不同时间（2、4、8、24h）后，CCK-8法测定细胞存活率；通过流式细胞术分析检测细胞内活性氧自由基（reactive oxygen species, ROS）的水平；紫外分光光度法检测细胞培养上清液中乳酸脱氢酶（ lactic dehydrogenase,LDH）的含量及超氧化物歧化酶（ superoxide dismutase, SOD）活性。结果随着H202浓度的增加及处理时间的延长，Ishikawa细胞存活率逐渐下降，细胞内ROS水平不断增高。其中，100μmol／LH2O2处理Ishikawa细胞4h后，细胞存活率为87．36％，细胞内ROS水平较对照组显著增加（P〈0．05），SOD活性明显低于对照组（P〈0．05），但LDH含量无明显增多（P〉0．05）。结论100μmol／LH2O2处理4h是建立人子宫内膜癌Ishikawa细胞氧化应激模型的最佳条件。

Objective To discuss the effects of various concentrations of hydrogen peroxide （ H2 O2 ） on the oxidative stress of Ishikawa cells, and determine the conditions for establishment of an oxidative stress model of endometrial cancer in vitro. Methods Ishikawa cells were treated with different concentrations （50, 100, 200 and 400 μmol/L） of H2O2 for various durations of time （2, 4, 8, and 24 h）. Cell viability was determined by CCK - 8 assay. The level of intracel- lular reactive oxygen species （ROS） was detected by flow cytometry. The content of lactic dehydrogenase （LDH） and the activity of superoxide dismutase （SOD） were measured by ultraviolet spectrophotometry. Results With the increase of H2 O2 concentrations and treatment time, the viability of Ishikawa cells was decreased but the level of intracellular ROS was increased. Meanwhile, LDH content within the supernatant of cell medium was increased but SOD activity was weak- ened. After exposure to 100 μmol/L of H2O2 for 4 h, the cell survival rate reached to 87.36% , while significant increa- ses in intracellular ROS level （ P 〈 0.05 ） and remarkable decreases in SOD activity were found （ P 〈 0.05 ） compared with the control. However, no significant changes were seen in LDH content （ P 〉 0.05 ）. Conclusions The optimal condition for establishment of an oxidative stress model of Ishikawa cells is treatment with 100 μmol/L of H2O2 for 4 h.