活性氧在δ-榄香烯诱导人结肠腺癌DLD-1细胞凋亡中的作用研究

目的探讨δ-榄香烯诱导人结肠腺癌DLD-1细胞凋亡的机制以及活性氧（reactive oxygen species，ROS）在δ-榄香烯诱导人结肠腺癌细胞DLD-1凋亡中的作用。方法采用MIT法考察了δ-榄香烯对人结肠腺癌DLD-1细胞和人正常肝胚胎WRL-68细胞增殖的影响；采用氯甲基二氯荧光素二乙酸酯（CM—H2DCFDA）定量检测凋亡过程中ROS的变化；流式细胞术JC-1标记检测凋亡细胞中线粒体膜电位（△φm）的影响。结果δ-榄香烯明显抑制DLD-1细胞的增殖，对WRL-68细胞的细胞毒作用不明显。200μM的δ-榄香烯作用DLD-1细胞1小时、3小时和6小时所引起的ROS含量的增加，分别接近空白对照组的7．8倍、18．2倍和21．8倍；这种作用可以被谷胱甘肽（glutathione，GSH）（ROS的清除剂）显著性抑制，即加入GSH后δ-榄香烯对ROS生成率（3．46％）与空白对照（2．11％）相比无显著性差异。流式细胞仪检测结果表明，线粒体膜电位（△φm）的降低呈时间依赖性，6小时、12小时、24小时线粒体膜电位降低量分别是空白组的（2．24±0．12）倍，（2．85±0．33）倍，（5．11±0．24）倍。结论△φm的降低表明δ-榄香烯的抗肿瘤作用是以诱导肿瘤细胞凋亡的方式产生的，且其凋亡过程提示可能与ROS的生成增加相关。

The role of ROS in apoptosis induced by δ-elemene in colorectal adenoarcinoma cells

Objective To investigate the apoptosis effect and the role of ROS in the mechanism of δ-elemene on Colorectal Adenoarcinoma Cells DLD-1 in vitro. Methods Cell proliferation was measured by MTT method. Apoptosis was further confirmed and quantified by the mitoehondrial probe JC-1 using flow cytometry. The rapid increase in intracellular reactive oxygen species （ROS） levels was involved in the mechanism of cell death. Results δ-elemene induced DLD-1 cell apoptosis and a weaker cytotoxic effect on WRL-68 cell. There was approximately 7. 8, 18.2 or 21.8 fold increase in generation of ROS in 200 μM δ-elemene for l h, 3h or 6h compared with vector control cells, the increase was statistically significanL Furthermore, The change of ROS induced by δ-elemene was inhibited by GSH to a level （3.46%） that was almost as low as the untreated control （2. 11% ）. the trends of increasing incidences of compromised △φm for 6h, 12h or 24h were （2. 24±0. 12）, （2. 85±0. 33 ）, （5.11±0. 24） fold with respected to the untreated control. Conclusions These results indicate that apoptosis pathway in DLD-1 cells by δ--elemene was associated with the ROS generation.