叠氮甲基蒽醌衍生物诱导KB细胞及其耐药株细胞的凋亡

Apoptosis of human carcinoma of mouth floor KB cells and multidrug resistant KBv200 cells induced by azide methyl anthraquinone derivative

目的 研究大黄素的蒽醌衍生物 6 叠氮甲基 1 羟基 3, 8 二甲氧基 9, 10 蒽醌 (AMAD 10)抑制KB细胞及其耐药株KBv200的生长和诱导细胞凋亡的作用。方法 细胞毒用MTT法测定;细胞内活性氧(ROS)和线粒体跨膜电位(ΔΨm)分别用DCFH DA和DiOC6荧光探针标记,流式细胞仪检测;AnnexinV染色和DNA琼脂糖凝胶电泳检测AMAD诱导凋亡作用。结果 AMAD浓度依赖性抑制KB和KBv200细胞的生长和引起活性氧明显增加,并能引起两种细胞ΔΨm时间依赖性的降低,AMAD可浓度依赖性地诱导两种细胞凋亡。结论 AMAD体外显著抑制KB及其耐药株KBv200的生长,这可能与其引起细胞内ROS增加、使ΔΨm降低,从而诱导细胞凋亡有关。

Aim To determine the effects of azide methyl anthraquinone derivative (AMAD) on growth inhibition and inducing apoptosis of multidrug resistant (MDR) KBv200 cells and parental drug-sensitive KB cells. Methods Cytotoxicity was determined by tetrazolium (MTT) assay. Reactive oxygen species (ROS) levels and mitochondrial membrane potential (ΔΨ_m) in cells were labeled with DCFH-DA and DiOC_6 and tested by flow cytometry. Annexin V stain and DNA ladder were used to examine the apoptosis of KB and KBv200 cells induced by AMAD. Results AMAD was shown to inhibit the growth of KB and KBv200 cells significantly in a concentration-dependent manner, with mean IC_50 of 0.36 and 0.45 μmol·L-1, respectively. The generation of ROS increased obviously after the cells were treated with AMAD for 12 h, up to the peak in 24 h, meanwhile the levels of ΔΨ_m were time-dependently decreased. DNA fragmentation appeared on the agarose gel. Annexin V stain showed AMAD induced apoptosis of KB and KBv200 cells also in a concentration-dependent manner. Conclusion AMAD showed inhibitory effect on both MDR KBv200 cells and parental drug-sensitive KB cells. The mechanism of action was associated with the increase of the cellular ROS level and the decrease of the mitochondrial membrane potential induced by AMAD, which result in cell apoptosis.