野艾蒿挥发油诱导HeLa细胞凋亡与坏死

Essential Oil from Artemisia lavandulaefolia Induces Apoptosis and Necrosis of HeLa Cells

目的:研究野艾蒿挥发油对HeLa细胞凋亡与坏死的影响。方法:MTT法检测细胞活力,显微镜观察细胞形态与结构的变化,DNA Ladder、流式细胞仪检测细胞凋亡,彗星电泳检测DNA损伤,Western blot检测Caspase-3和多聚ADP核糖聚合酶(PARP)表达。结果:MTT法检测结果显示野艾蒿挥发油对HeLa细胞的增殖有明显的抑制作用,呈剂量依赖性。野艾蒿挥发油处理HeLa细胞24 h后,100、200μg/mL实验组细胞体积缩小,核固缩,染色质凝集、片段化,表现为典型的凋亡特征;400μg/mL实验组细胞膜破裂、内含物外泄,表明细胞已坏死。此外,DNA Ladder结果显示,各实验组均有梯状条带;流式细胞仪检测结果也显示,各实验组均有凋亡峰。野艾蒿挥发油处理HeLa细胞6 h后,100μg/mL实验组彗星电泳中出现彗尾。Western-blotting结果显示,野艾蒿挥发油能够诱导HeLa细胞中Caspase-3蛋白表达量降低,同时PARP发生剪切、失活。结论:野艾蒿挥发油具有抑制HeLa细胞增殖的作用。低浓度野艾蒿挥发油诱导细胞凋亡,高浓度野艾蒿挥发油引起细胞坏死,其凋亡机制是依赖Caspase-3介导的PARP凋亡信号转导途径。

Objective： To investigate the effects of Artemisia lavandulaefolia essential oil on apoptosis and necrosis of HeLa cells. Methods： Cell viability was assayed using MTT method. The morphological and structure alterations in HeLa cells were observed by microscopy. Furthermore,cell apoptosis was measured by DNA Ladder and flow cytometry. DNA damage was measured by comet assay, and the protein expression was examined by Western blot analysis. Results： MTT assay displayed essential oil from Artemisia lavandulaefolia could inhibit the proliferation of HeLa cells in a dose-dependent manner. After treated with essential oil of Artemisia lavadulaefolia for 24 h,HeLa cells in 100 and 200 μg / mL experiment groups exhibited the typical morphology changes of undergoing apoptosis, such as cell shrinkage and nucleus chromatin condensed. However,the cells in the 400 μg / mL group showed the necrotic morphology changes including cytomembrane rupture and cytoplasm spillover. In addition,DNA Ladder could be demonstrated by DNA electrophoresis in each experiment group. Apoptosis peak was also evident in flow cytometry in each experiment group. After treating the HeLa cells with essential oil of Artemisia lavadulaefolia for 6 h,comet tail was detected by comet assay. Moreover,western blotting analysis showed that caspase-3 was activated and the cleavage of PARP was inactivated. Conclusion： Essential oil from Artemisia lavadulaefolia can inhibit the proliferation of HeLa cells in vitro. Low concentration of essential oil from Artemisia lavadulaefolia can induce apoptosis, whereas high concentration of the compounds result in necrosis of HeLa cells. And,the mechanism may be related to the caspase-3-mediated-PARP apoptotic signal pathway.