目的 探讨白头翁皂苷D体外抗肝癌作用及其相关机制。方法 采用MTT法考察白头翁皂苷D对人肝癌细胞BEL-7402增殖的影响,同时观察BEL-7402细胞形态的变化,采用集落形成实验考察白头翁皂苷D对BEL-7402细胞集落形成的影响;Hoechest 33342染色观察BEL-7402细胞凋亡的形态变化;流式细胞术检测BEL-7402细胞凋亡率和线粒体膜电位的变化;采用Western blotting法检测BEL-7402细胞Bcl-2和Caspase-3蛋白的表达。结果 MTT结果显示白头翁皂苷D对BEL-7402细胞增殖有抑制作用;集落形成实验结果显示白头翁皂苷D能明显抑制BEL-7402细胞集落的形成,白头翁皂苷D(25.00 μg/mL)能显著诱导BEL-7402细胞凋亡;白头翁皂苷D(18.75、25.00 μg/mL)能显著降低BEL-7402细胞线粒体膜电位;白头翁皂苷D(12.50、18.75、25.00 μg/mL)可显著上调BEL-7402细胞中Caspase-3的表达;白头翁皂苷D(25.00 μg/mL)可显著下调Bcl-2的表达。结论 白头翁皂苷D有良好的体外抗肝癌作用,其机制与调节线粒体途径凋亡相关蛋白Bcl-2、Caspase-3的表达有关。

Objective To research the antihepatocarcinoma effect in vitro of pulchinenoside D and mechanisms. Methods MTT assay was used to evaluate the proliferative inhibition of pulchinenoside D on hepatoma carcinoma cells (BEL-7402), and the morphology of BEL-7402 cells was also observed; Colony formation method was used to examine colony-forming role by pulchinenoside D on BEL-7402 cells; The morphological changes of drug-induced apoptosis of BEL-7402 were observed by Hoechst 33342 staining; Flow cytometry assay was used to analyze the apoptosis rate and mitochondrial membrane potential changes of BEL-7402 cells; Western blotting was employed to determine the expression levels of Bcl-2/Caspase-3 protein in BEL-7402 cells. Results MTT assay showed that pulchinenoside D could inhibit the proliferation of BEL-7402 cells, and colony formation method showed that pulchinenoside D could significantly inhibit the colony formation of BEL-7402 cells; Pulchinenoside D (25.00 μg/mL) could significantly induce the apoptosis of BEL-7402; Pulchinenoside D (18.75 and 25.00 μg/mL) could significantly reduce the mitochondrial membrane potential of BEL-7402 cells; Pulchinenoside D (12.50, 18.75, and 25.00 μg/mL) could significantly increase the expression of Caspase-3; Pulchinenoside D (25.00 μg/mL) could significantly down-regulate the expression of Bcl-2. Conclusion Pulchinenoside D has antitumor effects in vitro; The mechanism is involved in regulation of the mitochondrial pathway apoptosis-related protein expression of Bcl-2 and caspase-3.

国家自然科学基金资助项目(81273402);重大新药创制国家科技重大专项资助(2011ZX11102-001-19);江苏省自然科学基金资助(BK2012620);2013年苏州大学大学生创新创业训练计划(2013xj060)