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目的探讨白头翁皂苷D对人乳腺癌MCF-7细胞的体内外抗肿瘤作用及其机制。方法采用MTT法和吉姆萨染色,考察白头翁皂苷D对人MCF-7细胞体外增殖抑制的影响;建立MCF-7细胞裸鼠移植瘤模型,考察白头翁皂苷D体内抗肿瘤作用;HE染色和透射电镜实验考察肿瘤组织形态学和超微结构的变化;采用Western Blot法检测MCF-7细胞中Bcl-2、Caspase-3及PI3K/AKT/mTOR途径相关蛋白PI3K-p85、p-AKT、p-mTOR、p-p70S6K的表达。结果白头翁皂苷D可抑制MCF-7细胞的增殖,且呈剂量依赖关系;裸鼠体内实验结果显示白头翁皂苷D可抑制肿瘤的生长,30.0 mg·kg~(-1)剂量组瘤体质量明显降低,与模型组比较,差异有统计学意义(P<0.01);形态学观察结果也显示白头翁皂苷D对MCF-7细胞具有凋亡作用,且随着给药剂量的增加变化越明显;白头翁皂苷D(15.0,20.0,25.0μmol·L~(-1))不同剂量组均可抑制MCF-7细胞中Bcl-2、Caspase-3蛋白的表达,与对照组比较,差异均有统计学意义(P<0.01);白头翁皂苷D还可下调PI3K/AKT/mTOR信号传导通路相关蛋白PI3K-p85、p-AKT、p-mTOR、p-p70S6K蛋白的表达,20.0,25.0μmol·L~(-1)剂量组与对照组比较,差异均有统计学意义(P<0.05,P<0.01)。结论白头翁皂苷D对MCF-7细胞有显著的体内外抗肿瘤作用,其机制可能是通过下调PI3K/AKT/mTOR信号传导通路诱导细胞凋亡。
Objective To investigate the anti-tumor effect of Pulsatilla saponin D on human breast cancer MCF-7 cells in vitro and in vivo and its mechanism. Methods MTT assay and Giemsa staining were used to investigate the effect of Pulsatilla saponin D on the proliferation inhibition of human MCF-7 cells. The model of MCF-7 cells transplanted into nude mice was established. The anti-tumor effect of Pulsatilla saponin D was observed in vivo. HE staining and transmission electron microscopy The morphological and ultrastructural changes of the tumor tissues were examined. The expressions of Bcl-2, Caspase-3 and PI3K-p85, p-AKT and p-mTOR in MCF-7 cells were detected by Western Blot , P-p70S6K expression. Results Pulsatilla saponin D could inhibit the proliferation of MCF-7 cells in a dose-dependent manner. The results of in vivo experiments in vivo showed that Pulsatilla saponin D inhibited tumor growth, and the tumor mass of 30.0 mg · kg -1 group was significantly decreased (P <0.01). The results of morphological observation also showed that Pulsatilla saponin D had the apoptosis effect on MCF-7 cells, and the more obvious the change was with the increase of the dosage. Pulsanion saponins D (15.0,20.0,25.0μmol·L -1) could inhibit the expression of Bcl-2 and Caspase-3 protein in MCF-7 cells in different dose groups compared with the control group, the difference was statistically significant ( P <0.01). Pulsatilla saponin also down-regulated the expression of PI3K-p85, p-AKT, p-mTOR and p-p70S6K proteins in PI3K / AKT / mTOR signaling pathway, ) Dose group compared with the control group, the difference was statistically significant (P <0.05, P <0.01). Conclusion Pulsatilla saponin D has a significant antitumor activity against MCF-7 cells in vitro and in vivo. The possible mechanism is that PI3K / AKT / mTOR signaling pathway may induce apoptosis.