丹栀逍遥散含药血清对乳腺癌MCF-7细胞自噬的影响

Effect of Containing Serum of Danzhi Xiaoyao Powder on Autophagy of Human Breast Cancer Cell Line MCF-7

目的:观察丹栀逍遥散含药血清通过诱导自噬对乳腺癌细胞MCF-7的影响。方法:大鼠ig 8.99 g·kg-1丹栀逍遥散,制备含药血清,取对数生长期MCF-7细胞,以1×105个/m L密度接种于96孔板,每孔100μL,分为空白血清组和含药血清组,每组设3个复孔,分别作用12,24,48,72 h,噻唑蓝(MTT)法测定其对MCF-7细胞生长的抑制作用,单丹磺酰戊二胺(MDC)荧光染色观察细胞自噬泡的形成情况,免疫印迹法(Western blotting)检测自噬蛋白(Beclin1)的表达。结果:与空白血清组比较,15%丹栀逍遥散含药血清具有显著抑制MCF-7细胞生长的作用,确定其最佳作用时间为48 h。丹栀逍遥散含药血清作用细胞48 h后能够增强MDC染色荧光强度,增加自噬泡的形成并显著上调Beclin1的表达。结论:丹栀逍遥散含药血清能够显著抑制MCF-7的细胞生长,诱导细胞自噬性死亡可能是其抗乳腺癌的机制之一。

Objective： To investigate the effects of the containing serum of Danzhi Xiaoyao powder on human breast cancer cell line MCF-7 by regulation of autophagy. Method： Rats were intragastrically given with8. 99 g·kg- 1Danzhi Xiaoyao powder to prepare the containing serum. The exponential phase MCF-7 cells were treated with containing serum of Danzhi Xiaoyao powder was inoculated on 96-well plates with the density of 1 × 105 cells / m L,100 μL per well. The cells were divided into blank serum group and containing serum group,with 3double-wells in each group. They were treated for 12,24,48,72 h respectively,and then methyl thiazolyl tetrazolium（ MTT） assay was used to measure its inhibitory effects on the growth of MCF-7 cells; the formation of autophagic vacuoles was detected using monodansylcadaverine（ MDC） fluorescence microscope,and the expression of Bclin1 protein was detected using Western blotting. Result： Compared with the blank serum group,the growth of MCF-7 cells was significantly inhibited by 15% Danzhi Xiaoyao powder＇s containing serum,and the optimal condition was achieved after treatment for 48 h. At this time point,MDC staining fluorescence intensity and the formation of autophagic vacuoles were increased; at the same time,the expression of Beclin1 in MCF-7 cells was significantly up-regulated. Conclusion： Containing serum of Danzhi Xiaoyao powder could significantly inhibit MCF-7 breast cancer cells proliferation and induce autophagic death of cells. This may be one of the important mechanisms of its anti-breast cancer.