苦参碱诱导人乳腺癌Bcap-37细胞自噬及凋亡作用研究  

Antitumor Effect of Matrine in Human Breast Cancer Bcap-37 Cells by Apoptosis and Autophagy

作　　者：任莉莉[1] 蓝天[2] 王晓稼[1] 

机构地区：[1]浙江省肿瘤医院,浙江杭州310022 [2]杭州市中医院,浙江杭州310006

出　　处：《中华中医药学刊》2014年第11期2756-2759,I0013,共5页Chinese Archives of Traditional Chinese Medicine

基　　金：浙江省医药卫生科技计划项目(2013KYB04)

摘　　要：目的:研究苦参碱诱导人乳腺癌Bcap-37细胞自噬及凋亡的作用,为苦参碱用于乳腺癌治疗提供理论依据。方法:苦参碱(终浓度为0、0.5、1、1.5、2、2.5、3 mg/mL)处理乳腺癌Bcap-37细胞24、48、72 h,采用MTT法检测细胞增殖抑制率;Annexin V-FITC和碘化丙啶(PI)双染、流式细胞仪检测细胞凋亡率;用倒置显微镜和透射电镜观察细胞形态变化;运用Western-blot方法检测Cleaved PARP和Lc3b-Ⅰ/Lc3b-Ⅱ蛋白表达水平。苦参碱联合氯喹(chloroquine)(自噬抑制剂)处理Bcap-37细胞24 h后,运用MTT法检测细胞增殖抑制率的变化。实验数据采用SPSS 17.0软件进行统计,计量资料用均数±标准差(x珋±s)表示,多组均数间比较用方差分析,两组均数比较用T检验,均以P<0.01为差异显著的检验标准。结果:苦参碱可显著抑制乳腺癌Bcap-37细胞增殖,作用呈浓度-时间依赖性;苦参碱诱导细胞凋亡,且凋亡率与苦参碱浓度成正比;经苦参碱处理后,Bcap-37细胞胞浆内出现大量囊性小泡,经透射电镜扫描证实细胞发生自噬;Western-blot法检测到Cleaved PARP和Lc3b-Ⅱ蛋白表达均上调。苦参碱联合氯喹作用于Bcap-37细胞后MTT法检测细胞增殖抑制作用明显加强(P<0.01)。结论:苦参碱可显著抑制Bcap-37细胞增殖;促进细胞凋亡;诱导细胞发生自噬现象;自噬抑制剂氯喹可增强苦参碱抑制细胞增殖的作用,提示自噬现象是一种细胞保护性反应。Objective ： To study the antitumor effect of matrine on human breast cancer Bcap - 37 cells by inducing apoptosis and autophagy and to provide the basis for matrine to treat breast cancer. Methods ： Human breast cancer Bcap - 37 cells were cultured in vitro and treated by different final concentrations （0, 0. 5,1,1.5,2,2.5,3 mg/mL） of matrine, and then cultured for 24 h ,48 h and 72 h. MTT assay was used to detect viability of Bcap - 37 cells treated by different concentrations of matrine. AnnexinV - FITC/PI double staining was used to detect cell apoptosis rate. Inverted phase contrast microscope was used to observe cellular morphological changes, and transmission electron microscope was performed to further examine ultrastructural structure of the cells treated with matrine. Western - blot assay was used to detect the protein expression levels of LC3 - I and LC3 - Ⅱ. MTT assay was used to detect proliferation rate of Bcap - 37 cells treated by matrine （ 1 mg/mL） combined with chloroquine （ autophagy inhibitor ） . SPSS 17.0 software was used for statistical analysis and the data was expressed as x ± s. T - test and two - way ANOVA were used for statistical analysis. Results ： Matrine significantly decreased cell viability in Bcap - 37 cells in a dose - and time - dependent manner and induced apoptosis of Bcap -37 cells in a dose dependent manner. Abundant cytoplasmic vacuoles were observed in Bcap - 37 cells treated with matrine, and they were finally confirmed to be autophagie vacuoles demonstrated by electron microscopy. Meantime, Western - blot revealed that the expression levels of LC3 - Ⅱ protein were higher. When the specific autophagie inhibitor chloroqnine was applied, MTT method detected that the cell proliferation inhibition improved remark-ably（ P 〈 0. 01 ）. Conclusions：Matrine inhibited the proliferation of Bcap -37 cells in vitro and induced apoptosis and autophagy in Bcap - 37 cells. Conversely,inhibition of Matrine - induced autophagy by treating the cancer cells wit

关 键 词：苦参碱 乳腺肿瘤 凋亡 自噬 

分 类 号：R285.5[医药卫生—中药学]