[1]周静,李国兵,郑怡,等.莲心碱对乳腺癌细胞MDA-MB-231自噬功能的影响[J].第三军医大学学报,2015,37(07):629-633.
 Zhou Jing,Li Guobing,Zheng Yi,et al.Liensinine inhibits autophagy in breast cancer MDA-MB-231 cells[J].J Third Mil Med Univ,2015,37(07):629-633.
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
37卷
期数:
2015年第07期
页码:
629-633
栏目:
论著
出版日期:
2015-04-15

文章信息/Info

Title:
Liensinine inhibits autophagy in breast cancer MDA-MB-231 cells
作者:
周静李国兵郑怡银涛高宁
第三军医大学药学院生药学教研室;广州军区疾病预防控制中心
Author(s):
Zhou Jing Li Guobing Zheng Yi Yin Tao Gao Ning

Department of Pharmacognosy, College of Pharmacy, Third Military Medical University, Chongqing, 400038; Center for Disease Control and Prevention of Guangzhou Military Command, Guangzhou, Guangdong Province, 510507, China

关键词:
乳腺癌MDA-MB-231莲心碱自噬
Keywords:
breast cancer MDA-MB-231 cells liensinine autophagy
分类号:
R737.9;R966;R979.19
文献标志码:
A
摘要:

目的      观察莲心碱作用于乳腺癌细胞MDA-MB-231后,自噬表型的变化,初步论证莲心碱对乳腺癌细胞MDA-MB-231自噬功能的调控作用。      方法      透射电子显微镜观察莲心碱处理后MDA-MB-231的亚细胞结构变化,激光共聚焦显微镜观察pEGFP-LC3或ptfLC3(mRFP-EGFP-LC3)转染后自噬小体的形成和自噬流的变化,Western blot 检测自噬相关蛋白LC3B,SQSTM1/p62,Beclin1的表达,以及论证自噬流的变化。      结果      透射电镜显示,与对照组比较,莲心碱处理后MDA-MB-231细胞内有较多自噬空泡出现,部分分裂的线粒体,但未见到高密度的嗜锇性的自噬溶酶体结构;EGFP-LC3转染后,莲心碱组自噬体数量显著多于对照组(P<0.01);Western blot检测到自噬相关蛋白LC3B-Ⅱ,SQSTM1/p62表达呈现量效性和时效性的增多[20 μmol/L组或24 h组与对照组比较(P<0.01)],Beclin1未见显著变化[20 μmol/L组与对照组比较(P>0.05)];mRFP-EGFP-LC3转染后,对照组可见少量自噬体和自噬溶酶体,莲心碱和Bafilomycin A1处理组均可见大量自噬体结构,未见显著自噬溶酶体结构;Western blot证实莲心碱处理后增多的LC3B-Ⅱ不受Bafilomycin A1预处理的影响[莲心碱组LC3B-Ⅱ,SQSTM1/p62蛋白水平与Bafilomycin A1预处理加莲心碱组比较(P>0.05)]。      结论      莲心碱抑制自噬小体成熟,阻断乳腺癌细胞MDA-MB-231自噬降解作用。

Abstract:

Objective        To determine the effect of liensinine on the autophagic phenotypes in breast cancer MDA-MB-231 cells in order to investigate its regulative role on autophagic functions in the cells.       Methods       Transmission electron microscopy (TEM) was used to observe the ultrastructural changes in MDA-MB-231 cells after liensinine treatment (20 μmol/L for 24 h). Confocal laser scanning microscopy was employed to determine the formation of autophagosome and autophagic flux after the cells were transfected with pEGFP-LC3 or ptfLC3. Western blot analysis was used to detect the expression of autophagy related proteins, including LC3B, SQSTM1/p62 and Beclin1.       Results       TEM displayed there were more autophagic vacuoles and some fission mitochondria, but no high density osmiophilic autolysosomes in the MDA-MB-231 cells after liensinine treatment. Confocal laser scanning microscopy confirmed that there were more EGFP-LC3 puncta in the cells after pEGFP-LC3 transfection (P<0.01). Western blotting indicated that the expression levels of LC3B-Ⅱ and SQSTM1/p62 were increased in a dose- and time-dependent manner (20 μmol/L or 24 h vs control, P<0.01), and those of LC3B-Ⅰ and Beclin1 had no obvious change (20 μmol/L vs control, P>0.05) in the cells after pEGFP-LC3 transfection. While, in the cells after mRFP-EGFP-LC3, there were only a few of autophagosomes and autolysosomes in the control cells, but a lot of autophagosomes in the cells treated with liensinine and bafilomycin A1, but no autolysosomes. Western blotting indicated that bafilomycin A1 had no effect on liensinine induced enhancement of LC3B-Ⅱ and SQSTM1/p62 (P>0.05).       Conclusion        Liensinine suppresses the maturation of autophagosome, and therefore blocks the autophagic degradation in MDA-MB-231 cells.

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更新日期/Last Update: 2015-04-02