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中国临床药理学与治疗学 ›› 2014, Vol. 19 ›› Issue (8): 851-855.

• 基础研究 • 上一篇    下一篇

左旋一叶萩碱诱导人非小细胞肺癌A549细胞自噬机制的研究

张刚, 陈冬云, 程静, 吉兆宁   

  1. 皖南医学院附属弋矶山医院肿瘤内科,芜湖 241001,安徽
  • 收稿日期:2014-03-11 修回日期:2014-07-17 出版日期:2014-08-26 发布日期:2014-08-26
  • 通讯作者: 吉兆宁,男,教授,硕士研究生导师,研究方向:肿瘤化学及生物防治机理和临床应用。 Tel: 0553-5739060 E-mail: jzning@163.com
  • 作者简介:张刚,男,硕士研究生,研究方向:肿瘤化学及生物防治机理和临床应用。 Tel: 15855976955 E-mail: 921950782@qq.com
  • 基金资助:
    国家自然科学基金(81241102); 安徽省卫生厅中医药课题(2012zy59)

L-Securinine induced the human non small cell lung cancer A549 cell autophagy and its molecular mechanism

ZHANG Gang, CHEN Dong-yun, CHENG Jing, JI Zhao-ning   

  1. Cancer Center, Yijishan Hospital of Wannan Medical College, Wuhu 241001, Anhui, China
  • Received:2014-03-11 Revised:2014-07-17 Online:2014-08-26 Published:2014-08-26

摘要: 目的 研究左旋一叶萩碱诱导人非小细胞肺癌A549细胞自噬的机制。方法 常规体外培养人非小细胞肺癌A549细胞株,取对数生长期的细胞传代 24 h 后加入不同浓度(6.25、12.5、25、50、100、200 μmol/L)左旋一叶萩碱处理, 分别继续培养24、36、48 h 后,用CCK-8法测定细胞增殖活性;倒置显微镜下观察对照组和各实验组的细胞形态变化;分别用不同浓度(0、20 μmol/L)左旋一叶萩碱作用A549细胞 48 h 后,进行单丹磺酰尸胺(monodansylcadaverine,MDC)染色,分别通过荧光显微镜检测细胞自噬发生情况;荧光定量RT-PCR法检测Beclin 1在mRNA水平表达变化情况。结果 左旋一叶萩碱可以抑制A549细胞增殖,并且呈浓度和时间依赖性(P<0.05)。其中,21.204 μmol/L左旋一叶萩碱作用A549细胞48 h后,细胞死亡率接近50%;倒置显微镜观察到药物作用后细胞形态发生了明显变化;随着左旋一叶萩碱剂量增加,MDC阳性细胞的荧光强度及细胞内MDC荧光颗粒数目明显增加。另外,研究还发现随着药物浓度的上升,自噬基因Beclin 1的表达明显增强。结论 左旋一叶萩碱具有抑制人非小细胞肺癌A549细胞增殖、诱导细胞自噬性凋亡,其作用具有浓度和时间依赖性。左旋一叶萩碱上调自噬基因Beclin 1 mRNA的表达水平可能是其诱导A549细胞自噬性凋亡的机制之一。

关键词: 左旋一叶萩碱, 非小细胞肺癌, A549细胞, 自噬

Abstract: AIM: To explore the mechanism of L-securinine induced autophagy of human non small cell lung cancer A549 cell autophagy. METHODS: A549 cell lines were cultured in vitro and L-securinine at different concentrations (6.25,12.5,25,50,100,200 μmol/L) was added 24 h after cell lines were transferred. Then the plates were cultured for 24, 36 and 48 h. CCK-8 method was used to detect the antitumor effect of human non small cell lung cancer A549 in vitro. Inverted microscope was used to observe A549 cells treated with L-securinine morphological changes. The A549 cells were treated with different concentrations (0, 20 μmol/L) of L-securinine for 24 h.Then autophagy was analyzed by fluorescence microscope following staining with monodansylcadaverine (MDC). The mRNA levels of Beclin-1 were detected using real-time RT-PCR pre and post-treatment of L-securinine. RESULTS: The generation depression effects of A549 cells cultured in vitro were detected by CCK-8 method (P<0.05), and there were dosage-time dependent relationships. The morphology of cells become small and round, the process of cell division got less were observed. L-securinine-treated cells exhibited higher fluorescent density and more MDC-labeled particles in A549 cells compared with the control group. Beclin-1 expression enhanced with L-securinine concentration increased. CONCLUSION: L-securinine has an anti-tumor effect against human non small cell lung cancer A549 cell. The L-securinine can induce striking autophagy in A549 cell in vitro. The autophagy induced by L-securinine is related with upregulating the expression of autophagy gene Beclin-1.

Key words: L-securinine, non small cell lung cancer, A549 cell, autophagic

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