自噬对宫颈癌细胞增殖和迁移能力的影响

doi:10.11904/j.issn.1002-3070.2016.06.001

实用肿瘤学杂志 ›› 2016, Vol. 30 ›› Issue (6): 481-486.doi: 10.11904/j.issn.1002-3070.2016.06.001

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自噬对宫颈癌细胞增殖和迁移能力的影响

杨毅，丁妍，于爱清，严世荣

武汉大学附属中南医院基因诊断中心(武汉 442000)

收稿日期:2016-07-03 出版日期:2016-12-31 发布日期:2016-12-27
通讯作者: 严世荣,E-mail:249105714@qq.com
作者简介:杨毅，女，(1991-)，硕士研究生，从事基因诊断的相关研究
基金资助:
国家自然科学基金资助项目(81541147);湖北医药学院博士研究生启动金资助项目(2015QDJZR01)

Effects of autophagy on the proliferation and migration of cervical cancer cells

YANG Yi,DING Yan,YU Aiqing,YAN Shirong

Center for Gene Diagnosis,Zhongnan Hospital of Wuhan University,Wuhan 430000,China

Received:2016-07-03 Online:2016-12-31 Published:2016-12-27

摘要/Abstract

摘要： 目的观察自噬对宫颈癌细胞增殖迁移能力的影响，并探讨其可能机制。方法采用不同浓度雷帕霉素诱导人宫颈癌HeLa细胞，吖啶橙染色观察自噬小体的形成，Western blot检测自噬相关基因LC3B以及PI3K/Akt/mTOR通路蛋白的表达情况，采用RTCA仪器实时检测细胞的增殖能力，Transwell小室检测细胞的迁移能力。应用自噬抑制剂3-methyladenine(3-MA)阻断宫颈癌细胞自噬后，采用RTCA仪器实时检测细胞的增殖能力，Transwell小室检测细胞的迁移能力。构建带有绿色荧光蛋白的LC3B质粒，转入HeLa细胞，通过G418药物筛选转染阳性细胞，荧光显微镜观察LC3B的细胞定位，Western blot检测LC3B表达。RTCA仪器实时检测野生型和LC3B过表达HeLa细胞的增殖能力，Western blot检测PI3K/Akt/mTOR通路蛋白的表达。结果雷帕霉素诱导后，宫颈癌HeLa细胞自噬水平增高，增殖和迁移能力有所下降;PI3K/Akt/mTOR通路蛋白表达改变(P＜0.05)。自噬抑制剂3-MA诱导HeLa细胞增殖和迁移水平下降(P＜0.05)。荧光显微镜和Western blot结果显示构建的LC3质粒成功转入Hela细胞中并且能够抑制宫颈癌细胞的增殖(P＜0.05)。结论在一定范围内，随着自噬水平的升高，HeLa细胞增殖和迁移能力降低，可能是通过PI3K/Akt/mTOR信号通路发挥作用。通过基因靶向扰乱自噬关键基因(如LC3等)而改变宫颈癌细胞自噬水平可能为宫颈癌的治疗带来新策略。

关键词: 自噬, 宫颈癌细胞, 雷帕霉素, LC3

Abstract: Objective To investigate the effect of autophagy on the proliferation and migration of cervical cancer cells,as well as the underlining mechanisms.Methods Rapamycin was used to induce the autophagy in HeLa cells,formation of autophagosomes was observed by staining with acridine orange under fluorescence microscope.Western blot was used to detect the expression of LC3 and PI3K/Akt/mTOR in HeLa cells.The LC3 plasmid was transfected into HeLa cells.The distribution of LC3 in cells and the expression of LC3 was identified by fluorescence microscope and Western blot,respectively.The autophagy was inhibited with 3-methyladenine(3-MA)in HeLa cells.The cell proliferation was monitored by RTCA real-time instrument.Transwell chamber was carried out to assess cell migration.Results After 6h of rapamycin treatment,the expression of LC3B was increased in HeLa cells(P＜0.05).The proliferative and migration ability were weakened compared to wild type HeLa cells(P＜0.05).The same results in the presence of 3-MA.The expression of PI3K/AKT/mTOR pathway proteins were activated by rapamycin treatment and LC3 overexpression(P＜0.05).Conclusion Autophagy can suppress the proliferation and migration in cervical cancer cells,which may relate to PI3K/Akt/mTOR pathway.

Key words: Autophagy, Cervical cancer cells, Rapamycin, LC3B

中图分类号:

R737.3

杨毅，丁妍，于爱清，严世荣. 自噬对宫颈癌细胞增殖和迁移能力的影响[J]. 实用肿瘤学杂志, 2016, 30(6): 481-486.

YANG Yi,DING Yan,YU Aiqing,YAN Shirong. Effects of autophagy on the proliferation and migration of cervical cancer cells[J]. PRACTICAL ONCOLOGY JOURNAL, 2016, 30(6): 481-486.

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自噬对宫颈癌细胞增殖和迁移能力的影响

Effects of autophagy on the proliferation and migration of cervical cancer cells

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