mTOR通路抑制剂依维莫司对宫颈癌SiHa细胞恶性行为的影响  

Effect of the mTOR inhibitor everolimus on the malignant behavior of cervical cancer SiHa cells

作　　者：吴秋芳[1] 唐周舟[1] 张定富[1] 

机构地区：[1]湖北荆州华中科技大学同济医学院附属荆州医院肿瘤科,434020

出　　处：《临床肿瘤学杂志》2015年第8期685-689,共5页Chinese Clinical Oncology

摘　　要：目的：探讨依维莫司（ EVE）对人宫颈癌SiHa细胞增殖、凋亡及上皮间质转化（ EMT）的影响。方法常规培养SiHa细胞达对数生长期后，采用终浓度为1、10、100μmol／L EVE处理SiHa细胞，采用四甲基偶氮唑盐比色法测定对照组及不同浓度EVE处理24、48、72和96 h后的细胞增殖情况，分别采用Annexin V-FITC／PI双染联合流式细胞术和实时荧光定量聚合酶链反应（ qRT-PCR）检测不同浓度EVE处理96 h的早期和晚期凋亡率及凋亡相关基因（ Bax、Bad和Bcl-2）的表达情况，Western blotting和免疫荧光法检测各组处理96 h后的EMT相关标记分子包括上皮型钙粘附素（ E-cad）、纤维连接蛋白（ FN）和波形蛋白（ Vim）的水平。结果 EVE在1∽100μmol／L范围内对SiHa细胞有增殖抑制作用，且增殖抑制率呈剂量和时间依赖性，除1μmol／L作用24 h外，其余浓度及作用时间的增殖抑制率均高于对照组（ P＜0.05）；与对照组相比，EVE处理96 h后的早期、晚期凋亡率及Bax、Bad的mRNA水平均升高，而Bcl-2 mRNA水平降低，差异均有统计学意义（ P＜0.05），且呈剂量依赖性；EVE处理后的E-cad水平均高于对照组，而FN和Vim水平均低于对照组（ P＜0.05）。结论采用EVE阻断mTOR通路对宫颈癌SiHa细胞有毒性作用，不仅抑制其增殖且诱导凋亡，可能与抑制其EMT过程有关。Objective To investigate the effect of an mTOR pathway inhibitor everolimus（ EVE） on the proliferation, apopto-sis and epithelial mesenchymal transition（ EMT） of human cervical cancer SiHa cells. Methods After being cultured in normal cul-ture, the SiHa cells were treated with EVE at final concentrations of 1, 10 and 100 μmol/L. The cell proliferation was measured by methyl thiazolyl tetrazolium in the control group and different concentrations of EVE at 24, 48, 72 and 96 h after exposure. The early-and late-apoptosis rate was measured by Annexin V/PI double staining via flow cytometry, and the apoptosis related gene expression （ Bax, Bad and Bcl-2） was assessed by real-time fluorescence quantitative polymerase chain reaction. Western blotting and immunoflu-orescence assay were used to detect the levels of EMT related marker molecules, including E-cadherin（ E-cad） , Fibronectin（ FN） and Vimentin（Vim） at 96 h after EVE exposure. Results EVE had inhibitory effect on the proliferation of SiHa cells in the range of 1-100μmol/L in a dose-and time-dependent manner. Except for 1μmol/L at 24 h, the proliferation inhibition rates of the remaining concen-tration and action times were higher than that of the control group（ P〈0.05） . Compared with the control group, EVE treatment for 96 h could dose-dependently increase the apoptosis rate and mRNA levels of Bax and Bad but decrease Bcl-2 level（P〈0.05）. Compared with the control group, the level of E-cad was increased but the levels of Vim and FN were decreased after treatment with different con-centrations EVE for 96 h（ P〈0.05） . Conclusion Blocking mTOR pathway by EVE has a toxic effect on cervical cancer SiHa cells, which not only inhibit the proliferation and induce apoptosis, but may be related to the inhibition of EMT process.

关 键 词：依维莫司 宫颈癌 细胞增殖 凋亡 上皮间质转化 

分 类 号：R737.33[医药卫生—肿瘤]