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中华乳腺病杂志(电子版)

中华乳腺病杂志(电子版) ›› 2015, Vol. 09 ›› Issue (06) : 372 -375. doi: 10.3877/cma. j. issn.1674-0807.2015.06.005

论著

高表达twist 基因促进三阴性乳腺癌SUM159 细胞干性特征的初步研究
曾真1, 甘霖1, 王亚冬1, 闵捷1, 王婷1, 廖登辉1, 吕钢1,†()   
  1. 1.400011 重庆市第一中医院乳腺甲状腺科
  • 收稿日期:2015-06-15 出版日期:2015-12-01
  • 通信作者: 吕钢
  • 基金资助:
    2014 年重庆市科学技术委员会前沿与应用基础研究项目(cstc2014jcyjA83)

Preliminary study on high expression of twist gene promoting the stemness related characteristics of triple negative breast cancer SUM159 cells

Zhen Zeng1, Lin Gan1, Yadong Wang1, Jie Min1, Ting Wang1, Denghui Liao1, Gang Lyu1,†()   

  1. 1.Department of Breast and Thyroid Surgery, the First Hospital of Traditional Chinese Medicine of Chongqing Municipality, Chongqing 400011, China
  • Received:2015-06-15 Published:2015-12-01
  • Corresponding author: Gang Lyu
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曾真, 甘霖, 王亚冬, 闵捷, 王婷, 廖登辉, 吕钢. 高表达twist 基因促进三阴性乳腺癌SUM159 细胞干性特征的初步研究[J/OL]. 中华乳腺病杂志(电子版), 2015, 09(06): 372-375.

Zhen Zeng, Lin Gan, Yadong Wang, Jie Min, Ting Wang, Denghui Liao, Gang Lyu. Preliminary study on high expression of twist gene promoting the stemness related characteristics of triple negative breast cancer SUM159 cells[J/OL]. Chinese Journal of Breast Disease(Electronic Edition), 2015, 09(06): 372-375.

目的

探讨高表达twist 基因对乳腺癌SUM159 细胞干性特征的影响。

方法

用慢病毒构建过表达twist 基因的乳腺癌SUM159 细胞作为实验组(SUM159/twist),空载体构建对照组细胞(SUM159/vector)。 利用RT-PCR 检测twist 转染前后mRNA 水平变化;通过平板克隆形成、成球实验、流式细胞仪检测肿瘤干细胞比例,并用RT-PCR 检测干性相关基因SOX2、OCT4、BMI1 的mRNA 表达水平。mRNA 及细胞克隆数等计量资料以¯x±s 表示,组间比较采用t 检验。

结果

成功构建过表达twist 基因的乳腺癌SUM159 细胞,SUM159/twist 细胞中的twist mRNA 表达为2.04±0.15,高于SUM159/vector 细胞的0.49±0.45(t=-16.33,P ＜0.001);SUM159/twist 细胞的克隆菌落数目(92.75±4.85)明显高于SUM159/vector 细胞(44.50±5.19)(t=13.56,P＜0.001);成球实验显示:SUM159/twist 细胞形成的细胞球数目(46.75±4.11)显著高于SUM159/vector 细胞(22.50±3.41)(t=9.07,P＜0.001);利用流式细胞仪检测SUM159/twist 和SUM159/vector 细胞中的ALDH1+细胞比例发现:SUM159/twist 细胞中ALDH1+细胞比例为(9.63±0.89)%,显著高于SUM159/vector 细胞中的(2.31±0.21)%(t=13.85,P＜0.001);SUM159/twist 细胞中SOX2、OCT4、BMI1 的表达(12.39±0.63、13.35±1.56、6.48±0.96)均高于SUM159/vector 细胞(1.61±0.33、2.67±0.28、2.70±0.35)(t=-29.68,-11.61,6.43;P 均＜0.001)。

结论

twist 基因能够增强乳腺癌SUM159 细胞干性能力。

关键词: 乳腺肿瘤, 转录因子, 肿瘤干细胞

Objective

To investigate the impact of high expression of twist gene on the stemness related characteristics of breast cancer SUM159 cells.

Methods

We established a twist-overexpression breast cancer SUM159 cell line as experimental group (SUM159/twist), and the cells with blank vector construction served as control group (SUM159/vector). The twist mRNA level were detected before and after twist gene transfection using RT-PCR. The proportion of cancer stem cells was investigated by flat clone formation,mammosphere assay and flow cytometry. The mRNA expressions of stemness-related genes SOX2, OCT4 and BMI1 were detected by RT-PCR. All data were expressed as ¯x±s and processed by t test.

Results

SUM159 cells with twist gene overexpression were successfully established. The relative expression of twist mRNA in SUM159/twist cells was 2.04±0.15, significantly higher than 0.49±0.45 in SUM159/vector cells(t=-16.33,P＜0.001).The colony number of clone formation was 92.75±4.85 in SUM159/twist cells, significantly higher than 44.50±5.19 in SUM159/vector cells(t=13.56, P＜0.001).The mammosphere number was 46.75±4.11 in SUM159/twist cells and 22.50±3.41 in SUM159/vector cells, and the difference was statistically significant(t=9.07, P＜0.001). Flow cytometry results showed that the proportion of ALDH1+ was (9.63±0.89)% in SUM159/twist cells and (2.31±0.21)% in SUM159/vector cells, indicating a significant difference(t =13.85, P＜0.001). The relative mRNA expressions of SOX2, OCT4 and BMI1 in SUM159/twist cells were 12.39±0.63,13.35±1.56 and 6.48±0.96, significantly higher than 1.61±0.33,2.67±0.28 and 2.70±0.35 in SUM159/vector cells respectively(t=-29.68,-11.61,6.43,all P＜0.001).

Conclusion

The twist gene promotes the stemness related capability of breast cancer SUM159 cells.

Key words: Breast neoplasms, Transcription factor, Neoplastic stem cells
图1 twist 基因过表达后对乳腺癌SUM159 细胞克隆形成能力的影响
图2 twist 基因过表达后对乳腺癌SUM159 细胞成球能力的影响
图3 twist 基因过表达后对乳腺癌SUM159 细胞中ALDH1+细胞比例影响的流式细胞图 注:SSC-H 代表侧向散射角
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