NDRG2 Inhibits Migration of Breast Cancer Cells Through Upregulating PTEN Expression
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摘要:目的
利用具有相同遗传背景但不同转移能力的乳腺癌细胞模型MCF-7、LM-MCF-7以及MDA-MB-231细胞，研究NDRG2调控乳腺癌细胞迁移的作用及机制。
方法利用RT-PCR和蛋白免疫印迹法检测MCF-7、LM-MCF-7以及MDA-MB-231细胞中NDRG2和PTEN mRNA和蛋白表达水平；构建pCMV-NDRG2和pCMV-PTEN表达载体以及设计合成NDRG2和PTEN siRNA，通过转染过表达或沉默NDRG2和PTEN表达，Transwell迁移实验检测转染后细胞迁移能力改变，Western blot检测NDRG2和PTEN表达调控关系。
结果MCF-7细胞中NDRG2和PTEN mRNA水平和蛋白表达水平显著高于LM-MCF-7和MDA-MB-231细胞（P < 0.05）；在LM-MCF-7和MDA-MB-231中上调NDRG2表达可降低细胞迁移能力至13.02%和26.33%（P < 0.01）；在MCF-7细胞中沉默NDRG2表达可增强细胞迁移能力至354.62%（P < 0.01）；蛋白免疫印迹检测显示PTEN表达受NDRG2调控；沉默（或过表达）NDRG2表达，同时过表达（或沉默）PTEN，MCF-7（或LM-MCF-7）细胞迁移能力未发生显著变化（P > 0.05）。
结论乳腺癌细胞中，NDRG2的表达下调，导致PTEN表达下降，解除了对乳腺癌细胞迁移的抑制作用。NDRG2/PTEN信号途径的沉默是乳腺癌细胞获得高迁移能力的重要机制。
Abstract:ObjectiveTo investigate the effects of N-Myc downstream regulated gene 2 (NDRG2) on the migration of human breast cancer cells by using two parallel cell lines MCF-7 and LM-MCF with different metastatic abilities.
MethodsThe expression level of NDRG2 and PTEN in breast cancer cells were detected by RT-PCR and Western blot. The effects of overexpressed or down-regulated NDRG2 on the migration of breast cancer cells were investigated by Transwell migration test. The regulation mechanism between NDRG2 and PTEN on cell migration was detected via co-transfecting breast cancer cells with pCMV-NDRG2 and PTEN siRNA (or NDRG2 siRNA and pCMV-PTEN).
ResultsThe mRNA and protein expression levels of NDRG2 and PTEN in MCF-7 cells were higher than those in LM-MCF-7/MDA-MB-231 cells (P < 0.05). Overexpressed NDRG2 via transfecting LM-MCF-7 and MDA-MB-231 cells with pCMV-NDRG2 could inhibit cell migration ability to 13.02% and 26.33%(P < 0.01) respectively. Downregulated NDRG2 expression in MCF-7 cells could enhance cell migration ability to 354.62%(P < 0.01). Interestingly, the
Resultsof Western blot and Transwell assay revealed that the downregulation of NDRG2 expression resulted in the downregulation of PTEN expression. Co-transfecting LM-MCF-7/MDA-MB-231(or MCF-7) cells with pCMV-NDRG2/PETN siRNA (or pCMV-PTEN/NDRG2 siRNA), the migration ability of cells had no significant changes (P > 0.05).
ConclusionThe downregulation of the expression of NDRG2 could inhibit the expression of PTEN, which lead to the losing control of the migration of breast cancer cells. Downregulating NDRG2/PTEN pathway is a significant mechanism for breast cancer cells acquiring high migration ability.
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Key words:
- Breast neoplasms /
- NDRG2 /
- PTEN /
- Cell migration
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图 1 RT-PCR (A)和Western blot (B)法检测MCF-7、LM-MCF-7和MDA-MB-231细胞中NDRG2和PTEN的mRNA和蛋白表达水平
Figure 1 mRNA and protein levels of NDRG2 and PTEN in MCF-7, LM-MCF-7 and MDA-MB-231 cells examined via RT-PCR (A) and Western blot (B)
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图 2 双酶切鉴定pCMV-NDRG2和pCMV-PTEN质粒
Figure 2 pCMV-NDRG2 and pCMV-PTEN plasmid digested by EcoRⅠ/SalⅠand BamHⅠ/HindⅢ respectively
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图 3 Western blot和RT-PCR法检测干扰NDRG2表达的MCF-7细胞(A)和过表达NDRG2的LM-MCF-7细胞(B)和MDA-MB-231细胞(C)中的PTEN mRNA和蛋白表达水平
Figure 3 mRNA and protein expression levels of PTEN detected by RT-PCR and Western bot after transfecting MCF-7 cells with NDRG2 siRNA (A), or transfecting LM-MCF-7(B) and MDA-MB-231(C) cells with pCMV-NDRG2
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