NDRG2 Inhibits Migration of Breast Cancer Cells Through Upregulating PTEN Expression
-
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.
-
Key words:
- Breast neoplasms /
- NDRG2 /
- PTEN /
- Cell migration
-
-
![]()
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)
![]()
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
![]()
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
-
[1] Jin X, Mu P. Targeting Breast Cancer Metastasis[J]. Breast Cancer (Auckl), 2015, 9(Suppl 1): 23-34. https://www.broadinstitute.org/publications/broad7253
[2] Hu W, Fan C, Jiang P, et al. Emerging role of N-myc downstream-regulated gene 2 (NDRG2) in cancer[J]. Oncotarget, 2016, 7(1): 209-23. https://www.researchgate.net/publication/283292608_Emerging_role_of_N-myc_downstream-regulated_gene_2_NDRG2_in_cancer
[3] Lorentzen A, Lewinsky RH, Bornholdt J, et al. Expression profile of the N-myc Downstream Regulated Gene 2 (NDRG2) in human cancers with focus on breast cancer[J]. BMC Cancer, 2011, 11: 14. doi: 10.1186/1471-2407-11-14
[4] Mordalska A, Latek J, Ferenc T, et al. Evaluation of NDRG2 gene expression in primary papillary thyroid carcinoma and in metastases of this neoplasm to regional lymph nodes[J]. Thyroid Res, 2010, 3(1): 6. doi: 10.1186/1756-6614-3-6
[5] Park MY, Choi SC, Lee HS, et al. A quantitative analysis of N-myc downstream regulated gene 2 (NDRG 2) in human tissues and cell lysates by reverse-phase protein microarray[J]. Clin Chim Acta, 2008, 387(1-2): 84-9. doi: 10.1016/j.cca.2007.09.010
[6] Piepoli A, Cotugno R, Merla G, et al. Promoter methylation correlates with reduced NDRG2 expression in advanced colon tumour[J]. BMC Med Genomics, 2009, 2:11. doi: 10.1186/1755-8794-2-11
[7] Shi H, Li N, Li S, et al. Expression of NDRG2 in esophageal squamous cell carcinoma[J]. Cancer Sci, 2010, 101(5): 1292-9. doi: 10.1111/cas.2010.101.issue-5
[8] Skiriute D, Tamasauskas S, Asmoniene V, et al. Tumor grade-related NDRG2 gene expression in primary and recurrent intracranial meningiomas[J]. J Neurooncol, 2011, 102(1): 89-94. doi: 10.1007/s11060-010-0291-9
[9] Golestan A MSc, Mojtahedi Z PhD, Ghalamfarsa G PhD, et al. The Effects of NDRG2 Overexpression on Cell Proliferation and Invasiveness of SW48 Colorectal Cancer Cell Line[J]. Iran J Med Sci, 2015, 40(5): 430-9. https://www.researchgate.net/publication/280777221_The_Effects_of_NDRG2_Overexpression_on_Cell_Proliferation_and_Invasiveness_of_SW48_Colorectal_Cancer_Cell_Line?_sg=7iXZMZClCSwPNsJw8qwKdtN16ZFxqsL9uKTUoII-j4fzS7brc3coZpO05Z6a5m9FkREPlfJf4V3anzLIPXKmUZkPn9G4Rgu7VCKOjhZKIcA
[10] Kim YJ, Yoon SY, Kim JT, et al. NDRG2 suppresses cell proliferation through down-regulation of AP-1 activity in human colon carcinoma cells[J]. Int J Cancer, 2009, 124(1): 7-15. doi: 10.1002/ijc.v124:1
[11] Cao W, Zhang JL, Feng DY, et al. The effect of adenovirus-conjugated NDRG2 on p53-mediated apoptosis of hepatocarcinoma cells through attenuation of nucleotide excision repair capacity[J]. Biomaterials, 2014, 35(3): 993-1003. doi: 10.1016/j.biomaterials.2013.09.096
[12] Ma J, Liu W, Guo H, et al. N-myc downstream-regulated gene 2 expression is associated with glucose transport and correlated with prognosis in breast carcinoma[J]. Breast Cancer Res, 2014, 16(2):R27. http://or.nsfc.gov.cn/handle/00001903-5/361414
[13] Zhou X, Liu Y, You J, et al. Myosin light-chain kinase contributes to the proliferation and migration of breast cancer cells through cross-talk with activated ERK1/2[J]. Cancer Lett, 2008, 270(2): 312-27. doi: 10.1016/j.canlet.2008.05.028
[14] Ma J, Liu W, Yan X, et al. Inhibition of endothelial cell proliferation and tumor angiogenesis by up-regulating NDRG2 expression in breast cancer cells[J]. PLoS One, 2012, 7(2): e32368. doi: 10.1371/journal.pone.0032368
[15] Zheng J, Liu Q, Li Y, et al. NDRG2 expression regulates CD24 and metastatic potential of breast cancer cells[J]. Asian Pac J Cancer Prev, 2010, 11(6): 1817-21. http://www.sigmaaldrich.com/catalog/papers/21338239
[16] Baumann P, Thiele W, Cremers N, et al. CD24 interacts with and promotes the activity of c-src within lipid rafts in breast cancer cells, thereby increasing integrin-dependent adhesion[J]. Cell Mol Life Sci, 2012, 69(3): 435-48. doi: 10.1007/s00018-011-0756-9
[17] Zheng J, Li Y, Yang J, et al. NDRG2 inhibits hepatocellular carcinoma adhesion, migration and invasion by regulating CD24 expression[J]. BMC Cancer, 2011, 11: 251:1-9. http://www.docin.com/p-1397964537.html
[18] Guo Y, Ma J, Wu L, et al. Hyperthermia-induced NDRG2 upregulation inhibits the invasion of human hepatocellular carcinoma via suppressing ERK1/2 signaling pathway[J]. PLoS One, 2013, 8(4): e61079. doi: 10.1371/journal.pone.0061079
[19] Shon SK, Kim A, Kim JY, et al. Bone morphogenetic protein-4 induced by NDRG2 expression inhibits MMP-9 activity in breast cancer cells[J]. Biochem Biophys Res Commun, 2009, 385(2): 198-203. doi: 10.1016/j.bbrc.2009.05.038
[20] Kim A, Kim MJ, Yang Y, et al. Suppression of NF-kappaB activity by NDRG2 expression attenuates the invasive potential of highly malignant tumor cells[J]. Carcinogenesis, 2009, 30(6): 927-36. doi: 10.1093/carcin/bgp072
[21] Wang Y, Dai B. PTEN genomic deletion defines favorable prognostic biomarkers in localized prostate cancer: a systematic review and meta-analysis[J]. Int J Clin Exp Med, 2015, 8(4): 5430-7.
[22] Xu W, Yang Z, Zhou SF, et al. Posttranslational regulation of phosphatase and tensin homolog (PTEN) and its functional impact on cancer behaviors[J]. Drug Des Devel Ther, 2014, 8:1745-51. https://www.researchgate.net/publication/267624896_Posttranslational_regulation_of_phosphatase_and_tensin_homolog_PTEN_and_its_functional_impact_on_cancer_behaviors
[23] Zhang LL, Liu J, Lei S, et al. PTEN inhibits the invasion and metastasis of gastric cancer via downregulation of FAK expression[J]. Cell Signal, 2014, 26(5): 1011-20. doi: 10.1016/j.cellsig.2014.01.025
[24] Ebbesen SH, Scaltriti M, Bialucha CU, et al. Pten loss promotes MAPK pathway dependency in HER2/neu breast carcinomas[J]. Proc Natl Acad Sci U S A, 2016, 113(11): 3030-5. doi: 10.1073/pnas.1523693113
[25] Tian T, Nan KJ, Guo H, et al. PTEN inhibits the migration and invasion of HepG2 cells by coordinately decreasing MMP expression via the PI3K/Akt pathway[J]. Oncol Rep, 2010, 23(6): 1593-600. https://www.researchgate.net/publication/43352779_PTEN_inhibits_the_migration_and_invasion_of_HepG2_cells_by_coordinately_decreasing_MMP_expression_via_the_PI3KAkt_pathway?_sg=kyDcvnf4o-K1z45lR1eN1GMskR5Qe6glEFmz8XxveLvfz6BZz3qTLJMPUTblT5KRCL2GXtnR0B4lz7MPz8zufw
[26] Zhu L, Loo WT, Louis WC. PTEN and VEGF: possible predictors for sentinel lymph node micro-metastasis in breast cancer[J]. Biomed Pharmacother, 2007, 61(9): 558-61. doi: 10.1016/j.biopha.2007.08.015
下载:
计量
- 文章访问数: 1347
- HTML全文浏览量: 313
- PDF下载量: 561
