过表达WNT5B可抑制人乳腺癌MDA-MB-231细胞迁移和侵袭
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摘要
目的:探讨过表达WNT5B对人乳腺癌MDA-MB-231细胞迁移和侵袭能力的影响,并分析其可能的作用机制。方法:将含增强型绿色荧光蛋白(enhanced greenuorescent protein,EGFP)基因的WNT5B过表达重组载体pcDNA3.1-WNT5B+EGFP转染人乳腺癌MDA-MB-231细胞后,采用实时荧光定量PCR法和蛋白质印迹法分别检测细胞中WNT5B mRNA及蛋白的表达水平,划痕愈合实验和Transwell小室法检测细胞的迁移和侵袭能力,蛋白质印迹法检测上皮-间质转化(epithelial-mesenchymal transition,EMT)相关分子标志物N-cadherin、E-cadherin和Vimentin的表达。结果:与未转染组和转染空载体组相比,转染WNT5B过表达载体组MDA-MB-231细胞中WNT5B mRNA(P <0.01)和蛋白(P <0.05)表达水平明显上调;过表达WNT5B后,MDA-MB-231细胞的迁移(P <0.05)和侵袭(P <0.01)能力明显下降,E-cadherin的表达水平明显上调(P <0.05),N-cadherin和Vimentin的表达水平均明显下调(P值均<0.05)。结论:过表达WNT5B可抑制人乳腺癌MDA-MB-231细胞的迁移和侵袭,这种变化可能与其调控EMT的进程有关。
Objective: To investigate the effects of WNT5 B over-expression on the invasion and migration of human breast cancer MDA-MB-231 cells,and to explore its possible mechanisms.Methods: After the recombinant vector pcDNA3.1-WNT5 B+EGFP containing enhanced green fluorescent protein(EGFP) and WNT 5 B genes was transfected into human breast cancer MDA-MB-231 cells, the expression levels of WNT5 B mRNA and protein were examined by real-time fluorescent quantitative PCR and Western blotting, respectively. The abilities ofcell migration and invasion were analyzed by wound-healing test and Transwell chamber assay,respectively. Furthermore, the expression levels of epithelial-mesenchymal transition(EMT)-related molecules N-cadherin, E-cadherin and Vimentin were detected by Western blotting.Results: Compared with the untransfection and the empty vector-transfection groups, the expression levels of WNT5 B mRNA(P < 0.01) and protein(P < 0.05) in MDA-MB-231 cells transfected with the recombinant vector pcDNA3.1-WNT5 B+EGFP were up-regulated. The abilities of migration(P < 0.05) and invasion(P < 0.01) of MDA-MB-231 cells after WNT5 B over-expression were significantly decreased. The expression level of E-cadherin in MDAMB-231 cells with WNT5 B over-expression was up-regulated(P < 0.05), whereas the expression levels of N-cadherin and Vimentin were down-regulated(both P < 0.05).Conclusion: WNT5 B over-expression can inhibit the migration and invasion of breast cancer MDA-MB-231 cells, which may be related to the regulation of EMT process.
Objective: To investigate the effects of WNT5 B over-expression on the invasion and migration of human breast cancer MDA-MB-231 cells,and to explore its possible mechanisms.Methods: After the recombinant vector pcDNA3.1-WNT5 B+EGFP containing enhanced green fluorescent protein(EGFP) and WNT 5 B genes was transfected into human breast cancer MDA-MB-231 cells, the expression levels of WNT5 B mRNA and protein were examined by real-time fluorescent quantitative PCR and Western blotting, respectively. The abilities ofcell migration and invasion were analyzed by wound-healing test and Transwell chamber assay,respectively. Furthermore, the expression levels of epithelial-mesenchymal transition(EMT)-related molecules N-cadherin, E-cadherin and Vimentin were detected by Western blotting.Results: Compared with the untransfection and the empty vector-transfection groups, the expression levels of WNT5 B mRNA(P < 0.01) and protein(P < 0.05) in MDA-MB-231 cells transfected with the recombinant vector pcDNA3.1-WNT5 B+EGFP were up-regulated. The abilities of migration(P < 0.05) and invasion(P < 0.01) of MDA-MB-231 cells after WNT5 B over-expression were significantly decreased. The expression level of E-cadherin in MDAMB-231 cells with WNT5 B over-expression was up-regulated(P < 0.05), whereas the expression levels of N-cadherin and Vimentin were down-regulated(both P < 0.05).Conclusion: WNT5 B over-expression can inhibit the migration and invasion of breast cancer MDA-MB-231 cells, which may be related to the regulation of EMT process.
引文
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[15]赵鹏,刘冬,张卉,等.RBMS3通过Wnt/β-catenin信号通路抑制胃癌细胞侵袭及其上皮-间质转化[J].肿瘤,2017,37(10):1032-1040.
[2]LI Z,QIU Y,LU W,et al.Immunotherapeutic interventions of triple negative breast cancer[J].JTransl Med,2018,16(1):147.
[3]THRASIVOULOU C,MILLAR M,AHMED A.Activation of intracellular calcium by multiple Wnt ligands and translocation ofβ-catenin into the nucleus:a convergent model of Wnt/Ca2+and Wnt/β-catenin pathways[J].J Biol Chem,2013,288(50):35651-35659.
[4]KIM S,NIE H,NESIN V,et al.The polycystin complex mediates WNT/Ca2+signaling[J].Nat Cell Biol,2016,18(7):752-764.
[5]李佳纹,王诗卓,欧阳玲.WNT5B在卵巢上皮癌中的表达及意义[J].现代肿瘤医学,2017,25(24):4030-4033.
[6]HARADA T,YAMAMOTO H,KISHIDA S,et al.Wnt5b-associated exosomes promote cancer cell migration and proliferation[J].Cancer Sci,2017,108(1):42-52.
[7]SUN D,QIN L,XU Y,et al.Influence of adriamycin on changes in Nanog,Oct-4,Sox2,ARID1 and Wnt5b expression in liver cancer stem cells[J].World J Gastroentero,2014,20(22):6974-6980.
[8]WANG SH,CHANG JS,HSIAO JR,et al.Tumour cell-derived WNT5Bmodulates in vitro lymphangiogenesis via induction of partial endothelialmesenchymal transition of lymphatic endothelial cells[J].Oncogene,2017,36(11):1503-1515.
[9]ZHANG Y,L I N L,J I N Y,et al.Overexpression of WNT5B promotes COLO 205 cell migration and invasion through the JNK signaling pathway[J].Oncol Rep,2016,36(1):23-30.
[10]TAKESHITA A,IWAI S,MORITA Y,et al.Wnt5b promotes the cell motility essential for metastasis of oral squamous cell carcinoma through active Cdc42 and RhoA[J].Int J Oncol 2014,44(1):59-68.
[11]刘洪涛,周凡涵,申媛媛,等.WNT5B在乳腺癌中的表达及意义[J].中国病理生理杂志,2015,31(6):1032-1035.
[12]周凡涵,厉红元.WNT5B过表达对人乳腺癌MCF-7细胞活力及凋亡的影响[J].中国病理生理杂志,2016,32(9):1594-1598.
[13]YEUNG KT,YANG J.Epithel ialmesenchymal transition in tumor metastasis[J].Mol Oncol,2017,11(1):28-39.
[14]ILLAM SP,NARAYANANKUTTY A,MATHEW S E,et al.Epithelial mesenchymal transition in cancer progression:preventive phytochemi cals[J].Recent Pat Anticancer Drug Discov,2017,12(3):234-246.
[15]赵鹏,刘冬,张卉,等.RBMS3通过Wnt/β-catenin信号通路抑制胃癌细胞侵袭及其上皮-间质转化[J].肿瘤,2017,37(10):1032-1040.