miR-128-3p靶向TCF4抑制黑色素瘤A375细胞增殖、侵袭和上皮-间充质转化  

miR-128-3p targeting TCF4 in melanoma A375 cells inhibits proliferation, invasion and epithelial-mesenchymal transition

作　　者：侯绍蔚 张连清[2] 席海英[2] 赵爱玲[3] 侯恒[1] HOU Shao-wei;ZHANG Lian-qing;XI Hai-ying;ZHAO Ai-ling;HOU Heng(Shanxi Datong University Medical College,Datong 037009,China;Dermatology Department of Datong Fifth People’s Hospital,Datong 037009,China;Neurology Department of Datong Fifth People’s Hospital,Datong 037009,China)

机构地区：[1]山西大同大学医学院,山西大同037009 [2]大同市第五人民医院皮肤科,山西大同037009 [3]大同市第五人民医院神经内科,山西大同037009

出　　处：《中国病理生理杂志》2019年第11期1999-2005,共7页Chinese Journal of Pathophysiology

基　　金：山西省自然科学基金资助项目(No.2013011052-4)

摘　　要：目的:探究微小RNA-128-3p(miR-128-3p)对人黑色素瘤A375细胞增殖、侵袭和上皮-间充质转化的作用及其机制。方法:用miR-128 mimic和/或转录因子4(TCF4)表达质粒pcDNA-TCF4(pc-TCF4)转染A375细胞,RT-qPCR检测miR-128-3p的表达,Western blot法检测TCF4的表达,萤光素酶报告实验证明miR-128-3p和TCF4的靶向关系;Transwell检测细胞的侵袭能力;Western blot法检测上皮标志蛋白E-cadherin和间充质标志蛋白vimentin的表达及TCF4下游靶分子cyclin D和c-Myc的蛋白表达水平。结果:过表达miR-128-3p能显著抑制TCF4的表达(P<0.01),而转染pc-TCF4能明显减弱miR-128-3p对TCF4表达的抑制作用,miR-128-3p能显著减弱TCF4野生型质粒的萤光素酶活性(P<0.01);miR-128-3p能显著抑制A375细胞增殖(P<0.01),显著减少侵袭细胞数(P<0.01);除此之外,miR-128-3p还能显著降低vimentin、cyclin D和c-Myc的表达水平(P<0.01),显著升高E-cadherin(P<0.01)的表达水平。结论:miR-128-3p能抑制黑色素瘤A375细胞的增殖、侵袭和上皮-间充质转化,作用机制与靶向TCF4及抑制其下游靶基因的表达有关。AIM: To explore the effects of microRNA-128-3 p(miR-128-3 p) on proliferation, invasion and epithelial-mesenchymal transition(EMT) of human melanoma A375 cells and the possible mechanisms. METHODS: miR-128 mimic and/or transcription factor 4(TCF4) expression plasmid pcDNA-TCF4(pc-TCF4) were transfected into the A375 cells. The expression of miR-128-3 p was detected by RT-qPCR, and the expression of TCF4 was determined by Western blot. Luciferase reporter assay was used to confirm the relationship between miR-128-3 p and TCF4. The invasion ability of A375 cells was measured by Transwell assay. The expression levels of EMT-related proteins, including E-cadherin and vimentin, and the down-stream target molecules of TCF4, including cyclin D and c-Myc, were determined by Western blot.RESULTS: Over-expression of miR-128-3 p significantly inhibited the expression of TCF4(P<0.01), while transfection of pc-TCF4 significantly reduced the inhibitory effect, and miR-128-3 p also significantly reduced luciferase activity of wild-type TCF4 plasmid(P<0.01). miR-128-3 p significantly reduced positive cells labeled by EdU(P<0.01). Meanwhile, miR-128-3 p significantly decreased the number of invasive A375 cells(P<0.01). In addition, the protein levels of vimentin, cyclin D and c-Myc(P<0.01) were down-regulated by miR-128-3 p, but the protein level of E-cadherin was up-regulated(P<0.01).CONCLUSION: miR-128-3 p inhibits proliferation, invasion and EMT of A375 cells by inhibiting the expression of TCF4 and its downstream target genes.

关 键 词：黑色素瘤 微小RNA-128-3p 细胞侵袭 上皮-间充质转化 转录因子4 

分 类 号：R730.23[医药卫生—肿瘤] R739.5[医药卫生—临床医学]