MicroRNA-200c Promotes Tumor Cell Proliferation And Migration By Directly Targeting Dachshund Family Transcription Factor 1 By The Wnt/?-catenin Signaling Pathway In Nasopharyngeal Carcinoma

Objective The aim is to investigate the effect of microRNA-200c(miR-200c)on proliferation and migration of nasopharyngeal carcinoma cells,and to explore the target genes of miR-200 c downstream and the target regulation relationship.Thus elucidating the specific molecular regulation mechanism of miR-200 c on the biological function of nasopharyngeal carcinoma.Methods 1.Detect expression of miR-200 c in nasopharyngeal carcinoma tissues and cell lines by RT-qPCR.2.MiR-200c-inhibitor(miR-200c-in)was transfected into nasopharyngeal carcinoma CNE2 and SUNE1 cells.The transfection efficiency was observed and the stable expression cell lines were selected.3.The effect of miR-200 c level on the proliferation ability of tumor cells was detected by MTT and plate colony formation assays.4.The effect of miR-200 c on the migration ability of tumor cells was detected by transwell and wound-healing assays.5.Verify the downstream target of miR-200 c through Luciferase reporter assay The expression of DACH mRNA and protein level were detected by RT-qPCR and Western blotting,respectively.6.Expression of core protein in Wnt/?-catenin signaling pathway were tested by Western blotting.Result.1.The average expression level of miR-200 c in nasopharyngeal carcinoma tissues and cell lines: we detected 36 pairs of nasopharyngeal carcinoma tissues and adjacent normal tissues,the result indicated that the expression of miR-200 c in cancer tissues was significantly higher than that in normal tissues.Furthermore,compared with normal nasopharyngeal cell line NP69,the expression of miR-200 c was significantly increased in nasopharyngeal carcinoma cell lines,such as CNE2?HNE1?SUNE1.2.The cell growth rate slowed down,the number of flat clone formation decreased and the cell migration ability descended after transfection of miR-200c-in,compared with control group(transfected with miR-200c-NC).3.Luciferase reporter assay showed that DACH1 was confirmed to be the downstream target of miR-200 c.4.The results of RT-qPCR and western blotting showed that the expression of DACH1 mRNA and protein level were both increased when the expression of miR-200 c was down regulated.5.The changes of core protein of Wnt/?-catenin signaling pathway were detected by western blotting.The results showed that compared with the negative control group,the protein expression of c-Myc,cyclin D1,DACH1,?-catenin and GSK3? decreased after transfected with miR-200c-in.Conclusions Through this experiment research proves that miR-200 c is highly expressed in nasopharyngeal carcinoma tissues and cell lines,it can promote the proliferation and migration of tumor cells as a cancer gene.Inhibiting the expression of miR-200 c can decrease the proliferation and migration ability of tumor cells.DACH1 is the downstream target gene of miR-200 c.MiR-200 c inhibits the expression of DACH1 through the Wnt/?-catenin signaling pathway,thereby promoting the proliferation and migration of nasopharyngeal carcinoma cells.