MiR-613 Inhibits The Proliferation And Invasion Of Lung Adenocarcinoma Cell A549 By Targeting DCLK1

Objective: To investigate the effect of miR-613 on the proliferation and invasion of A549 cells,and to discover the potential target of miR-613 in lung adenocarcinoma cell line.Methods: The frozen cell line A549 donated by pathology laboratory of hebei medical university.A549 cells were transfected with miR-613 expression plasmid as miR-613 group,and blank control group(A549 cells cultured in normal culture)and negative control group(A549 cells infected with negative control virus).The lentivirus vector transfection efficiency was observed by fluorescence microscopy.Cell scratch assay and in vitro invasion assay were used to detect the proliferation and invasion of A549 cells.DIDNA and Starbase3.0 were used to predict the possible target genes of miR-613.DCLK1 wild-type 3 '-UTR luciferase plasmid and mutant plasmid were constructed and co-transfected with miR-613 mimics or miRNA of negative control in HEK293 cells.Luciferase activity was detected by luciferase assay.The effect of miR-613 on DCLK1 mRNA expression in A549 cells was detected by RT-PCR,and the effect of miR-613 on DCLK1 protein expression in A549 cells was detected by Western blot.Results:1.After 24 h of transfection,the transfection efficiency of lentivirus was observed under a fluorescence microscope.The miR-613 group and the negative control group expressed high-intensity fluorescence under a fluorescence microscope,and the infection efficiency was over 90%.2.At 12 h,the scratch distance of A549 cells in the miR-613 group(2.13±0.32)was significantly wider than that in the blank control group(1.33±0.19)and the negative control group(1.29±0.09),and the difference was statistically significant(P<0.05).At 24 hours,the scratch distance of miR-613 group(1.19±0.03)was significantly wider than that of the negative control group(0.13±0.11)and the blank control group(0.02±0.00),and the difference was statistically significant(P<0.05).The migration rate of miR-613 group was lower than that of blank control group and negative control group.3.After overexpression of miR-613,the number of invasive cells in A549 cells(27.26±1.06)was significantly lower than that in the blank control group(85.97±4.26)and the negative control group(87.43±3.17),and the difference was statistically significant(P<0.05).There was no significant difference between the blank control group and the negative control group(P>0.05).4.Bioinformatics software predicts DCLK1 3?-UTR has binding sites with miR-613,DCLK1 may be target gene of miR-613.5.Plasmid-WT transfected with miR-613 mimics showed significantly lower luciferase activity than the negative control group(P<0.01).Plasmid-Mut luciferase activity was not affected.6.After overexpression of miR-613,DCLK1 mRNA and protein levels in A549 cells decreased,with statistically significant differences(P<0.05),while no significant changes were observed in the negative control group and the blank control group(P>0.05).Conclusion:1.In this study,it was found that miR-613 significantly inhibited the proliferation and invasion of A549 cells.2.In A549 cells,miR-613 could inhibit DCLK1 mRNA and protein expression,and bioinformatics analysis showed that DCLK1 3?-UTR has binding sites with miR-613,miR-613 inhibits the proliferation of A549 cells may through DCLK1.3.Mir-613,as a tumor suppressor,regulates the proliferation and invasion of lung adenocarcinoma.This study will provide a new method for the precise targeted treatment of lung adenocarcinoma.