miR-218-5p Inhibits the Development of Colon Cancer by Regulating LAYN / 昆明医科大学学报

ABSTRACT

Objective To investigate the expression level of miR-218-5p and LAYN in colon cancer and the effects of their regulatory relationship on the progress of colon cancer.Methods RT-qPCR was used to detect the expression of miR-218-5p in colon cancer tissues,adjacent tissues,and cell lines.Western blot was used to detect the expression level of LAYN protein in colon cancer tissues,adjacent tissues,and cell lines.The in situ hybridization(ISH)was used to detect the expression of miR-218-5p in colon cancer and adjacent tissues.Immunohistochemical(IHC)experiment was used to detect the distribution and expression of LAYN protein in colon cancer tissues and adjacent tissues.The dual luciferase experiment was used to detect the binding relationship between miR-218-5p and 3'-rn-translation region(UTR)of LAYN.CCK-8 was used to detect the cell proliferation.Transwell was used to detect the invasion ability of cells.The flow cytometry was used to detect the apoptosis level of cells.Western blot was used to detect the expression level of cleaved Caspase3 in cells.Results The expression of miR-218-5p decreased in colon cancer tissues and colon cancer cell lines,with the lowest expression in HT-29 cells(P<0.0001),and the expression of LAYN increased and the highest expression in HT-29 cells(P<0.05).The double luciferase reporter gene experiment(P<0.01)combined with Western blot experiment(P<0.01)showed that miR-218-5p could bind to the 3'UTR region of LAYN and inhibit its protein expression.In cell experiments,overexpression of miR-218-5p inhibited cell proliferation(P<0.05),invasion(P<0.001),and increased apoptosis(P<0.01),and the expression of cleaved Caspase3 protein(P<0.01).Overexpression of LAYN promoted cell proliferation(P<0.05),invasion(P<0.001),and decreased apoptosis(P<0.0001)and the expression of cleaved Caspase3 protein(P<0.01),while overexpression of miR-218-5p and LAYN partially reversed the above changes.Conclusion The expression level of miR-218-5p is decreased in colon cancer tissues.Cell experiments show that miR-218-5p inhibits cell invasion and proliferation and promotes cell apoptosis through targeted regulation of LAYN.