Expression Of MicroRNA-935 In Pancreatic Cancer Tissues And Cells And Its Effects On Cell Proliferation,Migration And Apoptosis And Its Regulatory Mechanism

ObjectiveTo analyze the expression of miR-935 in pancreatic cancer tissues and cells,and investigate the impact of miR-935 on cell proliferation,cell migration and cell apoptosis of pancreatic cancer cells.To predict the direct targeting gene of miR-935,and explore the mechanism of its effect on the malignant biological behavior of pancreatic cancer cells.Methods(1)Fresh pancreatic cancer tissues and adjacent normal tissues were collected and HPDE6-C7 cells and PANC-1 cells were cultured;(2)Real-time PCRwas used to detect the expression of miR-935 in pancreatic carcinoma and pancreatic cancer cell line PANC-1;(3)Prediction of miR-935 direct targeting gene by bioinformatics database,and application of luciferase report assay to verify the rationality of target gene;(4)To transfect pancreatic cancer cell by miR-935 mimic,mir-935 inhibitor and miR-935 scramble;Cell proliferation ability was assessed by MTT assay and clongenic assay,Cell migration was evaluated using Transwell migration chambers,Cell apoptosis was quantified by means of flow cytometry,The expression of P21,P27,EMT related molecules and apoptosis related molecules in transfected cells were detected by qRT-PCR and Western blot,Silencing target gene expression in order to investigate the regulation of miR-935 and target genes.Results(1)compared with normal pancreatic tissues and cells,the expression of miR-935 was markedly up-regulated while INPP4A expression was obviously down-regulated in pancreatic cancer tissues and PANC-1 cells.(2)After transfection with miR-935 inhibitor,miR-935 was significantly suppressed,and suppression of miR-935 significantly inhibited cell proliferation,suppressed cell migration,and induced cell apoptosis of pancreatic cancer cells.(3)Suppression of miR-935 significantly resulted in a significantly increase in the expression of P27.Also,suppression of miR-935 resulted in the significantly expression changes of EMT markers;E-cadherin was significantly up-regulated,while N-cadherin,Snail and vimentin were markedly down-regulated.Besides,after suppression of miR-935,the expressions of apoptosis-related proteins were also changed;Bax were significantly up-regulated,while Bcl-2,pro-caspase-3 and active-caspase-3 were obviously down-regulated.(4)Importantly,opposite effects were obtained when miR-935 was overexpressed by transfection with miR-935 mimic.(5)INPP4A was a directly target of miR-935.(6)Silencing of INPP4A significantly counteracted the effects of miR-935 suppression on cell migration and apoptosis,as well as the expression changes of the above EMT-and apoptosis-related molecules.ConclusionsOur findings indicate that up-regulation of miR-935 may promote pancreatic cancer cell proliferation and migration,and inhibit cell apoptosis through targeting INPP4A.MiR-935 and INPP4A may serve as the potential targets in the therapy of pancreatic cancer.