Effects Of MiR-1264 On The Biological Fun Ction Of Laryngeal Carcinoma Hep2 Cells

Objective:To study the effect of mi R-1264 on the biological function of laryngeal carcinoma Hep2 cells;Using experiment to verify whether miR-1264 can down regulate the expression of tumor suppressor gene laryngeal carcinoma related gene 1 in laryngeal carcinoma.Methods:1.The biological behavioral effects of transient transfection of miR-1264 mimic/inhibitor on the proliferation,migration and apoptosis of Hep2 cells were observed by MTT,Transwell and flow cytometry.2.Binding sites of miR-1264 and LCRG1 gene 3’UTR with luciferase reporter gene experiment software prediction.3.The expression of miR-1264 in Hep2 cells after transient miR-1264 mimic/ inhibitor was detected by Real-time polymerase chain reaction assay.4.The expression of LCRG1 protein in transiently transfected mimic/inhibitor NC control group,miR-1264 mimic/inhibitor experimental group and blank group was determined by Western blot analysis.Results:1.MTT experimental results show that compared with NC control group and blank group,Hep2 cells transiently transfected with miR-1264 mimic,the number of living cells was significantly increased,proliferation ability was significantly increased(P<0.05);and mi R-1264 inhibitor after transfection,the number of live cells significantly decreased,the proliferation ability was significantly inhibited(P<0.05).The results showed that miR-1264 reduced the proliferation ability of Hep2 cells.2.Transwell experimental results showed that compared with NC control group and blank group,Hep2 cells were transiently transfected with mi R-1264 mimic,the migration and invasion of the cells increased significantly,migration and invasion ability was significantly enhanced(P<0.05);On the contrary,after transient miR-1264 inhibitor,the number of migrating and invasive cells decreased significantly,and the migration and invasion ability decreased(P<0.05).The flow cytometry results showed that: miR-1264 inhibitor experimental group,the percentage of G1 phase and the apoptosis rate were higher than that of NC control group and blank group.The down regulation of miR-1264 expression can induce the apoptosis of Hep2 cells,blocked the cell cycle in G1 phase.3.Dual luciferase assay showed that the first binding site of miR-1264 and LCRG1 3’UTR may be potential binding sites for miR-1264.4.Real-time PCR experimental results showed that the transient transfection of miR-1264 mimic can make the Hep2 cells miR-1264 expression was significantly up-regulated(P<0.05);in contrast,transient transfection of miR-1264 inhibitor down regulate the expression of miR-1264(P<0.05),indicating transient transfection success.5.Western blot results showed that the LCRG1 protein expression in the experimental group had no significant difference than that in the mimic NC control group and the blank group after Hep2 cells transfected miR-1264 mimic(P>0.05),the expression of LCRG1 protein in the mimic NC group and the blank group also showed no significant difference(P>0.05);after transient miR-1264 inhibitor,the expression of LCRG1 protein in miR-1264 inhibitor experimental group had no significant difference than that in inhibitor NC group and blank group(P>0.05).Conclusions:1.MiR-1264 can enhance the proliferation,migration,invasion and apoptosis of Hep2 cells.2.MiR-1264 may not be involved in the down-regulation of LCRG1 protein expression.