Expression And Significance Of Has-miR-93-5p In NSCLC Cell Line

Objective:To investigate the inhibitory effect of miR-93-5p on the proliferation,invasion and metastasis of non-small cell lung cancer(NSCLC)cell line,and to analyze the relationship between miR-93-5p and downstream target genes in order to provide an experimental basis for clinical prevention and treatment of NSCLC.Methods:miR-93-5p interference method 1 using lentivirus mediated preparation,expression of miR-93-5p in lung adenocarcinoma cell line A549 and human lung cancer cell line SK-MES-1(experimental group),the other is not mediated by the two kinds of cells as the control group.MTT method was used to detect the proliferation ability of each cell line,and the activity of miR-93-5p was detected by Western blot.2 the ability of single cell clone formation of NSCLC cells was detected by using the method of flat plate colony forming assay.Flow cytometry was used to detect the cell cycle arrest and apoptosis in two NSCLC cell lines.3 migration and invasion of NSCLC cells by Transwell migration and invasion assay in two kinds of cells.Analysis of the relationship between miR-93-5p and PTEN and RB1 regulation by 4.Western blot and double luciferase gene.Results:Compared with the control group(1.82 ± 0.099)(1.90 ± 0.099)inhibition experiment group A549(0.53.±0.008)and SK-MES-1(0.65 ±0.08)cell lines was significantly increased(79%,58%,P<0.001);compared with the control group(1.90 + 0.089).decreased the activity of miR-93-5p cells and SK-MES-1 cells in the experimental group A549(71%,66%,P<0.001);two cell lines in vitro and reduced ability to form clones(63%,75%,P<0.001);lentivirus mediated miR-93-5p interference after A549 cell cycle arrest in G1 phase cells(control group G1 48%,S 40%and G2/M 12%;A549 63%cells in G1 phase,S phase and G2/M 26%11%),SK-MES-1 cell cycle arrest in G1 phase(control group G149%,S 37%and G2/M 14%;SK-MES-1 cell G1 57%,S 30%and G2/M 13%).The expression of marker protein CDK2 and cyclin D in the two stage of the cell lines of G1 decreased significantly.The apoptosis rate of two cell lines were(26.2%,18.9%,P<0.001).The expression of apoptosis related protein Bcl2 was decreased in the two cell lines,while the expression level of Caspase 3 increased.MiR-93-5p interference significantly inhibited the migration ability of A549 and SK-MES-1 cell lines in vitro(inhibition rate 61.4%,,P<0.001).MiR-93-5p interference significantly inhibited the invasive ability of A549 and SK-MES-1 cells in vitro(62%and 88.5%,respectively,P<0.001).The results of blot Western expression after miR-93-5p interference in two cell lines of PTEN and RBI protein increased significantly,and dual luciferase reporter gene showed that miR-93-5p binds directly to PTEN and RBI 3 'non encoding region,thereby inhibiting two gene translation.Conclusion:miR-93-5p can interfere with cell cycle arrest and apoptosis induced by interference;miR-93-5p can inhibit the proliferation,NSCLC cell clone formation,migration and invasion;mechanism of invasion and migration effect of NSCLC proliferation,miR-93-5p binds directly to the target gene of PTEN and RBI 3 'non encoding region,inhibition of two target genes protein expression and realization.