Effect Of MiR-568 Targeting CDK4 On Growth And Proliferation Of Cervical Cancer Hela Cells

Objective: To investigate the relationship between expressions of miR-568 and cyclin-dependent kinase 4(CDK4), analyze whether CDK4 is the target gene of miR-568, and to elucidate the molecular mechanism of Cyclin D1 expression by miR-568 targeting CDK4 in cervical cancer Hela cells.Methods: First of all, 20 miRNAs are foud to be the potential targets of CDK4 by bioinformatics method, and the miR-568 is the best binding target of CDK4 by further calculation and analysis. In Hela-miR-568 mimics, Hela-miR-Scramble, Hela-miR-568 inhibitors, Hela, miR-568 expression was detected by q RT-PCR and CDK4, Cyclin D1 were detected by Western blot. To analyze whether miR-568 was combined with CDK4; the cell counting method to draw the growth curve, cell doubling time, and flow cytometry to detect the cell cycle distribution in the four cell lines.Results:(1) 20 miRNAs were found to bind with CDK4 by bioinformatics methods; and among these miRNAs, miR-568 was the target miRNA with the best specificity and the strongest stability for CDK4.(2) The growth rate of Hela-miR-568 mimics was significantly slower than that of Hela and Hela-miR-Scramble, and the growth rate of Hela-miR-568 inhibitors was significantly increased compared with Hela and Hela-miR-Scramble. The doubling time of Hela, Hela-miR-Scramble, Hela-miR-568 inhibitors and Hela-miR-568 mimics cells were 16 h, 16 h, 10 h, 26 h, respectively.(3) miR-568 had down expression in cell line Hela-miR-568 inhibitors, while CDK4 and Cyclin D1 had increased expression; miR-568 had increased expression in cell line Hela-miR-568 mimics, while CDK4 and Cyclin D1 had down-regulated expression; there was no significant difference in expression level of miR-568, CDK4 and Cyclin D1 in Hela cell line compare to Hela-miR-Scramble cell line.(4) Ratio of cells with G1, S phase in Hela-miR-568 inhibitors were higher than that of Hela-miR-Scramble. While ratio of cells with G2/M phase was lower than Hela-miR-Scramble, the difference were significant(P<0.05); Ratio of cells with G1, S phase in Hela-miR-568 mimics were lower than Hela-miR-Scramble. While ratio of cells with G2/M phase was higherer than Hela-miR-Scramble, the difference were significant(P<0.05).Conclusions:(1) CDK4 may be the target gene of miRNA-568.(2) miRNA-568 can targetedly bind to CDK4 and down-regulate expressions of Cyclin D1 and inhibits growth and proliferation of cervical cancer cells...