The Effect Of MiR-497-5p On The Proliferation Of Cervical Cancer Hela Cells By Targeting IKCa1 Gene

Objective: IKCal gene was highly expressed in cervical cancer, which affects the proliferation and apoptosis of cervical cancer Hela cells [1].In this study ,the main purpose was to predict the regulation miRNA of IKCal gene by bioinformatics software,to investigate the role of miRNA in the regulation of IKCal gene at the post transcriptional level and to detect effects of overexpression of miRNA on proliferation of cervical cancer Hela cells.Method:1.Predicting the upstream miRNA of IKCal gene by the bioinformatics software starBase V2.0 and select one of them as the research object after literature review. 2. In order to test whether the candidate miRNA can directly bind to the 3’UTR of IKCa1 mRNA ,we check the luciferase activity of the interaction between the candidate miRNA and IKCal mRNA 3 ’-UTR by the dual-luciferase reporter assay. 3.Transfection of Hela cells after identify the candidate miRNA targeting IKCa1 gene,taking PCR and Western bolt to detect the expression level of IKCal mRNA and IKCal protein,to examine the role of miRNA in the regulation of IKCal at the post transcriptional level. 4.CCK-8 method was used to detect the effect of miRNA on proliferation of Hela cells.Result: 1 .After literature review,miR-497-5p,which was selected as the research object to the follow experiment from miRNAs that the bioinformatics software starBase V2.0 predicted. 2.The results of luciferase reporter assay showed that the luciferase activity of the group of Hela cells which co-transfected with miR-497-5p mimics and IKCal-wt. 3.The results of PCR showed that the expression of IKCal mRNA in Hela cells transfected with miR-497-5p was significantly decreased,the results of Western bolt showed that IKCa1 protein was significantly decreased in Hela cells transfected with miR-497-5p mimics. 4.CCK-8 results showed that the proliferation activity of Hela cells decreased significantly after transfection with miR-497-5p mimics.Conclusion: 1.By starBase V2.0 prediction and literature review,miR-497-5p was selected as the research object;after co-transfected with IKCal-wt or IKCal-mut and miR-497-5p mimics or miR-497-5p mimics NC,taking luciferase reporter assay to identify that miR-497-5p could directly regulate IKCal gene in Hela cells. 2.The results of PCR and Western blot showed that miR-497-5p could inhibit the expression of IKCal mRNA and protein after transfected with miR-497-5p mimics,which redoublely indicated that miR-497-5p could targetting IKCal gene. 3.Taking CCK-8 test to check Hela cells proliferation after transfected with miR-497-5p mimics,the result showed that miR-497-5p could inhibit the proliferation of Hela cells. 4.This study identified that miR-497-5p can targeting IKCal, it showed that the down-regulation of miR-497-5p expression may be a mechanism of high expression of IKCal gene in cervical carcinomas; and miR-497-5p can inhibit the proliferation of cervical cancer Hela cells maybe related to it restrains the expression of IKCal gene,which could help to explore the possible molecular mechanism of cervical cancer.