The Study Of MiRNA-22 Which Plays An Important Role In Occurrence And Development Of Medulloblastoma

Objective:To construct Lentiviral Vector with miRNA-22,upregulate the expression of it in medullobalstoma cell lines——DAOY,ONS-76,analyse the expression of protein caspase-—3,watch the growtfh trend and use Flow cytometric to detect early apoptosis of the cell line DAOY.Then clarify the the mechanism of miRNA-22 molecules in medulloblastoma pathology.Methods:1.Lentiviral vector constructionBuild Lentiviral Vector with miRNA-22.2.Cell cultureSelect two medulloblastoma cell lines which are DAOY and ONS-76,and t hen culture them with Specific medium.3.Transfection experimentsTransfer lentiviral vectors with miRNA-22 or empty vector virus into cell lines.4.Western blotAfter transfection,These cells can be divided into four groups which are DAOY-K,DAOY-Z,ONS-76-K and ons-76-Z.Extract the cell protein from above-mentioned cells,determine Casepase-3 expression levels by WB.5.MTT assayDo MTT assays with DAOY-K and DAOY-Z,then Observe the growth and proliferation situation of them.6.FCMDetect the early apoptosis of DAOY-K and DAOY-Z by flow cytometry.The results were analysed by SPSS 17.0 statistical software.Results:1.The expression leavel of caspase-3 protein:caspase-3 of DAOY-Z increased significantly greater than DAOY-K as same as the ONS-76-Z and ONS-76-K(p<0.05).2.DAOY-K growth rate was obviously greater than DAOY-Z,and the results are statistically significant.3.Use flow cytometry to detect early apoptosis of DAOY-K and DAOY-Z,we found thatearly apoptosis of DAOY-Z was significantly higher than DAOY-K,the results are alsostatistically significant.Conclusions:A preliminary study of these experiments implied that miRNA-22 plays a tumor suppressor role by combining with related targets.