miRNA-99b inhibits the invasion ability of glioma cells by targeting mTOR / 中国药理学通报

ABSTRACT

Aim To detect the expression of miRNA-99b and mTOR in glioma tissues and to investigate the effect of miRNA-99b on the cell invasion ability of hu-man glioma cell line U251. Methods The expres-sions of miRNA-99b and mTOR mRNA in glioma tis-sues and normal brain tissues were detected by real-time PCR. After co-transfection with miRNA-99b mim-ics and wild or mutation type mTOR 3′UTR recombina-tion vector,the specific binding ability of miRNA-99b to 3′-UTR in mTOR gene was examined by luciferase gene reporter system. The expression levels of miRNA-99b,mTOR mRNA and mTOR protein in glioma cell line U251 after transfected with miRNA-99b mimics were measured by real-time PCR and Western blot,re-spectively. The cell invasion was measured by Tran-swell assay. The changes of mTOR and miRNA-99b expression levels in U251 cells after transfection with mTOR PsiCHECK were detected by real-time PCR. The correlation between the expression of miRNA-99b and prognosis was analyzed statistically. Results The expressed level of miRNA-99b was lower, and the ex-pression level of mTOR was higher in the glioma tissues than that in the normal brain tissues. The expression of miRNA-99b was up-regulated, and the expressions of mTOR mRNA and protein were down-regulated in U251 cells after transfection with miRNA-99b mimics. However, the abilities of invasion of U251 cells after transfection with miRNA-99b mimics were inhibited. The relative protein expression levels of mTOR in mTOR PsiCHECK group were significantly different from those in negative control group, but the relative expression levels of miRNA-99b had no signifi-cant difference compared with those in negative control group. Over-expression of mTOR restored the abilities of cell invasion in U251 cells, which was reduced by miRNA-99b. The Kaplan—Meier analysis and Log-Rank Test showed that there were significant differ-ences in overall survival (OS) between the miRNA-99b high-expression and low-expression group. Con-clusions The expression level of miRNA-99b is low in human glioma tissue. miRNA-99b may inhibit the cell invasion activity of glioma cell line U251 in vitro via inhibiting mTOR expression.