MiR-32 promotes proliferation and invasion of glioma by inhibiting the target gene KLF4

Abstract

Objective To explore the molecular mechanism through which miR-32 affects the proliferation and invasion of glioma cells by targeting KLF4. Methods The expression of miR-32 in glial cell lines (HA) and glioma cell lines (U87, U373MG, U251) was detected by real-time quantitative PCR (RT-qPCR). The effects of miR-32 on U87 glioma cell lines were detected by cell proliferation (MTT), cell invasion and scratch assay. Afterwards, the target gene of miR-32 was found by the Target Scan Human 7.1 online detection system and verified by the dual luciferase reporter assay. After successful verification, the RT-qPCR and Western blot (WB) assay were used to determine the regulation of target gene KLF4 mRNA by miR-32, and the effects of target gene KLF4 on glioma cell lines were detected by MTT, cell invasion and scratch assay. Finally, rescue experiments were used to explore how miR-32 regulated the target gene KLF4 and thus affected the proliferation and invasion of glioma. Results The results of RT-qPCR showed that the expression levels of miR-32 in U87, U373MG and U251 cells were significantly higher than those in HA cells (all P<0.01). Compared with the group transfected with miRNA-NC, the miR-32 expression level, proliferation, invasion and migration ability of U87 cells were significantly up-regulated in the group transfected with miR-32 mimic (all P<0.05). The expression levels of miR-32, proliferation, invasion and migration were significantly down-regulated in the group transfected with miR-32 inhibitor (all P<0.05). The bioinformatics and dual luciferase reporter results showed that KLF4 was a target gene of miR-32, and when miR-32 was overexpressed, the cell luciferase activity was significantly decreased, and the expression of KLF4 was down-regulated. When miR-32 was silenced, the luciferase activity was significantly increased, and the expression of KLF4 was up-regulated. The expression of KLF4 in U87, U373MG and U251 cells was significantly lower than that in HA cell lines (all P<0.01). The proliferation, invasion and migration of glioma cell lines were inhibited when KLF4 was highly expressed. The rescue experiments showed that KLF4 silencing reversed the down-regulation of proliferation and invasion of glioma cell induced by miR-32 silencing. Overexpression of KLF4 reversed the promotion of proliferation and invasion of glioma cells induced by miR-32 overexpression. Conclusion miR-32 promoted proliferation and invasion of glioma cells by inhibiting the expression of its target gene KLF4. Key words: Glioma; Cell proliferation; Neoplasm invasiveness; miR-32; KLF4