miR-181b过表达对胶质瘤U87细胞中MT1-MMP和TIMP3蛋白表达和细胞侵袭能力的影响

doi:10.13481/j.1671-587x.20150508

Abstract

Abstract:

Objective To explore the modulating effect of miR-181b overexpression on the expressions of membrane-type 1 metalloprotease (MT1-MMP) and tissue inhibitor of metalloprotease-3 (TIMP3)and invasive ability in glioma cells. Methods The human glioma U87 cells cultured in DMEM medium were divided into negative control (without treatment),empty vector (transfected with Lipofetamine 2000),scramble (transfected with scrambled has-miR-181b oligonucleotide) and miR-181b (transfected with has-miR-181b oligonucleotide) groups.The expression levels of miR-181b gene in the U87 cells in various groups were detected by qRT-PCR method;the expression levels of MT1-MMP and TIMP3 proteins in the U87 cells in various groups were examined by Western blotting method;the invasive ability of the U87 cells in various groups was determined by matrigel invasion assay;the inhibitory rate of chemotactic movement,migrated rate and cell adhesion rate in various groups were detected;the candidate target genes of miR-181b were predicted by Bioinformatics Database. Results The expression level of miR-181b mRNA in the U87 cells in miR-181b group was significantly higher than those in other three groups (P<0.01),resulting in miR-181b overexpression.Compared with other three groups,the expression level of MT1-MMP protein in the U87 cells in miR-181b group was decreased (P<0.01),and the miR-181b level was negatively correlated with the expression level of MT1-MMP protein (r=-0.787,P<0.05);while compared with other three groups,the expression level of TIMP3 protein in the U87 cells in miR-181b group was increased(P<0.01),and the miR-181b level was positively correlated with the expression level of TIMP3 protein (r=0.801,P<0.05).The matrigel invasion assay results showed that the number of transmembrane cells in miR-181b group was significantly lower than those in other three groups (P<0.05).The inhibitory rate of chemotactic movement was increased,the migrated rate and cell adhesion rate of U87 cells in miR-181b group were decreased compared with other three groups (P<0.05).The Bioinformatics Database prediction results revealed that one miR-181b-binding site within MT1-MMP 3' UTR and two binding sites of miR-181b within TIMP3 3' UTR were documented. Conclusion The miR-181b overexpression inhibits the invasive ability of glioma cells through downregulating the MT1-MMP expression and upregulating the TIMP3 expression;miR-181b serves as a critical regulator and may be an important therapeutic target for gliomas.

Key words: miR-181b, membrane-type 1 metalloprotease, tissue inhibitor of metalloprotease-3, glioma cell, invasion

CLC Number:

R739.4

LI Yunqian, ZHAO Liyan, SHI Yan, LIU Hui, MIAO Chunsheng, XU Wanzhen, YANG Zhiqing. Influence of overexpression of miR-181b in expressions of MT1-MMP and TIMP3 protein and invasive ability in glioma U87 cells[J].Journal of Jilin University(Medicine Edition), 2015, 41(05): 925-931.

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Influence of overexpression of miR-181b in expressions of MT1-MMP and TIMP3 protein and invasive ability in glioma U87 cells

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