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MicroRNA-16 inhibits glioma cell growth and invasion through suppression of BCL2 and the nuclear factor-κB1/MMP9 signaling pathway.

Yang TQ, Lu XJ, Wu TF, Ding DD, Zhao ZH, Chen GL, Xie XS, Li B, Wei YX, Guo LC, Zhang Y, Huang YL, Zhou YX, Du ZW - Cancer Sci. (2014)

Bottom Line: MicroRNA-16 decreased glioma malignancy by downregulating NF-κB1 and MMP9, and led to suppressed invasiveness of human glioma cell lines SHG44, U87, and U373.Our results also indicated that upregulation of miR-16 promoted apoptosis by suppressing BCL2 expression.Finally, the upregulation of miR-16 in a nude mice model of human glioma resulted in significant suppression of glioma growth and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery and Brain and Nerve Research Laboratory, The First Affiliated Hospital of Soochow University, Suzhou, China.

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miR-16 downregulated BCL2 expression. (a) Analyzing the homology between microRNA-16 (miR-16) and BCL2 mRNA sequences. (b) MicroRNA-16 levels are inversely correlated with BCL2 expression in the same human brain glioma samples. (c) Luciferase assay revealed reduced relative luciferase activities in 293T cells stably overexpressing miR-16 following transfection of BCL2 3′-UTR using pMIR and pMIR-REPORT vectors (P < 0.01). FL, firefly luminescence; RL, Renilla luminescence. (d) Western blot analysis showed that miR-16 led to obvious downregulation of BCL2 expression.
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fig03: miR-16 downregulated BCL2 expression. (a) Analyzing the homology between microRNA-16 (miR-16) and BCL2 mRNA sequences. (b) MicroRNA-16 levels are inversely correlated with BCL2 expression in the same human brain glioma samples. (c) Luciferase assay revealed reduced relative luciferase activities in 293T cells stably overexpressing miR-16 following transfection of BCL2 3′-UTR using pMIR and pMIR-REPORT vectors (P < 0.01). FL, firefly luminescence; RL, Renilla luminescence. (d) Western blot analysis showed that miR-16 led to obvious downregulation of BCL2 expression.

Mentions: The microRNA.org database (http://www.microrna.org/microrna/get Mrna.do?gene=596&utr=5412&organism=9606) shows that BCL2 is a target of post-transcriptional repression by miR-16 (Fig.3a). Our data showed that miR-16 expression was inversely correlated with BCL2 in the same tissue samples of human glioma (Fig.3b). A dual-luciferase reporter assay system was used to validate whether miR-16 directly recognizes the 3′-UTR region of BCL2 mRNA. As shown in Figure3(c), the relative luciferase activities of the pre-miR-16 groups transfected with BCL2 3′-UTR constructs were significantly decreased, compared to those transfected with pMIR-BCL2 constructs, for the 293T cell line, implying that BCL2 is a direct target of miR-16. To examine if miR-16 facilitates apoptosis in human glioma cells, flow cytometry data indicated that increasing miR-16 expression induced early apoptosis, compared with the negative control oligonucleotide and the mock, and that the proportion of early apoptotic cells in the miR-16 treatment group was markedly increased (P < 0.01, n = 3) (Fig. S1). We further investigated the mechanism of transfection of the miR-16 mimics and inhibitors on inducing apoptosis in human glioma cells. BCL2 was detected by Western blot. Data showed that the upregulation of miR-16 led to obvious downregulation of BCL2 expression (Fig. 3d), and that miR-16 induces apoptosis in human glioma cells in vitro.


MicroRNA-16 inhibits glioma cell growth and invasion through suppression of BCL2 and the nuclear factor-κB1/MMP9 signaling pathway.

Yang TQ, Lu XJ, Wu TF, Ding DD, Zhao ZH, Chen GL, Xie XS, Li B, Wei YX, Guo LC, Zhang Y, Huang YL, Zhou YX, Du ZW - Cancer Sci. (2014)

miR-16 downregulated BCL2 expression. (a) Analyzing the homology between microRNA-16 (miR-16) and BCL2 mRNA sequences. (b) MicroRNA-16 levels are inversely correlated with BCL2 expression in the same human brain glioma samples. (c) Luciferase assay revealed reduced relative luciferase activities in 293T cells stably overexpressing miR-16 following transfection of BCL2 3′-UTR using pMIR and pMIR-REPORT vectors (P < 0.01). FL, firefly luminescence; RL, Renilla luminescence. (d) Western blot analysis showed that miR-16 led to obvious downregulation of BCL2 expression.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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fig03: miR-16 downregulated BCL2 expression. (a) Analyzing the homology between microRNA-16 (miR-16) and BCL2 mRNA sequences. (b) MicroRNA-16 levels are inversely correlated with BCL2 expression in the same human brain glioma samples. (c) Luciferase assay revealed reduced relative luciferase activities in 293T cells stably overexpressing miR-16 following transfection of BCL2 3′-UTR using pMIR and pMIR-REPORT vectors (P < 0.01). FL, firefly luminescence; RL, Renilla luminescence. (d) Western blot analysis showed that miR-16 led to obvious downregulation of BCL2 expression.
Mentions: The microRNA.org database (http://www.microrna.org/microrna/get Mrna.do?gene=596&utr=5412&organism=9606) shows that BCL2 is a target of post-transcriptional repression by miR-16 (Fig.3a). Our data showed that miR-16 expression was inversely correlated with BCL2 in the same tissue samples of human glioma (Fig.3b). A dual-luciferase reporter assay system was used to validate whether miR-16 directly recognizes the 3′-UTR region of BCL2 mRNA. As shown in Figure3(c), the relative luciferase activities of the pre-miR-16 groups transfected with BCL2 3′-UTR constructs were significantly decreased, compared to those transfected with pMIR-BCL2 constructs, for the 293T cell line, implying that BCL2 is a direct target of miR-16. To examine if miR-16 facilitates apoptosis in human glioma cells, flow cytometry data indicated that increasing miR-16 expression induced early apoptosis, compared with the negative control oligonucleotide and the mock, and that the proportion of early apoptotic cells in the miR-16 treatment group was markedly increased (P < 0.01, n = 3) (Fig. S1). We further investigated the mechanism of transfection of the miR-16 mimics and inhibitors on inducing apoptosis in human glioma cells. BCL2 was detected by Western blot. Data showed that the upregulation of miR-16 led to obvious downregulation of BCL2 expression (Fig. 3d), and that miR-16 induces apoptosis in human glioma cells in vitro.

Bottom Line: MicroRNA-16 decreased glioma malignancy by downregulating NF-κB1 and MMP9, and led to suppressed invasiveness of human glioma cell lines SHG44, U87, and U373.Our results also indicated that upregulation of miR-16 promoted apoptosis by suppressing BCL2 expression.Finally, the upregulation of miR-16 in a nude mice model of human glioma resulted in significant suppression of glioma growth and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery and Brain and Nerve Research Laboratory, The First Affiliated Hospital of Soochow University, Suzhou, China.

Show MeSH
Related in: MedlinePlus