MicroRNA-145 Is Downregulated in Glial Tumors and Regulates Glioma Cell Migration by Targeting Connective Tissue Growth Factor

Abstract

Glioblastomas (GBM), the most common and aggressive type of malignant glioma, are characterized by increased invasion into the surrounding brain tissues. Despite intensive therapeutic strategies, the median survival of GBM patients has remained dismal over the last decades. In this study we examined the expression of miR-145 in glial tumors and its function in glioma cells. Using TCGA analysis and real-time PCR we found that the expression of miR-145/143 cluster was downregulated in astrocytic tumors compared to normal brain specimens and in glioma cells and glioma stem cells (GSCs) compared to normal astrocytes and neural stem cells. Moreover, the low expression of both miR-145 and miR-143 in GBM was correlated with poor patient prognosis. Transfection of glioma cells with miR-145 mimic or transduction with a lentivirus vector expressing pre-miR 145 significantly decreased the migration and invasion of glioma cells. We identified connective tissue growth factor (CTGF) as a novel target of miR-145 in glioma cells; transfection of the cells with this miRNA decreased the expression of CTGF as determined by Western blot analysis and the expression of its 3′-UTR fused to luciferase. Overexpression of a CTGF plasmid lacking the 3′-UTR and administration of recombinant CTGF protein abrogated the inhibitory effect of miR-145 on glioma cell migration. Similarly, we found that silencing of CTGF decreased the migration of glioma cells. CTGF silencing also decreased the expression of SPARC, phospho-FAK and FAK and overexpression of SPARC abrogated the inhibitory effect of CTGF silencing on cell migration. These results demonstrate that miR-145 is downregulated in glial tumors and its low expression in GBM predicts poor patient prognosis. In addition miR-145 regulates glioma cell migration by targeting CTGF which downregulates SPARC expression. Therefore, miR-145 is an attractive therapeutic target for anti-invasive treatment of astrocytic tumors.

A selective screening platform reveals unique global expression patterns of microRNAs in a cohort of human soft-tissue sarcomas

“…32 This miRNA is downregulated in glial tumors and regulates glioma cell migration by targeting connective tissue growth factor. 33 In breast cancer cells, miR-145 mediates the epithelial to mesenchymal transition by targeting Oct4. 34 MiR-145 is regulated by DNA methylation and p53 gene mutation in prostate cancer.…”

Interference on cytoplasmic polyadenylation element-binding proteins affects the invasion ability of glioma stem cells

“…Because overexpressed CPEBs remarkably promote the growth, development, and angiogenesis of tumor cells (Hansen et al, 2009;D'Ambrogio et al, 2013), we inhibited CPEBs using siRNA-3 for transfection. Characterized by a wide range of host and regular expressions of exogenous genes, lentivirus has been extensively used in gene therapy (Lee et al, 2013). In this study, transfection with a CPEB RNA-interfering lentiviral vector generated stable siRNA, and more than 70% of cells in the experiment group emitted green fluorescence, suggesting that the lentivirus carrying the specific target interference sequence worked well after transfection.…”

“…The pSRL-SIH1-H1-GFP plasmid (Tianjin Ssier Biotechnology Co., Ltd., Tianjin, China) was used as the lentiviral vector (Lee et al, 2013). Escherichia coli strain DH5α [TaKaRa Biotechnology (Dalian) Co., Ltd., Shiga, Japan] was used as competent cells (Xiao et al, 2008).…”

“…The siRNA sequence with the most evident interfering effect on CPEBs was identified by western blotting according to which DNA oligo was prepared, with the sticky ends identical to the restriction sites of the pSRL-SIH1-H1-GFP plasmid (Lee et al, 2013). The products were then transfected into E. coli competent cells, and preparation of lentiviral vector was confirmed through cloning, extraction, digestion, identification, and sequencing of the cultured plasmid as well as comparison with positive clones (Ji et al, 2007;Kang et al, 2007) (Figure 1).…”

Molecular Mechanism of MiR-136-5p Targeting NF-κB/A20 in the IL-17-Mediated Inflammatory Response after Spinal Cord Injury

“…miRNA-145, which binds to the target gene CTGF (connective tissue growth factor) 3’ non-variable coding region, inhibits the migration of malignant glial cells. Researchers have also determined that a high expression of miRNA-145 predicted good patient tolerance [26]. Moreover, one study showed that miRNA-221/222 plays a role in the initiation and invasion of glioblastoma [27].…”