RESUMO
BACKGROUND: Abnormal expression of lncRNA is involved in a diversity of diseases and plays a vital role in targeted therapy. However, few studies have been conducted on lncRNA PART1 in glioma. We aimed to investigate the function and the potential regulatory mechanism of lncRNA PART1/miR-374b/SALL1 axis in glioma. METHODS: qRT-PCR and western blotting detected genes and proteins expression. Dual-luciferase reporter assay was performed to examine the binding relationship of lncRNA PART1, miR-374b, and SALL1. MTT assay and clone formation assay were performed to detect the cell viability and proliferation. Transwell assay detected glioma cell migration. In vivo tumor development experiments detected changes in tumor size, volume, and weight of the tumor after overexpression of lncRNA PART1. Immunohistochemistry was used to detect ki-67, E-cadherin, and N-cadherin expression. RESULTS: The expression of lncRNA PART1 and SALL1 were down-regulated and miR-374b was up-regulated in different glioma cell lines. Overexpression of lncRNA PART1 inhibited glioma cell proliferation, migration, and epithelial mesenchymal transition (EMT). LncRNA PART1 targeted miR-374b to promote SALL1 expression. The knockdown of miR-374b inhibited glioma cell proliferation and migration and EMT by SALL1. What's more, overexpression of miR-374b or knockdown of SALL1 reversed the inhibitory effect of lncRNA PART1 on the proliferation, migration, and EMT of glioma cells. Furthermore, overexpression of lncRNA PART1 inhibited glioma growth in vivo. CONCLUSION: LncRNA PART1 inhibited glioma proliferation and migration via miR-374b/SALL1 axis. These results might provide new insights for comprehending the complex lncRNA-miRNA network in gliomas.
Assuntos
Glioma , MicroRNAs , RNA Longo não Codificante , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/patologia , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
BACKGROUND: Glioma is the most common cancer in the central nervous system. Previous studies have revealed that the miR-376 family is crucial in tumour development; however, its detailed mechanism in glioma is not clear. METHODS: Cellular mRNA or protein levels of miR-376a, SIRT1, VEGF and YAP1 were detected via qRT-PCR or Western blotting. We analysed the proliferation, angiogenesis and migration abilities of glioma cell lines using colony formation, tube formation and Transwell assays. A luciferase assay was performed to determine whether miR-376a could recognize SIRT1 mRNA. Xenograft experiments were performed to analyse the tumorigenesis capacity of glioma cell lines in nude mice. The angiogenesis marker CD31 in xenograft tumours was detected via immunohistochemistry (IHC). RESULTS: miR-376a expression was lower in glioma cells than in normal astrocytes. miR-376a mimic inhibited SIRT1, YAP1, and VEGF expression and suppressed the proliferation, migration and angiogenesis abilities of the glioma cell lines LN229 and A172, whereas miR-376a inhibitor exerted the opposite functions. In a luciferase assay, miR-376a inhibited the luciferase activity of WT-SIRT1. SIRT1 overexpression upregulated YAP1 and VEGF in glioma cells and promoted proliferation, migration and angiogenesis. Xenografts with ectopic miR-376a expression exhibited lower volumes and weights and a slower growth curve. Overexpression of miR-376a inhibited YAP1/VEGF signalling and angiogenesis by inhibiting SIRT1 in xenograft tissues. CONCLUSION: miR-376a directly targets and inhibits SIRT1 in glioma cells. Downregulation of SIRT1 resulted in decreased YAP1 and VEGF signalling, which led to suppression of glioma cell proliferation, migration and angiogenesis.
RESUMO
Glioma is characterized by high morbidity, high mortality and poor prognosis. Recent studies exhibited that lncRNA CCAT2 is overexpressed in glioma and promotes glioma progression, but the specific molecular biological mechanism remains to be determined. We performed qRT-PCR to evaluate the expression of related genes, Western blotting analysis to measure protein levels, colony formation assay to detect the proliferative ability of glioma cells, flow cytometry to measure cell apoptosis, bioinformatics analysis and dual luciferase assay to verify the binding sites and the targeted regulatory relationship in A172 and U251 cell lines and tube formation assay to determine endothelial angiogenesis. LncRNA CCAT2 and VEGFA were highly expressed, while miR-424 was expressed at low levels in NHA cells. Furthermore, knockdown of lncRNA CCAT2 decreased cell proliferation, increased cell apoptosis and inhibited endothelial angiogenesis in glioma. Moreover, lncRNA CCAT2 shared a complementary sequence with miR-424 which in turn directly bound to the 3'-UTR of VEGFA. Further investigation indicated that lncRNA CCAT2 promoted cell proliferation and endothelial angiogenesis by inducing the PI3K/AKT signalling pathway in glioma. The oncogenic lncRNA CCAT2 is highly associated with the development of glioma and exerts its function by upregulating VEGFA via miR-424.
Assuntos
Regulação Neoplásica da Expressão Gênica/genética , Glioma/metabolismo , MicroRNAs/metabolismo , Neovascularização Patológica/metabolismo , RNA Longo não Codificante/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Regiões 3' não Traduzidas , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Células Endoteliais/metabolismo , Técnicas de Silenciamento de Genes , Glioma/genética , Humanos , MicroRNAs/genética , Neovascularização Patológica/genética , Oncogenes/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/genética , RNA Interferente Pequeno , Transdução de Sinais/genética , Regulação para Cima , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
BACKGROUND: Glioma is a common malignant tumor of the human central nervous system, and the pathological characteristics include invasive growth, angiogenesis, and so on. Ectopic expression of miR-503 works as a critical factor in cancer cell proliferation, migration, and capillary-like tube formation. The potential mechanisms of miR-503 in angiogenesis of glioma cells are still not reported. METHODS: The expression levels of miR-503, LRIG2, and VEGFA mRNA and protein were performed by quantitative reverse transcription-PCR or Western blot assay. Dual-Luciferase reporter gene assay was used to determine the interaction between miR-503 and LRIG2. The concentration of VEGFA was measured using the ELISA method. The cell proliferation, migration, and angiogenesis of cocultured HCMEC/D3 cells were analyzed by MTT assay, transwell detection, and tube formation assay, respectively. RESULTS: The expression levels of LRIG2 and VEGFA were reduced in glioma cells with miR-503 overexpression and enhanced with miR-503 inhibition. Moreover, cell proliferation, migration, and angiogenesis of cocultured HCMEC/D3 cells were alleviated with miR-503 mimics transfection. VEGFA and miR-503 inhibitor promoted cell proliferation, cell migration, and angiogenesis. Luciferase reporter gene assay revealed that miR-503 could directly target LRIG2. Furthermore, knockdown of LRIG2 or addition of VEGF inhibitor bevacizumab could abrogate the effect of miR-503 inhibitor on VEGFA expression, as well as the promotion of cell proliferation, migration, and angiogenesis. CONCLUSION: MiR-503 mediated LRIG2 suppression and regulated the expression of VEGFA, thereby reducing cell proliferation, migration, and angiogenesis of glioma cells. These results provide new insight into the action mechanism of miR-503-modulated signaling pathway in angiogenesis of glioma cells.
