Effect of silencing lncRNA MALAT1 on formation of vasculogenic mimicry of glioma
Objective To investigate the effect of downregulation of lncRNA metastasis-associated lung ad-enocarcinoma transcript 1(MALAT1)on glioma vasculogenic mimicry.Methods lncRNA-MALAT1 ex-pression levels in human glioma cells U251,U87 and normal astrocytes NHA were detected by qPCR.The Kyoto Encyclopedia of Genes and Genomes(KEGG)and Gene Ontology(GO)analyzed the signaling pathway and biological function of lncRNA-MALAT1.The silencing efficiency was detected by qPCR.The fragment sequence with the highest silencing efficiency was selected as the experimental group(LV-MALAT1 group),recombinant lentivirus infected human glioma cells U251 and U87 carrying meaningless fragments were selected as negative control group(LV-NC group),and the untreated human glioma cells U251 and U87 were selected as BLANK control group.The expression level of lncRNA-MALAT1 in the 3 groups was detected and analyzed.The VM formation ability of U251 and U87 cells in vitro was observed and analyzed by 3-dimensional cell culture.Cell proliferation and migration were evaluated by CCK-8 assay and scratch assay.Transwell assay was used to observe the migration and invasion ability of cells.The ex-pression of vasculogenic mimicry(VM)related proteins VE-cadherin,MMP2,VEGF and PI3K/AKT/FOXO1 signal transduction pathway protein and its phosphorylated protein were detected by Western blot.Results lncRNA-MALAT1 was highly expressed in glioma cells U251 and U87 compared with NHA cells(P<0.000 1).KEGG and GO enrichment analysis showed that lncRNA-MALAT1 was involved in the cellular molecular biological processes of tumor cell signaling pathway,such as PI3K/AKT pathway,cell adhesion,migration and extracellular matrix generation.The results of in vitro VM formation ability test showed that compared with U251 and U87 cells LV-NC group,the number and length of VM formation in LV-MALAT1 group were significantly reduced,with statistical significance(P<0.01).Western-blot re-sults showed that compared with U251 and U87 cells LV-NC group,the expressions of VM-related pro-teins VE-cadherin,MMP2 and VEGF in LV-MALAT1 group were significantly decreased,with statistical significance(P<0.05).The results of CCK-8 experiment showed that after 48 h of cell culture,compared with U251 and U87 cells in LV-NC group,the cell growth rate of LV-MALAT1 group was significantly decreased(P<0.01).The scratch test results showed that compared with U251 and U87 cells LV-NC group,the scratch healing area of LV-MALAT1 group was significantly reduced after 24 h and 48 h(P<0.000 1).The results of Transwell experiment showed that compared with U251 and U87 cells in LV-NC group,the number of cell migration and invasion in LV-MALAT1 group was significantly reduced(P<0.001).Compared with U251 and U87 cells in LV-NC group,the expressions of PI3K,Phospho-PI3K,AKT and Phospho-AKT in LV-MALAT1 group were significantly decreased,and the expressions of FOXO1 and Phospho-FOXO1 were significantly increased(P<0.05).Conclusion Down-regulation of lncRNA-MALAT1 inhibited the ability of glioma cells U251,U87 vasculogenic mimicry and significantly reduced their proliferation,migration and invasion,the mechanism of which may be related to the altera-tion of PI3K/AKT/FOXO1 pathway.
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