H19 expression is elevated in many human tumors including glioblastomas, suggesting an oncogenic role for the long noncoding RNA; yet the upregulation of H19 in glioblastomas remains unclear. Here we report that hypoxia significantly stimulated H19 expression in glioblastoma cell lines, which was related to hypoxia-inducible factors 1α (Hif-1α). Hif-1α promoted H19 expression in U87 and U251 cells. Meanwhile PTEN is an advantageous factor to affect H19 expression, through attenuating Hif-1α stability. Hif-1α also positively correlates with H19 in human glioblastoma samples depending on PTEN status. ChIP and luciferase reporter assays showed that Hif-1α induced H19 transcription through directly binding to the H19 promoter. Furthermore, Hif-1α upregulated specific protein 1 (SP1) expression in glioblastomas cells in vitro and in vivo, and SP1 also strongly interacted with the H19 promoter to promote H19 expression under hypoxia. We also showed that H19 acts as a molecular sponge that binds miR-181d, relieving inhibition of β-catenin expression. Therefore, H19 participates in hypoxia-driven migration and invasion in glioblastoma cells. In summary, our results uncover the mechanisms that stimulate H19 expression under hypoxia to promote malignant effects in glioblastomas and suggest H19 might be a promising therapeutic target.

BackgroundGlioblastoma multiforme (GBM) is the most common and aggressive form of astrocytoma among adult brain tumors. Multiple studies have shown that long non-coding RNAs (lncRNAs) play important roles in acting as molecular sponge for competing with microRNAs (miRNAs) to regulate downstream molecules in tumor progression. We previously reported that miR155 host gene (miR155HG), an lncRNA, and its derivative miR-155 promote epithelial-to-mesenchymal transition in glioma. However, the other biological functions and mechanisms of miR155HG sponging miRNAs have been unknown. Considering ANXA2 has been generally accepted as oncogene overexpressed in a vast of cancers correlated with tumorigenesis, which might be the target molecule of miR155HG sponging miRNA via bioinformatics analysis. We designed this study to explore the interaction of miR155HG and ANXA2 to reveal the malignancy of them in GBM development.MethodsThe expression of miR155HG was analyzed in three independent databases and clinical GBM specimens. Bioinformatics analysis was performed to assess the potential tumor-related functions of miR155HG. The interaction of miR155HG and miR-185 and the inhibition of ANXA2 by miR-185 were analyzed by luciferase reporter experiments, and biological effects in GBM were explored by colony formation assays, EDU cell proliferation assays, flow cytometric analysis and intracranial GBM mouse model. Changes in protein expression were analyzed using western blot. We examined the regulatory mechanism of ANXA2 on miR155HG in GBM by gene expression profiling analysis, double immunofluorescence staining, chromatin immunoprecipitation and luciferase reporter assays.ResultsWe found that miR155HG was upregulated in GBM tissues and cell lines. Bioinformatic analyses of three GBM databases showed that miR155HG expression levels were closely associated with genes involved in cell proliferation and apoptosis. Knocking down miR155HG suppressed GBM cell proliferation in vitro, induced a G1/S-phase cell cycle arrest, and increased apoptosis. We also found that miR155HG functions as a competing endogenous RNA for miR-185. Moreover, miR-185 directly targets and inhibits ANXA2, which exhibits oncogenic functions in GBM. We also found that ANXA2 promoted miR155HG expression via STAT3 phosphorylation.ConclusionOur results demonstrated that overexpressed miR155HG in GBM can sponge miR-185 to promote ANXA2 expression, and ANXA2 stimulates miR155HG level through phosphorylated STAT3 binding to the miR155HG promoter. We establish the miR155HG/miR185/ANXA2 loop as a mechanism that underlies the biological functions of miR155HG and ANXA2 in GBM and further suggest this loop may serve as a therapeutic target and/or prognostic biomarker for GBM.Electronic supplementary materialThe online version of this article (10.1186/s13046-019-1132-0) contains supplementary material, which is available to authorized users.

Enhancer of zeste homolog 2 (EZH2) is the catalytic unit of polycomb repressive complex 2 (PRC2) which epigenetically silences many genes involved in tumor-suppressive mechanisms via the trimethylation of lysine 27 of histone H3 (H3K27me3). We recently found that overexpression of EZH2 was associated with poor outcome of glioblastoma (GBM). In this study, we examined the antitumor effects of the EZH2 inhibitor GSK343 on glioma cells in vitro and in vivo. The proliferation and cell cycle of glioma cells was measured. Wound healing assay and transwell invasion assay were performed to evaluate the capacity of migration and invasion of glioma cells. Western blot, qPCR, immunoprecipitation and fluorescent staining were used to test the levels of EZH2 and associated proteins. Spheroid formation assay and clonogenic assays were conducted to assess the stemness of glioma stem cells. Finally, the effect of GSK343 was measured through a nude mice model with intracranially xenotransplanted glioma. We found that GSK343 reduced proliferation, attenuated cell motility and reversed epithelial-mesenchymal transition in U87 and LN229 glioma cells. GSK343 also suppressed the stemness of cell lines and patient derived glioma stem cells. Further, GSK343 inhibited histone H3K27 methylation and upregulated the expression of EZH2 target genes thereby regulating the levels of markers involved in epithelial-mesenchymal transition and stemness. Taken together, our results indicate that GSK343 could be a potential drug against glioblastoma.