Profiling and Bioinformatics Analyses of Differential Circular RNA Expression in Glioblastoma Multiforme Cells Under Hypoxia

Abstract

The hypoxia microenvironment is highly associated with GBM’s malignant phenotypes. CircRNAs were reported involved in GBM’s biological characteristics and regulated by HIF-1α. However, the differential expression profile and role of circRNAs in GBM cells under hypoxia are still unclear. The expression profiles of circRNAs in LN229 and T98G under hypoxia were explored via circRNA sequencing analysis. Those circRNAs significantly dysregulated both in LN229 and T98G and could be found in circBase were selected and validated by qRT-PCR, RNase R digestion reaction, and Sanger sequencing. Normal cell line and fresh GBM tissues were also used for qRT-PCR validation. The roles of differentially expressed circRNAs were evaluated by bioinformatics analyses. There were 672 dysregulated circRNAs in LN229 and 698 dysregulated circRNAs in T98G. GO analysis indicated that the alteration of circRNA expression related to GBM cell’s biogenesis and metabolism. KEGG analysis demonstrated that TGF-β signaling pathway, HIF-1 signaling pathway, and metabolism-related signaling pathway were closely associated with differentially expressed circRNAs under hypoxia. These results were confirmed by GSEA analysis. The 6 selected and dysregulated circRNAs both in LN229 and T98G including hsa_circ_0000745, hsa_circ_0020093, hsa_circ_0020094, hsa_circ_0000943, hsa_circ_0004874, and hsa_circ_0002359 were validated by qRT-PCR. Inhibition of hsa_circ_0000745 inhibited GBM cell’s proliferation, migration, and invasion. HIF-1α centered circRNA-miRNA-mRNA networks analysis showed that the 6 validated circRNAs could cross-talk with 11 related miRNAs. The circRNA expressions are dysregulated in GBM cell under hypoxia. The 6 validated circRNAs could participate in GBM’s development and progression when hypoxia occurs. They might be the candidates for prognostic markers and adjuvant therapeutics of GBM in the future.