miR-566 regulated the β-catenin and HIF-1α pathway through VHL. (A) U87 and LN229 cells were treated with lenti-AS-566 or VHL plasmid, and TOPFLASH activity was measured. The data are shown as the mean ± SD of 3 independent experiments. *, P < 0.05, compared with vector control. (B, D) Cytoplasmic and nuclear proteins were prepared from lenti-AS-566 or VHL plasmid treated U87 and LN229 cells with or without the addition of CoCl2. Cytoplasmic and nuclear β-catenin and HIF-1α expression levels were detected. β-actin and Histone H2A.X were used as loading controls for cytoplasmic and nuclear proteins, respectively. (C, E) U87 cells were infected with lenti-AS-566 or VHL plasmid in the presence of LiCl, CoCl2 or vehicle for 24 h. Confocal images show the translocation of β-catenin and HIF-1α in U87 cells. DAPI was used to stain the nuclei. Bar, 10 μm.