Figure 2.

Hypoxia-induced downregulation of UHRF1 contributes to the occurrence of EMT. (a) Western blot analysis for UHRF1, vimentin and HIF-1α in HepG2 cells grown under normoxia and hypoxia for 24 h. (b) qRT-PCR analysis for the mRNA expression level of UHRF1 in HepG2 cells grown under normoxia and hypoxia for 24 h. (c) Western blot analysis for UHRF1, vimentin and HIF-1α in Hep3B cells grown under normoxia and hypoxia for 24 h. (d) qRT-PCR analysis for the mRNA expression level of UHRF1 in Hep3B cells grown under normoxia and hypoxia for 24 h. (e) Immunocytochemistry for HIF-1α and UHRF1 in a HepG2 spheroid. (f) Western blot analysis for UHRF1, vimentin and HIF-1α in UHRF1-overexpressing HepG2 cells. (g and h) Effects of UHRF1 overexpression on the migratory and invasive properties of HepG2 cells grown under hypoxia for 24 h. The migration and invasion assay were performed using the Transwell chamber. (I and J) Effects of UHRF1 deficiency on the migratory and invasive properties of HepG2 cells grown under hypoxia for 24 h. The migration and invasion assay were performed using the Transwell chamber. (k) Effects of UHRF1 deficiency on tumorigenic capacity of HepG2 cells in vivo. shCont- and shUHRF1-HepG2 cells were subcutaneously injected to the right flank of athymic BALB/c female nude mice (n = 5 per group). After 5 weeks, tumor mass was photographed under a light microscope (up). The average of tumor volumes was measured at 1-week intervals (down). (l) Quantification of lung metastasis determined by counting the number of foci on the lung surface after tail vein injection of shCont- and shUHRF1-HepG2 cells (n = 4 per group). (m) H&E staining of lung sections after tail vein injection of shCont- and shUHRF1-HepG2 cells. β-actin was used as a loading control. Results from three independent experiments are expressed as means ± SEMs. (*P < 0.05, **P < 0.01).