Western blotting was conducted as previously described.19 (link),20 (link) Briefly, the total proteins were extracted from the cells with RIPA lysis buffer, separated in 8–12% SDS-PAGE gels, and transferred onto PVDF (polyvinylidene fluoride) membranes (Millipore, Billerica, MA, USA). After blocking with 5% nonfat milk at room temperature for 1 hour, the membranes were incubated with the relevant primary antibodies followed by an appropriate secondary antibody. The primary antibodies in the current study included anti-UBE2O (1:800, Abcam), anti-E-cadherin, anti-Vimentin, anti-N-cadherin, anti-Snail, anti-Zeb1 and anti-Zeb2 (1:800, Cell Signaling Technology, Danvers, MA, USA). Anti-β-Actin (1:1000, Beyotime, Shanghai, China) was used as a loading control. The blots were visualized using ECL reagents (Pierce, Rockford, IL, USA) and a chemiluminescence imaging system (Bio-Rad, Richmond, CA, USA).