Fig 2. ZHX1 regulated the proliferation of CCA cells.

(A) The effect of knockdown or overexpression of ZHX1 on the level of mRNA was examined by real-time PCR. For ZHX1 knockdown, CCA cells were treated with 100 nM of scrambled siRNA (SCR) or ZHX1 siRNA. SCR siRNA-treated samples were used as controls. For the gain-of-function study, ZHX1-overexpressing CCA cells (ZHX1-over) were generated from HuCCT1 cells, and empty control vector-expressing (Mock) cells were used as a control. GAPDH was used as an internal control. (B) Knockdown and overexpression efficiencies were determined by western blotting. (C) ZHX1 protein levels were quantified using image J software, and β-actin was used as an internal control. SCR siRNA-treated controls in the knockdown study, and Mock cells in the gain-of-function study were used as controls. (D) Cell proliferation was measured 3 to 5 days after SCR or ZHX1 siRNA transfection. SCR siRNA-treated controls in the knockdown study and Mock cells in the gain-of-function study were used as controls to calculate relative cell proliferation. Bar graphs show the means ± SEs of three independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.