Figure 1

Simvastatin induces p21 and p27-dependent G0/G1 cell cycle arrest in HCC cell lines. Simvastatin suppressed cell growth in HCC cells. HepG2 and Hep3B cells were treated with various concentrations of simvastatin for 24 and 48 h. (a) Cell growth inhibition was measured by CCK-8 assay. (b) Cell death rates were determined by viable cell counting. These data are presented as percentages of vehicle-treated cells (*un-treated versus treated of HepG2 or Hep3B cells for 24 h or 48 h). (c and d) Simvastatin-induced HCC cell G0/G1 phase arrest. HepG2 and Hep3B cells were treated with simvastatin (0, 5, 10 or 20 μg/ml) for 48 h, and then cell cycle distributions were analyzed by PI staining and flow cytometry. (e) Simvastatin-induced G0/G1 phase-related protein expression in HCC cells. The cells were treated with simvastatin (0, 5, 10 or 20 μg/ml) for 24 h, and then the cell lysates were harvested for analysis of the expression of the cell cycle-related proteins p21, p27, cyclin D1, cyclin E1 and β-actin by immunoblotting. (f and g) Simvastatin-induced p21- and p27-dependent G0/G1 cell cycle arrest in HCC cells. HepG2 cells were transfected with p21, p27 or control siRNA for 24 h and then treated with 10 μg/ml simvastatin for 48 h. The cells were then harvested, and their DNA content and protein expression were analyzed by flow cytometry and immunoblotting using p21, p27 and β-actin antibodies. Data are expressed as the mean±S.E.M. of three independent experiments. Statistically significant differences between the un-treated and treated groups are indicated. *P<0.05, **P<0.01, ***P<0.001