Flu induced G0/G1 arrest in 4T1 and MDA-MB-231 cells. (A and E) 4T1 and MDA-MB-231 cells were treated with 0.1% DMSO and indicated concentrations of Flu for 24 hours. The cells were then incubated with 50 μg/mL PI and analyzed by FCM. Images shown are representative from three independent experiments. (B and F) Quantified histograms of cell cycle distributions after Flu treatment for 24 hours in 4T1 and MDA-MB-231 cells. (C and G) Effects of Flu on the expressions of proteins related in G0/G1 regulation. The expressions of CDK2, CDK4, cyclin E, cyclin D1, p21 and p27 after Flu treatment for 24 hours at indicated concentrations are shown. Molecular weight of bands are shown to the right of each image (kilo Dalton, kD). β-actin served as the internal control. Images shown are representatives from three independent experiments. (D and H) Protein expressions from (C and G) were quantified using Image J and normalized to β-actin expression. *P<0.05; **P<0.01; ***P<0.001.