Figure 3.

FoxM1 expression and its effect on cell growth in colorectal cancer (CRC) cell lines. A: Upper panel: LOVO, Cl-11, COLO-320, CX1, DLD-1, and HCT-15 cells were lysed and proteins were immunoblotted with FoxM1 and β-actin. Lower panel: The data obtained from the immunoblot analyses of FoxM1 were used to evaluate relative expression by spot densitometry. B: CRC cells were incubated with 5 and 10 μmol/L thiostrepton for 48 hours. Cell proliferation assays were performed using MTT as described in Materials and Methods. Data are reported as means ± SD of three independent experiments, with replicates of six wells for all of the doses and vehicle control for each experiment. *P < 0.05, Student's t-test. C: Thiostrepton-induced increase in subG1/Apo fraction of CRC cells. LOVO, Colo-320, DLD-1, and HCT-15 CRC cells were treated with 5 and 10 μmol/L thiostrepton for 48 hours. Thereafter, the cells were washed, fixed, and stained with propidium iodide and were analyzed for DNA content by flow cytometry as described in Materials and Methods. At least three independent experiments were performed for all of the cell lines. *P < 0.05, Student's t-test. D: Thiostrepton-mediated apoptosis in CRC cell lines.LOVO, COLO-320, DLD-1, and HCT-15 CRC cells were treated with 5 and 10 μmol/L thiostrepton for 48 hours and cells were subsequently stained with fluorescein-conjugated annexin-V and propidium iodide and analyzed by flow cytometry. Right lower (RL) quadrant denotes annexin v postive cells depicting alive apoptotic cells while right upper (RU) quadrant denotes annexin V and propidium iodide positive cells depicting dead apoptotic cells. Apoptosis is measured as the sum of RL and RU. X-axis denotes number of annexin V positive cells and y-axis denotes number of propidium iodide positive cells.