Figure 5

Proapoptotic role of TSA in MDA-MB231 cells. (A) Cells transfected with NF-κBLuc were exposed to 2.5 μ M TSA in medium containing 10% FBS. The histograms indicate the absolute values for ratios of Firefly/Renilla luciferase activity. The data are the means±s.e. of five independent experiments performed in triplicate. ^***P<0.001 vs MCF-7 control value; ^ΔΔΔP<0.001 vs MDA-MB231 control value. (B) The cells were exposed to 2.5 μ M TSA for 1 or 2 days (d), and western blots of total proteins were carried out. The numbers at the bottom indicate the fold variations relative to the respective values for controls. Immunoblot with anti-vinculin antibody was used for normalisation. The blots are representative of experiments repeated three times. (C) TSA-treated cells were transfected with siRNA for Met, and western blots of total proteins were carried out. Anti-Met antibody revealed both the precursor (170 kDa) and the β-chain (145 kDa). Immunoblot with anti-vinculin antibody was used for normalisation. The blots are representative of experiments repeated three times. (D) The histograms show the percentages of apoptotic and necrotic cells obtained by cytofluorimetric analysis of TSA-treated cells. □, percentage of apoptotic cells; ▪, percentage of necrotic cells. The data are the mean±s.e. of three independent experiments. ^*P<0.05, ^**P<0.005 and ^***P<0.001 vs control values at 1 and 2 days. (E) The cells were treated with TSA, and total protein extracts were prepared and used for western blots. The numbers at the bottom indicate the fold variations relative to controls. Immunoblot with anti-vinculin antibody was used for normalisation. The blots are representative of experiments repeated three times.