Myc overexpression prevents apoptosis in melanoma cells.
(A) Detection of apoptosis by Annexin V vs propidium iodide assay as evaluated by flow cytometry after 24 h of PTX exposure. The mean percentages±SD reported in each cytogram represent early apoptotic (upper left), late apoptotic (upper right), necrotic (low right) and living (low left) cells. The cytograms reported are representative of three different experiments with similar results. (B) Mitochondrial membrane potential analyzed by JC-1 staining. ΔΦm in untreated and PTX treated cells are shown. Valinomycin (Val) was used as positive control to detect depolarized mitochondrial membrane potential. Data are average of at least three separate experiments. Bars represent standard deviation. (C) Caspase 3 activity evaluated by the Enz-check kit in control, Myc(−), Myc(+) and MYC silenced M14 cells after 24 h exposure to PTX. Data are reported as percent increase vs untreated cells and are average of three separate experiments. Bars represent standard deviation. (D) Western blot analysis of bcl-2 protein expression in the three cell lines after PTX treatment. Each lane was loaded with 40 µg of proteins from cell lysate. The experiment was repeated three times showing similar results. A representative blot is shown.