Effects of FRNK overexpression on Gem-induced apoptosis in Panc-1 cells. Parental Panc-1 cells and empty vector-transfected (Vector) and pcDNA3.1-FRNK plasmid-transfected (Pool 1) cells were treated with or without 10 μM Gem for 72 h and subjected to cell apoptosis analysis by Hoechst staining (the arrow indicates the apoptotic cells) (A), flow cytometry analysis of Annexin-V labeling (B) and western blot analysis of cleaved caspase-3 protein expression (C). Bars represent the mean of three independent experiments ± SE. *, P < 0.05, vs. parental cells without Gem treatment; #, P < 0.05, vs. parental cells or vector cells with Gem treatment. D, Western blot analysis was used to detect the expression of Bad, p-Bad (pS136), p-Bad (pS112), Bcl-2, Bax and survivin in parental cells and vector and pool 1 cells. The membranes were probed with anti-β-actin antibody to ensure even loading of proteins in each lane.