Effects of FAS siRNA on drug-induced FAS expression in HMC-1 cells and responsiveness against demethylating agents. (A) HMC-1.2 cells were kept in control medium (untransfected cells) or were transfected with siRNA against FAS (200nM) using lipofectin. Cells were then incubated in control medium (black-lined open histograms), 5-azacytidine, or decitabine (each 5μM, gray histograms) at 37°C for 30 hours. Thereafter, CD95 expression was analyzed by flow cytometry. Expression of FAS was compared with staining reactions produced by isotype-matched control antibodies (gray-lined open histograms). (B-C) HMC-1.2 cells were kept in control medium (untransfected cells) or were transfected with siRNA against luciferase (200nM) or against FAS (200nM) using lipofectin. After 1 hour, cells were incubated in control medium, 5-azacytidine, or decitabine (each 5μM) at 37°C for 30 hours. After incubation, annexin V/PI staining (B) or active caspase-3 staining (C) was performed by flow cytometry. Results show the percentage of annexin V/PI-positive cells determined by flow cytometry (B), and the percentage of active caspase-3–positive cells (C).