The six-well plated cultured cells were incubated with the red caspase-1 detection probe, 660-YVAD-FMK (FLICA) following the manufacturer’s instructions (ImmunoChemistry Technologies, USA). Cells were washed with 4°C PBS before adding the 500 μl Annexin-binding buffer containing 2.5 μl of Annexin-V fluorescein isothiocyanate (FITC) and Hoechst (5 μg/ml) for 30 min of incubation. Finally, we added 1 μl propidium iodide (PI) solution for a further 5-min incubation in the dark. The stained samples were imaged live in an In Cell Analyzer 1000 high-content analysis system (GE Healthcare Life Sciences) equipped with a CCD camera and a 10×/0.45 NA objective. The 51008 polychroic mirror set was used in conjunction with the following excitation (×) and emission (m) filter combinations: 405/20×, 535/50m for Hoechst 33342, 475/20×, 535/50 m for Annexin-V FITC, 475/20×, 620/60 m for PI and 620/60×, 700/75 m for FLICA 660-YVAD-FMK. For the analysis, we defined a segmentation based on nuclei staining with Hoechst 33342. Then we established four groups in order to classify five populations: (1) Live cells were defined as negative cells for PI and normal staining with Hoechst
