Immunoblotting. Western blot analyses were conducted using primary antibodies specific for murine forms of total/cleaved caspase 9, cleaved caspase 3 (Asp 175), total caspase 8, and cleaved caspase 8 (Asp387) (Cell Signaling, Beverly, MA). Mouse b-actin antibody (Sigma) was also used to verify equal loading. Cells were washed twice with ice-cold phosphate-buffered saline (PBS), lysed in boiling lysis buffer (1% [wt/vol] SDS, 1 mM sodium orthovanadate, and 10 mM Tris, pH 7.4), boiled for 5 min, and sonicated briefly, the resultant lysate centrifuged at 12,000 3 g for 10 min at 4°C and protein concentration measured with a BCA Protein Assay Kit (ThermoFisher, Pittsburgh, PA). Total cellular proteins (40 lg) were separated on Bio-Rad precast 4-20% polyacrylamide gels (Bio-Rad) and transferred to a polyvinylidene difluoride membrane (Amersham, Arlington Heights, IL). After incubating with blocking buffer (Li-Cor, Lincoln, NE) for 1 h at 25°C, membranes were incubated with murine and/or rabbit primary antibodies (1:1000 dilution in Li-Cor blocking buffer) to immobilized proteins of interest overnight at 4°C. Blots were washed three times of 10 min with Tris-buffered saline and Tween 20 (50 mM Tris-HCl, 150 mM NaCl, and 0.1% Tween 20, pH 7.5) and then incubated with secondary IRDye 680 goat anti-rabbit and/or IRDye 800CW goat anti-mouse IgG antibodies (Li-Cor) (1:2000 dilution in Li-Cor blocking buffer) for 1 h at 25°C. After washing three times, infrared fluorescence from these two antibody conjugates were simultaneously measured using a Li-Cor Odyssey Infrared Imaging System.