Figure 5. MEK2- and FAS Ligand-Dependent Caspase-8 Activation Is Necessary for Sorbitol-Induced Caspase-7 Activation.
(A–G) Immunoblotting analyses with the indicated antibodies.
(A) Caspase-9−/−, caspase-12−/−, caspase-8−/− 3T3 fibroblast, and their WT counterpart cells were treated with or without sorbitol for 6 h. 3Y1 cells were pretreated with or without caspase-8 inhibitor Ac-IETD-CHO (10 μM) for 30 min before sorbitol treatment for 6 h.
(B–C) 3Y1 cells were treated with or without sorbitol for the indicated time. Cytosol and membrane fractions of the cells were prepared (C).
(D) MEK2+/+ and MEK2−/− cells were treated with or without sorbitol for 6 h.
(E) 3Y1 cells were treated with or without sorbitol for the indicated time. Serum-free media were collected for protein precipitation.
(F–G) Incubation with or without a neutralizing FAS ligand antibody (4 μg/ml) before sorbitol treatment of 3Y1 cells (F) or MEK2−/− cells (G) for 6 h was followed by cell lysate preparation.