Overexpression of catalase in the mitochondrial or cytosolic compartment increases sensitivity of HepG2 cells to tumor necrosis factor-α-induced apoptosis

The sensitivity of HepG2 cells overexpressing catalase in either the cytosolic or mitochondrial compartment to tumor necrosis factor-α (TNF-α) and cycloheximide was studied. Cells overexpressing catalase in the cytosol (C33 cells) and especially in mitochondria (mC5 cells) were more sensitive to TNF-α-induced apoptosis than were control cells (Hp cells). The activities of caspase-3 and -8 were increased by TNF-α, with the highest activities found in mC5 cells. Sodium azide, an inhibitor of catalase, reduced the increased sensitivity of mC5 and C33 cells to TNF-α to the level of toxicity found with control Hp cells. Azide also decreased the elevated caspase-3 activity of mC5 cells. A pan-caspase inhibitor prevented the TNF-α-induced apoptosis and toxicity produced by catalase overexpression. Addition of H₂O₂ prevented TNF-α-induced apoptosis and caspase activation, an effect prevented by simultaneous addition of catalase. TNF-α plus cycloheximide increased ATP levels, with higher levels in C33 and mC5 cells compared with Hp cells. TNF-α did not produce apoptosis in mC5 cells maintained in a low energy state. TNF-α signaling was not altered by the overexpression of catalase, as activation of nuclear factor κB and AP-1 by TNF-α was similar in the three cell lines. These results suggest that catalase, overexpressed in the cytosolic or especially the mitochondrial compartment, potentiates TNF-α-induced apoptosis and activation of caspases by removal of H₂O₂.