Caspase‐8 activation requires functional mitochondria in camptothecin‐induced apoptosis in intestinal epithelial cells

Abstract

Both TNF‐alpha and camptothecin (CPT) activate caspase‐8 in IEC‐6 cells, but the degree of activation is lower in response to CPT compared to TNF‐alpha. The mechanism by which TNF‐alpha induces receptor‐mediated caspase‐8 activation is well established, but that of CPT which damages DNA is unknown. The present study explored the mechanisms by which caspase‐8 is activated in CPT‐induced apoptosis. IETD‐FMK, an inhibitor of caspase‐8, attenuated activation of caspase‐9 and effector caspases, blocked poly (ADP‐ribose) polymerase (PARP) cleavage, and inhibited apoptosis. Mitochondrial electron transport chain inhibitors (antimycin A, rotenone), and an inhibitor of protein synthesis (cycloheximide) prevented CPT‐induced activation of caspase‐8, ‐9 and ‐3. DEVD‐FMK, an inhibitor of caspase‐3, significantly inhibited CPT‐induced activation of caspase‐9, but had no effect on activation of caspase‐8. Inhibition of caspase‐9 by LEHD‐FMK, an inhibitor of caspase‐9, decreased caspase‐8 activation. Together, these data show that mitochondrial function is essential to the activation of caspase‐8 in the absence of receptor‐mediated signaling in CPT‐induced apoptosis. (Supported by NIDDK grant DK‐16505)