Protection against Fas receptor-mediated apoptosis in hepatocytes and nonparenchymal cells by a caspase-8 inhibitor in vivo: Evidence for a postmitochondrial processing of caspase-8

Lymphocytes can kill target cells including hepatocytes during various infl ammatory diseases by Fas receptor-mediated apoptosis. Caspase-8 is activate d at the receptor level, thereby initiating the processing of downstream ef fector caspases. The aim of this study was to investigate the time course o f caspase-8 activation and to evaluate the efficacy of the caspase-8 inhibi tor IETD-CHO in a model of Fas-induced apoptosis in vivo. C3Heb/FeJ mice we re treated with the anti-Fas antibody Jo-2 (0.6 mg/kg). Western blot analys is demonstrated increased cytochrome c in the cytosol (20 min), which was f ollowed by the progressive activation of caspase-3, -9 (40-120 min), and ca spase-8 (120 min). At 90 and 120 min, extensive hemorrhage was observed, in dicating damage to sinusoidal lining cells. In addition, high plasma ALT le vels (997 +/- 316 U/L) and histological evaluation indicated severe parench ymal cell injury. Parenchymal and nonparenchymal cells showed a similar inc rease in caspase-3 activity and DNA fragmentation. Treatment with IETD-CHO (10 mg/kg) attenuated the increase in caspase-3 activity and DNA fragmentat ion by 80-90% and completely prevented hemorrhage and parenchymal cell dama ge. IETD-CHO also prevented the early release of mitochondrial cytochrome c and the processing of caspase-3, -8, and -9. Thus, our data support the hy pothesis that Fas-mediated apoptosis is dependent on caspase-8 activation i n hepatocytes and nonparenchymal cells. However, the bulk of procaspase-8 i s processed late, suggesting that only a small amount of procaspase-8 may a ctually be activated at the Fas receptor. This initial signal may be amplif ied by further activation of caspase-8 by effector caspases, i.e., after mi tochondrial activation. Caspase-8 is a promising therapeutic target for inh ibition of Fas-mediated apoptosis.