Hepatocytes sensitized to tumor necrosis factor-alpha cytotoxicity undergo apoptosis through caspase-dependent and caspase-independent pathways.
Hepatocytes can be sensitized to tumor necrosis factor (TNF)-alpha toxicity by repression of NF-kappaB activation or inhibition of RNA synthesis. To determine whether both forms of sensitization lead to TNF-alpha cytotoxicity by similar mechanisms, TNF-alpha-induced cell death in RALA255-10G hepatocytes was examined following infection with an adenovirus, Ad5IkappaB, that blocks NF-kappaB activation or following cotreatment with actinomycin D (ActD). TNF-alpha treatment of Ad5IkappaB-infected cells resulted in 44% cell death within 6 h. ActD/TNF-alpha induced no death within 6 h but did lead to 37% cell death by 24 h. In both instances, cell death occurred by apoptosis and was associated with caspase activation, although caspase activation in ActD-sensitized cells was delayed. CrmA and chemical caspase inhibitors blocked Ad5IkappaB/TNF-alpha-induced cell death but did not inhibit ActD/TNF-alpha-induced apoptosis. A Fas-associated protein with death domain (FADD) dominant negative decreased Ad5IkappaB/TNF-alpha- and ActD/TNF-alpha-induced cell death by 81 and 47%, respectively. However, downstream events differed, since Ad5IkappaB/TNF-alpha but not ActD/TNF-alpha treatment caused mitochondrial cytochrome c release. These results suggest that NF-kappaB inactivation and inhibition of RNA synthesis sensitize RALA255-10G hepatocytes to TNF-alpha toxicity through distinct cell death pathways that diverge below the level of FADD. ActD-induced hepatocyte sensitization to TNF-alpha cytotoxicity occurs through a FADD-dependent, caspase-independent pathway of apoptosis.
Jones
B E
BE
Department of Medicine, Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Lo
C R
CR
Liu
H
H
Srinivasan
A
A
Streetz
K
K
Valentino
K L
KL
Czaja
M J
MJ
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D017209:Apoptosis; D002352:Carrier Proteins / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D003574:Cytochrome c Group / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D004789:Enzyme Activation; D053200:Fas-Associated Death Domain Protein; D020672:I-kappa B Proteins; D008099:Liver / Q000166:cytology / Q000502:physiology*; D016328:NF-kappa B / Q000378:metabolism; D011003:Ploidies; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000633:toxicity*
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D061945:Caspase Inhibitors; D003574:Cytochrome c Group; C505535:Fadd protein, rat; D053200:Fas-Associated Death Domain Protein; D020672:I-kappa B Proteins; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases