Importance of FADD signaling in serum deprivation- and hypoxia-induced cardiomyocyte apoptosis.
Although cardiomyocyte (CM) apoptosis has been well described in both in vitro and in vivo models of ischemic heart disease, the intracellular pathways leading to CM death have not been fully characterized. To define the role of death receptor signaling in CM apoptosis, we constructed recombinant adenoviral vectors carrying wild-type (wt) or dominant negative (dn) forms of the death receptor adaptor protein FADD (Fas-associated death domain protein) and used these vectors to transduce rat neonatal CMs in models of hypoxia- and serum deprivation (SD)-induced apoptosis. The combination of SD and hypoxia induced rapid activation of caspase-3 and -8 as well as DNA fragmentation, reaching a plateau within 4-8 h. Adenoviral expression of FADD-dn inhibited caspase-8 activation as well as hypoxia/SD-induced apoptosis at 24 h in an moi (multiplicity of infection)-dependent manner. In contrast, adenoviral expression of FADD-wt increased apoptosis and caspase-3 activity in CMs under both normoxic and hypoxic conditions. Surprisingly, FADD-dn, as well as the specific caspase-8 inhibitor benzyloxycarbonyl-IETD-fluoromethylketone also inhibited the activation of caspase-9 and -3 in CMs subjected to hypoxia/SD. These data suggest a primary role for FADD/caspase-8 signaling that is necessary and sufficient for apoptosis of CMs subjected to hypoxia/SD.
Chao
Wei
W
Program in Cardiovascular Gene Therapy, Cardiovascular Research Center, the Department of Anesthesia and Critical Care, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, USA.
Shen
Yan
Y
Li
Ling
L
Rosenzweig
Anthony
A
D048868:Adaptor Proteins, Signal Transducing; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000502:physiology*; D002352:Carrier Proteins / Q000502:physiology*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D015687:Cell Hypoxia / Q000502:physiology*; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053200:Fas-Associated Death Domain Protein; D006321:Heart / Q000502:physiology*; D007700:Kinetics; D009206:Myocardium / Q000166:cytology* / Q000201:enzymology; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*
D048868:Adaptor Proteins, Signal Transducing; D000590:Amino Acid Chloromethyl Ketones; D002352:Carrier Proteins; D061945:Caspase Inhibitors; D016895:Culture Media, Serum-Free; D015853:Cysteine Proteinase Inhibitors; C505535:Fadd protein, rat; D053200:Fas-Associated Death Domain Protein; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C505410:Casp3 protein, rat; C505528:Casp8 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases