Inhibition of caspase-8 attenuates neuronal death induced by limbic seizures in a cytochrome c-dependent and Smac/DIABLO-independent way.
There is increasing evidence that neuronal cell death induced by seizures occurs via extrinsic (death receptors) and intrinsic (mitochondria) pathways. Caspase-8 cleaves Bid, which releases cytochrome c, bridging the "extrinsic" and "intrinsic" pathways. Cleavage of Bid may amplify caspase-8-induced neuronal cell death following seizures. In the present study, we explored the effect of an inhibitor of caspase-8 (z-IETD-fmk) on the release of Smac/DIABLO and cytochrome c from mitochondria. Rats received intra-amygdaloid injection of kainic acid (KA) to induce seizures for 1 h. The seizures were then terminated by diazepam (30 mg/kg). The damaged and surviving neurons in hippocampus were observed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and cresyl violet staining, the expression of caspase-8, Bid, XIAP, caspase-9, cytochrome c and Smac/DIABLO were detected with immunofluorescence and Western blot. The cleavage of caspase-8 and Bid increased at 0 h, cytosolic fraction of cytochrome c and Smac/DIABLO increased by 2 h, cleavage of caspase-9 was detected by 4 h, TUNEL-positive neurons appeared at 8 h and reached a maximum at 24 h following administration of diazepam in the ipsilateral CA3 subfield of hippocampus. Inhibition of caspase-8 significantly decreased neuronal cell death, accompanied by reduction of t-Bid, cleaved caspase-9 and cytosol cytochrome c. Smac/DIABLO from mitochondria was not affected. These results suggest that seizures can lead the translocation of cytochrome c into the cytosol, and the activation of caspase-8 occurs upstream the mitochondria release of cytochrome c and Smac/DIABLO. Inhibition of caspase-8 attenuated neuronal cell death following seizures by decreasing mitochondria release of cytochrome c but not Smac/DIABLO.
Li
Tianfu
T
Institute of Neurology, Huashan Hospital, University of Fudan, 200040 Shanghai, China.
Lu
Chuanzhen
C
Xia
Zuoli
Z
Xiao
Baoguo
B
Luo
Yumin
Y
D000818:Animals; D051017:Apoptosis Regulatory Proteins; D015153:Blotting, Western; D002352:Carrier Proteins / Q000502:physiology*; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D045304:Cytochromes c / Q000502:physiology*; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007839:Functional Laterality / Q000502:physiology; D020287:In Situ Nick-End Labeling; D007276:Injections, Intraventricular; D008032:Limbic System / Q000187:drug effects / Q000502:physiology*; D008297:Male; D024101:Mitochondrial Proteins / Q000502:physiology*; D009474:Neurons / Q000187:drug effects*; D009842:Oligopeptides / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012640:Seizures / Q000201:enzymology; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D061945:Caspase Inhibitors; C489013:DIABLO protein, rat; D004791:Enzyme Inhibitors; D024101:Mitochondrial Proteins; D009842:Oligopeptides; D051636:X-Linked Inhibitor of Apoptosis Protein; C403753:benzyloxycarbonyl-isoleucyl-glutamyl-threonyl-aspartic acid fluoromethyl ketone; D045304:Cytochromes c; C505528:Casp8 protein, rat; C506435:Casp9 protein, rat; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases