Induction of caspase-3-like protease may mediate delayed neuronal death in the hippocampus after transient cerebral ischemia.
Delayed neuronal death after transient cerebral ischemia may be mediated, in part, by the induction of apoptosis-regulatory gene products. Caspase-3 is a newly characterized mammalian cysteine protease that promotes cell death during brain development, in neuronal cultures, and in other cell types under many different conditions. To determine whether caspase-3 serves to regulate neuronal death after cerebral ischemia, we have (1) cloned a cDNA encoding the rat brain caspase-3; (2) examined caspase-3 mRNA and protein expression in the brain using in situ hybridization, Northern and Western blot analyses, and double-labeled immunohistochemistry; (3) determined caspase-3-like activity in brain cell extracts; and (4) studied the effect of caspase-3 inhibition on cell survival and DNA fragmentation in the hippocampus in a rat model of transient global ischemia. At 8-72 hr after ischemia, caspase-3 mRNA and protein were induced in the hippocampus and caudate-putamen (CPu), accompanied by increased caspase-3-like protease activity. In the hippocampus, caspase-3 mRNA and protein were predominantly increased in degenerating CA1 pyramidal neurons. Proteolytic activation of the caspase-3 precursor was detected in hippocampus and CPu but not in cortex at 4-72 hr after ischemia. Double-label experiments detected DNA fragmentation in the majority of CA1 neurons and selective CPu neurons that overexpressed caspase-3. Furthermore, ventricular infusion of Z-DEVD-FMK, a caspase-3 inhibitor, decreased caspase-3 activity in the hippocampus and significantly reduced cell death and DNA fragmentation in the CA1 sector up to 7 d after ischemia. These data strongly suggest that caspase-3 activity contributes to delayed neuronal death after transient ischemia.
Chen
J
J
Department of Neurology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, USA.
Nagayama
T
T
Jin
K
K
Stetler
R A
RA
Zhu
R L
RL
Graham
S H
SH
Simon
R P
RP
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D001710:Biotin; D020170:Caspase 1; D053148:Caspase 3; D020169:Caspases; D002540:Cerebral Cortex / Q000201:enzymology; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053938:DNA Fragmentation; D018076:DNA, Complementary; D003856:Deoxyuracil Nucleotides; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D006624:Hippocampus / Q000166:cytology* / Q000378:metabolism; D002546:Ischemic Attack, Transient / Q000201:enzymology*; D008297:Male; D008969:Molecular Sequence Data; D017072:Neostriatum / Q000201:enzymology; D009474:Neurons / Q000166:cytology / Q000201:enzymology*; D009842:Oligopeptides / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D013194:Staining and Labeling; D013379:Substrate Specificity
D015853:Cysteine Proteinase Inhibitors; D018076:DNA, Complementary; D003856:Deoxyuracil Nucleotides; D004792:Enzyme Precursors; D009842:Oligopeptides; D012333:RNA, Messenger; C108637:aspartyl-glutamyl-valyl-aspartal; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; C096429:L 709049; D001710:Biotin; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D020170:Caspase 1