Two caspase-2 transcripts are expressed in rat hippocampus after global cerebral ischemia.
Caspase family genes play a critical role in the initiation and execution of programmed cell death. Programmed cell death is an important contributor to neuronal loss following cerebral ischemia. We have performed a series of experiments to investigate the role of a specific caspase, caspase-2, in the development of delayed neuronal death following transient global ischemia in the rat. A rat ischemic brain cDNA library was screened, and two splice-variants of caspase-2 mRNA were identified, caspase-2S and caspase-2L, which were highly homologous with the sequences of human and mouse caspase-2S and caspase-2L genes, respectively. RT-PCR demonstrated an increase in expression of both caspase-2S and caspase-2L mRNA at 8, 24 and 72 h of reperfusion after global ischemia. The ratio of the two PCR fragments did not change significantly throughout the time course of reperfusion. Western blot with monoclonal antibody specific to the pro-apoptotic caspase-2L splice variant revealed an increase in procaspase-2 (51 kDa) protein from 4 to 72 h following ischemia compared with sham-operated controls. Furthermore, an approximately 30-kDa cleavage product appeared at 8 h and increased with increasing duration of reperfusion. Thus, caspase-2L is both translated and activated following transient global ischemia. Finally, intraventricular administration of the caspase-2-like inhibitor (VDVAD-FMK) 30 min before induction of ischemia decreased the number of CA1 neurons staining positively for DNA damage (Klenow-labeling assay) and increased the number of healthy-appearing CA1 neurons (cresyl violet) compared with vehicle-treated controls. Taken together, the data suggest that caspase-2 induction and activation are important mediators of delayed neuronal death following transient global ischemia.
Jin
Kunlin
K
Buck Institute for Age Research, Novato, California, USA.
Nagayama
Tetsuya
T
Mao
Xiaoou
X
Kawaguchi
Kenji
K
Hickey
Robert W
RW
Greenberg
David A
DA
Simon
Roger P
RP
Graham
Steven H
SH
D017398:Alternative Splicing; D000818:Animals; D015153:Blotting, Western; D002545:Brain Ischemia / Q000201:enzymology* / Q000473:pathology; D053143:Caspase 2; D020169:Caspases / Q000096:biosynthesis* / Q000235:genetics; D004791:Enzyme Inhibitors / Q000008:administration & dosage; D006624:Hippocampus / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D007150:Immunohistochemistry; D007276:Injections, Intraventricular; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015424:Reperfusion; D020133:Reverse Transcriptase Polymerase Chain Reaction
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D012333:RNA, Messenger; D053143:Caspase 2; D020169:Caspases