Apoptotic retinal neuronal death by ischemia-reperfusion is executed by two distinct caspase family proteases.
OBJECTIVE
To evaluate possible roles of caspase-1 and caspase-3 in retinal ischemia-reperfusion injury.


METHODS
Retinal ischemia was induced in rats by increasing the intraocular pressure to 110 mm Hg for 60 minutes. Expression of caspase-1 and caspase-3 was studied at the mRNA and protein levels using immunohistochemical staining, western blot analysis, semiquantitative reverse transcription-polymerase chain reaction, and assay of the enzymatic activities. Apoptotic retinal neurons were detected by the TdT-dUTP terminal nick-end labeling (TUNEL) method. To study the roles of the caspases in retinal ischemia-reperfusion injury, an inhibitor of caspase-1, acetyl-tyrosyl-valyl-alanyl-aspart-1-al (Ac-YVAD-CHO; total dose, 10(-7) moles) and that of caspase-3, acetyl-aspartyl-glutamylvalyl-aspart-1-al (Ac-DEVD-CHO; total dose, 10(-7) moles) was injected intravitreally and the number of TUNEL-positive cells was compared with the number in sections not treated with the inhibitors.


RESULTS
In the inner nuclear layer (INL), caspase-3-like immunoreactivity was predominantly detected, whereas caspase-1-like immunoreactivity was more predominant in the outer nuclear layer (ONL). Expression of caspase-1 and -3 was upregulated at the protein and gene levels 24 hours after reperfusion. Intravitreal injection of Ac-DEVD-CHO decreased the number of TUNEL-positive cells more significantly in the INL than in the ONL (P < 0.01) at 24 hours, whereas, intravitreal injection of Ac-YVAD-CHO was more effective in decreasing the number in the ONL (P < 0.05).


CONCLUSIONS
These findings suggest a possibility that cell-type-specific activation of caspases takes place in retinal ischemia-reperfusion injury, and such caspase may induce retinal neuronal cell death.
Katai
N
N
Department of Ophthalmology, Shinshu University School of Medicine, Matsumoto, Japan.
Yoshimura
N
N
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D020170:Caspase 1 / Q000235:genetics / Q000378:metabolism*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000235:genetics / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D020287:In Situ Nick-End Labeling; D008297:Male; D009475:Neurons, Afferent / Q000201:enzymology* / Q000473:pathology; D009842:Oligopeptides / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000201:enzymology* / Q000473:pathology; D012160:Retina / Q000201:enzymology* / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015854:Up-Regulation
D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D017931:DNA Primers; D009842:Oligopeptides; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal; C096429:L 709049; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1