Apoptosis and caspases after ischemia-reperfusion injury in rat retina

PURPOSE. Extensive cell loss in the retinal ganglion cell layer (RGCL) and the inner nuclear layer (INL) was noted in a rat model of retinal ischemia- reperfusion injury by transient elevated intraocular pressure (IOP). The po ssible involvement of apoptosis and caspases was examined in this model of neuronal loss. METHODS. Transient elevated IOP was induced in albino Lewis rats through th e insertion of a needle into the anterior chamber connected to a saline col umn. Elevated IOP at 110 mm Hg was maintained for 60 minutes. Groups of ani mals were euthanatized at various times after reperfusion, and their retina s were evaluated by morphology, agarose gel electrophoresis of DNA, in situ terminal deoxynucleotidyl transferase-mediated biotin-deoxyuridine triphos phate nick-end labeling (TUNEL), immunohistochemistry of caspases II (ICH1) and III (CPP32), and morphometry. YVAD.CMK, a tetrapeptide inhibitor of ca spases, was used to examine the involvement of caspases. RESULTS. A marked ladder pattern in retinal DNA gel analysis, typical of in ternucleosomal DNA fragmentation and characteristic of apoptosis, was prese nt 12 and 18 hours after reperfusion. Labeling of nuclei in the RGCL and th e inner nuclear layer (INL) by TUNEL was noted between 8 and 18 hours after reperfusion. Histologic and ultrastructural features typical of apoptosis were also observed in the inner retina after ischemia. TVAD.CMK administere d during the ischemic period inhibited apoptotic fragmentation of retinal D NA and ameliorated the tissue damage. When administered intravitreally 0, 2 , or 4 hours after reperfusion, YVAD.CMK was also effective in preserving t he inner retina but had no significant effect when administered G or 8 hour s after reperfusion. The inner retina showed transient elevated immunoreact ivity of caspases II and III 4 and 8 hours after reperfusion. CONCLUSIONS. Retinal ischemia-reperfusion after transient elevated IOP indu ced apoptosis of cells in the retinal ganglion cell layer and the INL. Casp ases may have a pivotal role in the early events of the apoptotic pathway(s ). Rescue by using anti-apoptotic agents after ischemia-reperfusion is feas ible.