Assessment of retinal oxygen metabolism, visual function, thickness and degeneration markers after variable ischemia/reperfusion in rats

Abstract

After total retinal ischemia induced experimentally by ophthalmic vessel occlusion followed by reperfusion, studies have reported alterations in retinal oxygen metabolism (MO₂), delivery (DO₂), and extraction fraction (OEF), as well as visual dysfunction and cell loss. In the current study, under variable durations of ischemia/reperfusion, changes in these oxygen metrics, visual function, retinal thickness, and degeneration markers (gliosis and apoptosis) were assessed and related. Additionally, the prognostic value of MO₂ for predicting visual function and retinal thickness outcomes was reported. Sixty-one rats were divided into 5 groups of ischemia duration (0 [sham], 60, 90, 120, or 180 min) and 2 reperfusion durations (1 h, 7 days). Phosphorescence lifetime and blood flow imaging, electroretinography, and optical coherence tomography were performed. MO₂ reduction was related to visual dysfunction, retinal thinning, increased gliosis and apoptosis after 7-days reperfusion. Impairment in MO₂ after 1-h reperfusion predicted visual function and retinal thickness outcomes after 7-days reperfusion. Since MO₂ can be measured in humans, findings from analogous studies may find value in the clinical setting.

Keywords: Ophthalmic vessel occlusion; Retinal ischemia; Retinal oxygen metabolism.

Figure 1.. Study design and groups

Diagram of the sequence of various quantities assessed after ophthalmic vessel occlusion (OVO). Rats underwent unilateral OVO or sham surgery. Animals were divided into 5 ischemia duration groups (0 [sham], 60, 90, 120 and 180 minutes) and 2 reperfusion duration (1 hour, 7 days) groups. ERG: Electroretinography, OCT: spectral-domain optical coherence tomography, PO₂: vascular oxygen tension.

Figure 2.. Retinal blood flow and oxygen delivery following ophthalmic vessel occlusion

Measurements of retinal oxygen metrics stratified by ischemia duration of 0 (sham), 60, 90, 120 and 180 minutes, and reperfusion durations of 1 hour or 7 days. (A) Total retinal blood flow (TRBF) and (B) oxygen delivery (DO₂). The data are presented as mean ± SD. Different of animals were evaluated in each group. * denotes significant difference compared to the sham group.

Figure 3.. Retinal oxygen metabolism and extraction fraction following ophthalmic vessel occlusion

Measurements of retinal oxygen metrics stratified by ischemia duration of 0 (sham), 60, 90, 120 and 180 minutes, and reperfusion durations of 1 hour or 7 days. (A) Oxygen metabolism (MO₂) and (B) Oxygen extraction fraction (OEF). The data are presented as mean ± SD. Different of animals were evaluated in each group. * denotes significant difference compared to the sham group.

Figure 4.. Retinal electrophysiological function following ophthalmic vessel occlusion

Measurements of (A) a- and (B) b-wave amplitude after 7 days of reperfusion, stratified by duration of ischemia. The data are presented as mean ± SD. * denotes significant difference compared to the sham group.

Figure 5.. Retinal gliosis following ophthalmic vessel occlusion

(A) Glial fibrillary acidic protein (GFAP) in representative retinal sections of 0-minute (sham), 60-minute, 90-minute, 120-minute, and 180-minute ischemia duration groups. Active gliosis is depicted as the green color, and cell nuclei are depicted as the blue color (DAPI). (B) GFAP score (gliosis scoring system ranging from 0-3) measured after 7 days of reperfusion, stratified by duration of ischemia. The data are presented as mean ± SD. NFL: nerve fiber layer, RGCL: retinal ganglion cell layer, IPL: inner plexiform layer, INL: inner nuclear layer, OPL: outer plexiform layer, ONL outer nuclear layer. * denotes significant difference compared to the sham group.

Figure 6.. Retinal apoptotic cells following ophthalmic vessel occlusion

(A) TUNEL staining, depicted as the green color, colocalized with cell nuclei, presented as blue color (DAPI) in representative retinal sections. The white arrows indicate co-expression in the same cell. (B) The number of TUNEL-positive cells measured after 7 days of reperfusion, stratified by duration of ischemia. The data are presented as mean ± SD. * denotes significant difference compared to the sham group.

Figure 7.. Retinal thickness following ophthalmic vessel occlusion

Total retina thickness (TRT) measured after 7 days of reperfusion, stratified by duration of ischemia. The data are presented as mean ± SD. * denotes significant difference compared to the sham group.

Figure 8.. Relationships of MO₂, b-wave amplitude and total retinal thickness impairment indices following ophthalmic vessel occlusion

Impairment indices of b-wave amplitude and total retinal thickness (TRT) after 7-days of reperfusion are plotted as a function of MO₂ impairment index after 1-hour reperfusion.