Protective effect of alpha-melanocyte-stimulating hormone (α-MSH) on the recovery of ischemia/reperfusion (I/R)-induced retinal damage in a rat model

Abstract

The present study demonstrates capacity of α-MSH to augment recovery from ischemia/reperfusion (I/R)-induced retinal damage in vivo and correlation of its protective effects with expression of heme oxygenase-1 (HO-1). Used techniques include ocular ischemia and reperfusion, electroretinography, histology, electron microscopy, and molecular-biological techniques. The results demonstrate the α-MSH-mediated inhibition of I/R-induced functional deterioration of the retina. Outcomes suggest that the protective effects of α-MSH occur mainly through HO-1-dependent pathways but HO-1-independent mechanisms may also contribute to protection. The observation that post-ischemic treatment with α-MSH exhibits therapeutic efficacy in the same range as pre-ischemic treatment, is a novel result. This outcome suggests a highly conserved protective role for α-MSH as a major stress response mechanism--and offers the possibility for development of novel therapeutic strategies utilizing this hormone, in particular in treatment of conditions resulting from I/R injury, such as deterioration of retinal microcirculation. The merit of the study lies in the fact that I/R injury contribute significantly to the severity of retinopathies. However, currently there are no evidence-based treatments for retinal I/R injury available for clinical use. Our finding suggests that α-MSH may have a very wide range of uses in the prevention of I/R-mediated pathologies.

Fig. 1

Electroretinograms (group I-a) of baseline (a) and ischemic/reperfused (b) eyes of vehicle-treated (control) animals and ischemic/reperfused eyes of rats treated with 50, 250, 500, and 1,000 μg/kg α-MSH (c–f, respectively)

Fig. 2

Effects of α-MSH doses on ERG b waves (group I-a). Shown here are percentages of mean values of b waves measured in electroretinogramms of baseline and ischemic/reperfused eyes of vehicle-treated (control) animals and ischemic/reperfused eyes of rats treated with 50, 250, 500, and 1,000 μg/kg α-MSH with standard error of the mean (SEM). Percentages were calculated relative to control baseline values. Horizontal lines indicate comparisons. The significance of comparisons is indicated as follows: ns not significant; *p < 0.05; **p < 0.01; ***p < 0.001

Fig. 3

Effect of α-MSH on I/R-induced retinal edema (group I-b). Sagittal sections of the rat retina showing the structure of layers in nonischemic retinal tissue of vehicle-treated (control) animals (a), I/R-injured retinal tissue from vehicle-treated (control) rats (b), and I/R-injured retinal tissue from animals treated with 500 μg/kg α-MSH (c). d The effect of α-MSH on the extent of retinal edema following I/R injury, in the inner plexiform layer. Thickness was measured using a manual scale on each glass slides. Mean ± SEM; *p < 0.05 vs. control I/R

Fig. 4

EM studies (group I-b). a–c Mitochondria in I/R-injured inner retinal cells from rats not treated with α-MSH. These images demonstrate disintegration of the mitochondria due to formation of interior vacuoles (arrows). Mitochondria in I/R-injured inner retinal cells from animals treated with 500 μg/kg α-MSH are shown in (d–g). No vacuolization is seen here

Fig. 5

Western blot analysis for HO-1 protein expression (group I-b). The HO-1 content of I/R-injured bulbi is represented in arbitrary units with SEM in animals not receiving α-MSH (control) and rats treated with 500 μg/kg α-MSH. *p < 0.05

Fig. 6

Effects of SnPP and post-ischemic α-MSH treatment on ERG voltage profiles (a) and ERG b-waves (b) obtained in part II experiments (group II). Results are shown for five groups including: nonischemic retinas from vehicle-treated rats (Control Baseline) and I/R-injured retinas from vehicle- (Control Ischemia), α-MSH- (MSH Ischemia), SnPP + α-MSH- (SnPP + MSH Ischemia), and SnPP-treated rats (SnPP Ischemia). Shown on (a) are representative ERG spikes; shown on (b) are mean percentage values (relative to control baseline) for b waves measured in electroretinograms of the aforementioned groups. Horizontal lines indicate comparisons. The significance of comparisons is indicated as follows: ns not significant; *p < 0.05; **p < 0.01; ***p < 0.001

Fig. 7

Relative HO activities in ocular tissue from eyes harvested in part II experiments (group II). Results are shown for five groups including: nonischemic retinas from vehicle-treated rats (Control Baseline) and I/R-injured retinas from vehicle- (Control Ischemia), α-MSH- (MSH Ischemia), SnPP + α-MSH- (SnPP + MSH Ischemia), and SnPP-treated rats (SnPP Ischemia). Activity of the enzyme in ocular tissue of an aforementioned group is shown as percentage of bilirubin production by a particular group relative to control baseline eye values with SEM