Oxidative Stress Mediates the Spermidine-Induced Degeneration of Retinal Pigment Epithelium in Rats

Abstract

Purpose: : The degeneration of retinal pigment epithelial (RPE) cells is a crucial event in dry age-related macular degeneration (dry AMD) and gyrate atrophy. Spermidine, a metabolite of ornithine, is shown to inhibit the growth of RPE cells in vitro. This study was performed: 1) to elucidate the mechanisms underlying spermidine-induced RPE cell death and 2) to establish a new animal model of retinal degeneration.

Methods: : ARPE-19 cells were cultured in the presence or absence of spermidine with/without fetal bovine serum (FBS), aminoguanidine (a polyamine oxidase inhibitor), N-acetylcysteine (NAC, an antioxidant) or aldehyde dehydrogenase (ALDH, a metabolic enzyme for acrolein). Cell viability was assessed by MTS assay 24 hours after the treatment. For an in vivo study, spermidine (10, 20 and 30 nmol/eye) was injected into the vitreous body of adult Brown Norway rats, and then the permeability of the blood-retinal barrier (BRB) was measured by vitreous fluorophotometry 1, 3, 7 and 28 days after injection of spermidine. The scotopic electroretinogram (ERG) was also measured 2, 6, 13 and 27 days after injection. The eyes were subjected to histological examinations after measurements of permeability.

Results: : Spermidine caused cell death of ARPE-19 in a concentration-dependent manner, and this effect was prevented by removal of 10%FBS from the media. Aminoguanidine, NAC or ALDH significantly inhibited spermidine-induced cell death. Spermidine significantly increased the permeability of outer BRB from day 7 to 28, and decreased the amplitude of ERG a- and b-waves from day 6 to 27 after the intravitreous injection. Histological analysis showed that spermidine induced vacuolation, atrophy and loss of RPE cells, and then caused the disruption of photoreceptor outer segments. Simultaneous intravitreous administration of NAC with spermidine prominently inhibited the functional and morphological changes induced by spermidine.

Conclusions: : The present study demonstrated that intravitreous administration of spermidine induced the dysfunction and death of RPE cells followed by the degeneration of photoreceptor cells. These effects may be mediated by oxidative stress generated during polyamine oxidation. This animal model would be useful to understand the pathophysiology of dry AMD and gyrate atrophy.

Keywords: retinal pigment epithelium • oxidation/oxidative or free radical damage • degenerations/dystrophies