Polyamine oxidase: a novel mediator of hyperoxia-induced retinal neuro-vascular injury

Abstract

Purpose: Retinopathy of prematurity (ROP) is a potentially blinding eye disease that affects premature infants and often leads to lifelong vision impairment. Hyperoxia treatment is shown to induce neurovascular damage in the developing retina of neonates. Using an oxygen-induced retinopathy mouse model of ROP, we have demonstrated that deletion of arginase 2 reduced hyperoxia-induced retinal neurodegeneration through the regulation of polyamine metabolism. In the present study, we evaluated the therapeutic benefit of inhibiting a novel target in polyamine oxidase (PAO) using MDL 72527, during hyperoxia-induced retinal neuro-vascular injury.

Methods: Newborn wild-type (WT) mice were exposed to 70% oxygen from postnatal day (P) 7 to P12, treated daily from P6 with either vehicle or MDL 72527 (39 mg/kg of body weight, in saline, i.p.) and sacrificed at different stages of hyperoxia. Cryostat sections were used for neurodegeneration studies using immunolabeling of retinal neurons. Analysis of vaso-obliteration (N=10-12), vessel sprouting (N=7-9) and microglial activation (N=6-9) was performed on retinal flatmounts. Mixed rat retinal neuronal cultures (7 DIV) were subjected to normoxia (21%) or hyperoxia (50%) with or without MDL 72527 (50 uM) (N=6-8) and processed for Western blotting.

Results: We have previously shown that inhibition of PAO significantly reduced neuronal death in hyperoxia (2 days) treated retina. Current immunolabeling studies showed improved synaptic contacts and bipolar cell morphology following PAO inhibition. Moreover, hyperoxia-treated retinal neuronal culture showed activation of neuro-inflammatory signaling as studied by increased p-NFkB, p-ERK1/2 and cytochrome-C levels, which were significantly reduced by PAO inhibition. Immunolabeling for Iba-1 showed that activation of microglia was significantly reduced and number of resting microglia were increased on P9 following PAO inhibition during hyperoxia (p<0.05). Blockade of PAO during hyperoxia showed significantly reduced avascular area on P9 (reduced by 18±2%, p<0.01) and P12 (reduced by 13±4%, p<0.01) and increased vascular sprouting on P9 (increased by 38±5%, p<0.01) compared to vehicle-treated mice.

Conclusions: Our data suggest that PAO inhibition offers a new therapeutic strategy for limiting hyperoxia-induced retinal neuro-vascular injury.