Treatment with polyamine oxidase inhibitor reduces excitotoxicity-mediated retinal neuro-inflammation

Abstract

Purpose : Neuronal injury to retina is the major cause of vision impairment in blinding diseases worldwide. Cellular damage resulting from polyamine catabolism has been demonstrated to be a major player in many neurodegenerative conditions that affect the brain.We have previously shown that inhibition of polyamine oxidase (PAO) using MDL 72527 significantly reduced retinal neurodegeneration and cell death signaling pathways in hyperoxia-mediated retinopathy. In the present study, we investigated the efficacy of PAO inhibition in limiting NMDA (N-Methyl-D-aspartate) induced excitotoxic retinal damage.

Methods : Adult mice (8-10 weeks old) were given intravitreal injections (40 nmoles) of NMDA or NMLA (N-Methyl-L-aspartate, control). Intraperitoneal injection of MDL 72527 (40 mg/kg body weight/day) or vehicle (normal saline) was given 24 h before NMDA or NMLA treatment, and continued until animals were sacrificed (varied from 1-7 days). Retinal cryostat sections were prepared for morphometry analysis, immunostaining and TUNEL assay. Analyses of retinal ganglion cell (RGC) layer cells and microglia were performed on retinal flatmounts. Fresh frozen retinal samples were used for Western blotting analysis.

Results : A marked decrease in RGC survival was observed in NMDA treated retinas compared to their sham and NMLA treated controls, as studied by retinal flatmount analysis of NeuN-positive neurons in RGC layer. Treatment with MDL 72527 significantly reduced this RGC loss in NMDA treated retinas (p<0.05). This was confirmed by TUNEL labelling studies and morphometry analysis of retinal sections. Further studies demonstrated increased activation of microglia (studied by Iba1 immunostaining) in response to NMDA, in vehicle treated retinas, suggesting the involvement of inflammatory signaling in NMDA-mediated neuronal injury. However, microglial activation was abrogated by MDL 72527 treatment. Analysis of signaling pathways during excitotoxic injury revealed activation of NF-κB and reduced levels of pro-survival molecules p-ERK and p-CREB, which were normalized with PAO inhibition.

Conclusions : Our data demonstrate that treatment with polyamine oxidase inhibitor reduces NMDA-induced neurotoxicity, limits microglial activation and promotes cell survival in the retina, thus offering a new therapeutic target for retinal neuro-inflammatory disease conditions.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.