Abstract
Purpose :
Endothelin-1 (ET-1) is a vasoactive peptide whose levels are elevated both in the aqueous humor and circulation of primary open angle glaucoma patients as well as in animal models of glaucoma. Intravitreal ET-1 treatment in rodents has been shown to produce retinal ganglion cells (RGCs) loss, axonal injury, and disruption of nerve fiber layer. However, the precise mechanisms underlying these effects are still not completely understood. The purpose of the study was to assess mitochondrial mechanisms underlying ET-1 mediated neurodegeneration of RGCs in culture as well as in the Morrison model of ocular hypertension in rats.
Methods :
Primary RGCs isolated from rat pups were treated with ET-1 (100 nM) for 24 h and reactive oxygen species were detected with the CellRox Green reagent and mitochondrial membrane potential was measured with the JC-1 dye. Mitophagy was assessed in the RGCs by treating with MitoTracker (labels mitochondria in live cells) and LysoTracker (tracks lyosomes in live cells). To confirm these findings in vivo, intraocular pressure (IOP) elevation was carried out in one eye of retired breeder Brown Norway rats. Two weeks post-IOP elevation, retina sections from IOP-elevated eyes and contralateral eyes were analysed for expression of LC3B (a marker of autophagosomes), LAMP-1 (lysosomal marker) and TOM20 (mitochondrial marker).
Results :
ET-1 treatment of primary RGCs for 24 h produced a significant decrease (n=3, p < 0.05) in reactive oxygen species and a decrease in mitochondrial membrane potential. A decreased co-localization of MitoTracker and LysoTracker was found in primary RGCs treated with ET-1, indicative of decreased mitophagy. IOP elevation for 2 weeks in rats, produced a significant decrease in co-localization of LC-3B and TOM-20 (n=4, p<0.05) as well as LAMP-1 and TOM-20 in ganglion cell layer and nerve fiber layer.
Conclusions :
Treatment of primary RGCs with ET-1 produced an elevation in reactive oxygen species which could result in mitochondrial damage. Following IOP elevation in rats, a decreased colocalization of TOM-20 with LC-3B as well as with LAMP-1 could be indicative of ET-1 mediated decrease in mitophagy in RGCs. Mitochondrial damage in conjunction with reduced mitophagy will exacerbate oxidative damage and neuronal injury, which could be one of the mechanisms underlying ET-1 mediated neurodegeneration of RGCs in glaucoma.
This is a 2021 ARVO Annual Meeting abstract.