The Effects of Subretinal Delivery of Nrl Gene on Light-induced Retinopathy in a Rat Model
Date Issued
2010
Date
2010
Author(s)
Weng, Li-Ya
Abstract
Abstract
Progressive retinal atrophy (PRA) is a large group of canine inherited retinal diseases. Both PRA and retinitis pigmentosa (RP), the human homologue of PRA, share a similar clinical pathology and phenotype. Photoreceptor apoptosis leads to clinical signs of progressive loss of peripheral and night vision (rod cells) leading to loss of central and day vision (cone cells). There is no clinically effective treatment for inherited retinopathy to date. Light-induced retinopathy provides fast, synchronic photoreceptor apoptosis to mimic the inherited retinal degeneration. The mutation of neural retina leucine zipper (Nrl) gene has been shown to induce RP. 8 week-old Sprague-Dawley rats were maintained in 1500-2000 lux of cyclic light for 2 days. A mixture of Nrl gene and FuGene was introduced into eyes by subretinal injection immediately after light exposure. Retinal function was evaluated by electroretinography (ERG) before light exposure and weekly for 4 weeks (28 days) after gene delivery. Photoreceptor cell death was quantified by TUNEL assay at 1, 3, 5, and 7 days after light exposure and by measuring thickness of the outer nuclear layer (ONL) in the retina before exposure and each week after gene delivery. Retinas were harvested for immunohistochemistry studies every week till 1 month (28 days). Decrements of amplitude of ERG waves and ONL thickness, and the positive TUNEL results showed that the light-induced retinopathy in a rat model was successfully established. The Nrl gene transfection was detected by green fluorescence protein (GFP) and observed in cytoplasm of retinal pigmented epithelium (RPE), outer segment (OS) and ONL during 4 weeks after delivery, then diminished progressively. Significant retinal function preservation was observed at 3 weeks after gene delivery, the preservation of retinal ONL was also observed in the Nrl gene treated group at the first three weeks after gene delivery. TUNEL assay showed similar results of photoreceptor protection against apoptosis as well. In retinal immunohistochemistry, Nrl gene activated the expression of rhodopsin in the mature neuroretinal cells. Some transfected cells expressed positive cell markers of RPE65 for RPE, and syntaxin for amarcrine cells and horizontal cells, respectively. Based on this study, subretinal delivery of Nrl gene may provide functional and morphological preservation against photoreceptor degeneration following light-induced retinal degeneration.
Subjects
Nrl Gene
Light-induced Retinopathy
inherital retinal degneration
SDGs
Type
thesis
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