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Samantha R. De Silva, Laurence Brown, Qisheng You, Mark Hankins, Robert MacLaren; Assessing Transduction Of The Degenerate Retina With AAV2/5 Using In Vivo Imaging And Immunocytochemistry. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1393.
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Artificial vision using gene therapy aims to restore light sensitivity to the retina in patients with advanced retinal degeneration, in which photoreceptors are no longer present. Our purpose was to assess the efficacy of transducing degenerate retina with AAV 2/5.GFP and to examine which retinal cell types were transduced. Since the degenerate retina is known to undergo remodelling, we also aimed to establish which cell and synapse specific markers could still be effectively used for immunocytochemistry in this tissue.
AAV2/5.GFP was injected into the subretinal space of 8 eyes of 4 rdrd mice. 23 days after injection, fundal imaging of the mice was carried out using scanning laser ophthalmoscopy (SLO). After harvesting and sectioning eyes, the retinal cell types transduced were identified using immunocytochemistry.
SLO imaging revealed widespread transduction of the retina with a large area of fluorescence in all injected mice. Immunolabelling with Calbindin, PKC alpha, CHx10, Glutamate synthetase, GFAP and GAD 65 were all effective in the degenerate retina. We demonstrated significant transduction of cells in the inner nuclear layer, these were predominantly horizontal cells and Muller cells with sparse bipolar cell transduction.The figure illustrates Calbindin staining of the degenerate retina showing colocalisation with GFP in horizontal cells.
Subretinal injection of AAV 2/5 effectively transduces horizontal cells and Muller cells in the inner nuclear layer of the degenerate retina. These cells could therefore potentially be a target of gene therapy for artificial vision to restore light sensitivity to the degenerate retina.
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