A Novel Method Combining Vitreous Aspiration and Intravitreal AAV2/8 Injection Results in Retina-Wide Transduction in Adult Mice.
Invest Ophthalmol Vis Sci
; 57(13): 5326-5334, 2016 Oct 01.
Article
em En
| MEDLINE
| ID: mdl-27784063
ABSTRACT
PURPOSE:
Gene therapies to treat eye disorders have been extensively studied in the past 20 years. Frequently, adeno-associated viruses were applied to the subretinal or intravitreal space of the eye to transduce retinal cells with nucleotide sequences of therapeutic potential. In this study we describe a novel intravitreal injection procedure that leads to a reproducible adeno-associated virus (AAV)2/8-mediated transduction of more than 70% of the retina.METHODS:
Prior to a single intravitreal injection of a enhanced green fluorescent protien (GFP)-expressing viral suspension, we performed an aspiration of vitreous tissue from wild-type C57Bl/6J mice. One and one-half microliters of AAV2/8 suspension was injected. Funduscopy, optical coherence tomography (OCT), laser scanning microscopy of retinal flat mounts, cryosections of eye cups, and ERG recordings verified the efficacy and safety of the method.RESULTS:
The combination of vitreous aspiration and intravitreal injection resulted in an almost complete transduction of the retina in approximately 60% of the eyes and showed transduced cells in all retinal layers. Photoreceptors and RPE cells were predominantly transduced. Eyes presented with well-preserved retinal morphology. Electroretinographic recordings suggested that the new combination of techniques did not cause significant alterations of the retinal physiology.CONCLUSIONS:
We show a novel application technique of AAV2/8 to the vitreous of mice that leads to widespread transduction of the retina. The results of this study have implications for virus-based gene therapies and basic science; for example, they might provide an approach to apply gene replacement strategies or clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in vivo. It may further help to develop similar techniques for larger animal models or humans.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Doenças Retinianas
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Sucção
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Transdução Genética
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Terapia Genética
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Dependovirus
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Vetores Genéticos
Tipo de estudo:
Diagnostic_studies
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Prognostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2016
Tipo de documento:
Article