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A novel method combining vitreous aspiration and intravitreal AAV2/8 injection results in retina-wide transduction in adult mice


Da Costa, Romain; Röger, Carsten; Segelken, Jasmin; Barben, Maya; Grimm, Christian; Neidhardt, John (2016). A novel method combining vitreous aspiration and intravitreal AAV2/8 injection results in retina-wide transduction in adult mice. Investigative Ophthalmology & Visual Science [IOVS], 57(13):5326-5334.

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.

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.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > University Hospital Zurich > Ophthalmology Clinic
04 Faculty of Medicine > Center for Integrative Human Physiology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:1 October 2016
Deposited On:08 Mar 2017 12:55
Last Modified:02 Feb 2018 12:25
Publisher:Association for Research in Vision and Ophthalmology
ISSN:0146-0404
OA Status:Hybrid
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1167/iovs.16-19701
PubMed ID:27784063

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