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D. C. Chung, V. Lee, L. H. Vandenberghe, D. Cheng, Z. Wei, J. M. Wilson, J. Bennett; Impact of Single Amino Acid Modifications of Adeno-Associated Virus (AAV) Capsid and Viral Dose on Transduction Patterns After Transuterine Subretinal Delivery of AAV Vectors to the Fetal Retina. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3019.
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© ARVO (1962-2015); The Authors (2016-present)
To compare transduction patterns after transuterine subretinal delivery of transgenes through novel AAVs with wildtype or modified capsids to the fetal retina in mice. It is proposed that viral tropism and transduction characteristics could be altered by single nucleotide viral capsid modifications or viral dosage. Optimization of retinal transduction properties could lead to increased efficacy in treatments designed for early onset retinal degenerative diseases.
Wild-type C57Bl6 mice underwent timed pregnancies to determine the date of conception. At embryonic day 14.5-15.5 (E14.5-15.5), transuterine injections were conducted to place a wildtype or modified AAV2 vector carrying the enhanced green fluorescent protein marker gene driven by the CMV promoter into the embryonic subretinal space. Adeno-associated viral vectors tested were AAV2/6, AAV2/6.1, AAV2/6.1.2, AAV2/6.2. Vectors were injected at a high dose (2.8E9gc - 2.4E10gc) and low dose (5.5E8gc). Following successful gestation and delivery, pups were maintained for an additional 28 days. Animals were then euthanized, eyes enucleated, cryoprotected and sectioned. Retinal sections were examined on a fluorescent microscope for GFP expression.
The cellular specificity and efficiency of transduction exhibited slight differences in expression patterns after high titer injections. At high titer, photoreceptor transduction was seen with all vectors used, however transduction with AAV2/6.2 was significantly reduced, as measured by eGFP expression. At low titer, all vectors showed expression in retinal pigment epithelial cells only. All injected mice developed normally and without apparent toxicity secondary to the transuterine injection. The majority of dams were able to rear subsequent litters without difficulty.
Viral titer and AAV capsid modification can alter tropism and expression efficiency after transuterine subretinal delivery to the fetal retina. Such differences could be exploited to optimize gene targeting to the fetal retina and influence therapies for congenital or pediatric retinal degenerative diseases.
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