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Rod Outer Segment Development Influences AAV-Mediated Photoreceptor Transduction After Subretinal Injection.

ABSTRACT: Vectors based on the adeno-associated virus (AAV) are currently the preferred tools for delivering genes to photoreceptors (PR) in small and large animals. AAVs have been applied successfully in various models of PR dystrophies. However, unknown barriers still limit AAV's efficient application in several forms of severe PR degenerations due to insufficient transgene expression and/or treated cells at the time of injection. Optimizations of PR gene therapy strategies will likely benefit from the identification of the cellular factors that influence PR transduction. Interestingly, recent studies have shown that the AAV transduction profile of PRs differs significantly between neonatal and adult mouse retinas after subretinal injection. This phenomenon may provide clues to identify host factors that influence the efficiency of AAV-mediated PR transduction. This study demonstrates that rod outer segments are critical modulators of efficient AAV-mediated rod transduction. During retinal development, rod transduction correlated temporally and spatially with the differentiation order of PRs when vectors were introduced subretinally but not when introduced intravitreally. All subretinally injected vectors had an initial preference to transduce cones in the absence of formed rod outer segments and then displayed a preference for rods as the cells matured, independently of the expression cassette or AAV serotype. Consistent with this observation, altered development of rod outer segments was associated with a strong reduction of rod transduction and an increase in the percentage of transduced cones by 2- to 2.8-fold. A similar increase of cone transduction was observed in the adult retinal degeneration 1 (rd1) retina compared to wild-type mice. These results suggest that the loss of rod outer segments in diseased retinas could markedly affect gene transfer efficiency of AAV vectors by limiting the ability of AAVs to infect dying rods efficiently. This information could be exploited for the development of more efficient AAV-based PR gene delivery procedures.

SUBMITTER: Petit L

PROVIDER: S-EPMC5488363 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Rod Outer Segment Development Influences AAV-Mediated Photoreceptor Transduction After Subretinal Injection.

Petit Lolita L Ma Shan S Cheng Shun-Yun SY Gao Guangping G Punzo Claudio C

Human gene therapy 20170601 6

Vectors based on the adeno-associated virus (AAV) are currently the preferred tools for delivering genes to photoreceptors (PR) in small and large animals. AAVs have been applied successfully in various models of PR dystrophies. However, unknown barriers still limit AAV's efficient application in several forms of severe PR degenerations due to insufficient transgene expression and/or treated cells at the time of injection. Optimizations of PR gene therapy strategies will likely benefit from the ...[more]

PMID: 28510482

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Project description:Transplantation of photoreceptor precursor cells (PPCs) into the retina represents a promising treatment for cell replacement in blinding diseases characterized by photoreceptor loss. In preclinical studies, we and others demonstrated that grafted PPCs integrate into the host outer nuclear layer (ONL) and develop into mature photoreceptors. However, a key feature of light detecting photoreceptors, the outer segment (OS) with natively aligned disc membrane staples, has not been studied in detail following transplantation. Therefore, we used as donor cells PPCs isolated from neonatal double transgenic reporter mice in which OSs are selectively labeled by green fluorescent protein while cell bodies are highlighted by red fluorescent protein. PPCs were enriched using CD73-based magnetic associated cell sorting and subsequently transplanted into either adult wild-type or a model of autosomal-dominant retinal degeneration mice. Three weeks post-transplantation, donor photoreceptors were identified based on fluorescent-reporter expression and OS formation was monitored at light and electron microscopy levels. Donor cells that properly integrated into the host wild-type retina developed OSs with the formation of a connecting cilium and well-aligned disc membrane staples similar to the surrounding native cells of the host. Surprisingly, the majority of not-integrated PPCs that remained in the sub-retinal space also generated native-like OSs in wild-type mice and those affected by retinal degeneration. Moreover, they showed an improved photoreceptor maturation and OS formation by comparison to donor cells located on the vitreous side suggesting that environmental cues influence the PPC differentiation and maturation. We conclude that transplanted PPCs, whether integrated or not into the host ONL, are able to generate the cellular structure for effective light detection, a phenomenon observed in wild-type as well as in degenerated retinas. Given that patients suffering from retinitis pigmentosa lose almost all photoreceptors, our findings are of utmost importance for the development of cell-based therapies.

Dataset Information

Rod Outer Segment Development Influences AAV-Mediated Photoreceptor Transduction After Subretinal Injection.

Publications

Rod Outer Segment Development Influences AAV-Mediated Photoreceptor Transduction After Subretinal Injection.

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