NDLI: Modulation of transgene expression in retinal gene therapy by selective laser treatment.

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Modulation of transgene expression in retinal gene therapy by selective laser treatment.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Dalal, Roopa Chalberg, Thomas W. Lavinsky, Daniel Huie, Philip Marmor, Michael Mandel, Yossi Palanker, Daniel
Description	Country affiliation: United States Author Affiliation: Lavinsky D ( Department of Ophthalmology, Stanford University, Stanford, California, USA. Daniellavinsky@gmail.com)
Abstract	PURPOSE: To develop a method for modulation of transgene expression in retinal pigment epithelium (RPE) using scanning laser that spares neurosensory retina. METHODS: Fifteen pigmented rabbits received subretinal injection of recombinant adeno-associated virus (rAAV-2) encoding green fluorescent protein (GFP). GFP expression was measured using confocal scanning laser ophthalmoscopy (cSLO) fluorescence imaging and immunohistochemistry. To reduce the total expression in RPE by half, 50% of the transfected RPE cells were selectively destroyed by microsecond exposures to scanning laser with 50% pattern density. The selectivity of RPE destruction and its migration and proliferation were monitored using fluorescein angiography, spectral-domain optical coherence tomography (SD-OCT), and light, transmission, and scanning electron microscopy. 5-Bromo-2'-dioxyuridine (BrdU) assay was performed to evaluate proliferation of RPE cells. RESULTS: RPE cells were selectively destroyed by the line scanning laser with 15 µs exposures, without damage to the photoreceptors or Bruch's membrane. RPE cells started migrating after the first day, and in 1 week there was complete restoration of RPE monolayer. Selective laser treatment decreased the GFP fluorescence by 54% as compared to control areas; this was further decreased by an additional 48% following a second treatment 1 month later. BrdU assay demonstrated proliferation in approximately half of the RPE cells in treatment areas. CONCLUSIONS: Microsecond exposures produced by scanning laser destroyed RPE cells selectively, without damage to neural retina. Continuity of RPE layer is restored within days by migration and proliferation, but transgene not integrated into the nucleus is not replicated. Therefore, gene expression can be modulated in a precise manner by controlling the laser pattern density and further adjusted using repeated applications.
ISSN	01460404
e-ISSN	15525783
Journal	Investigative Opthalmology & Visual Science
Issue Number	3
Volume Number	54
Language	English
Publisher	Association for Research in Vision and Ophthalmology
Publisher Date	2013-03-13
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Gene Expression Genetic Therapy Laser Therapy Retinal Diseases Surgery Retinal Pigment Epithelium Metabolism Transgenes Genetics Animals Disease Models, Animal Microscopy, Electron, Scanning Rabbits Pathology Ultrastructure Tomography, Optical Coherence Comparative Study Research Support, Non-u.s. Gov't Discipline Ophthalmology
Content Type	Text
Resource Type	Article
Subject	Ophthalmology Sensory Systems Cellular and Molecular Neuroscience

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