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X. Cai; Partial Rescue of Rds Haploinsufficiency Phenotype With DNA Nanoparticles. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1690.
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© ARVO (1962-2015); The Authors (2016-present)
Mutations in the retinal degeneration slow (Rds) gene are associated with multiple retinal diseases. The rds+/- haploinsufficiency phenotype involves progressive vision loss with early-onset rod degeneration (including dysmorphic outer segments (OSs)) followed by late-onset cone degeneration. Here, we tested the ability of subretinally-injected nanoparticles containing normal mouse peripherin/rds (NMP) cDNA to rescue this phenotype.
Three vectors, containing promotors for either chicken beta-actin (CBA-NMP), mouse opsin (MOP-NMP), or human interphotoreceptor retinoid-binding protein (hIRBP-NMP), were constructed to express full-length NMP cDNA and compacted as acetate nanoparticles. The NMP C-terminal sequence was modified from Glu-341 to proline (P341Q) to allow detection of expressed, transfected Rds on an endogenous Rds background, using the monoclonal antibody 3B6. At postnatal day 5, mice were injected subretinally either with nanoparticles (in 0.4 µl saline) or with saline alone; contralateral eyes served as uninjected controls. Gene expression was analyzed by qRT-PCR and immunocytochemistry at 2, 7, 14, 21, and 30 days post-injection (PI); functional rescue was evaluated by electroretinography (ERG) at PI-30.
Total Rds transcription levels (NMP and endogenous) in all nanoparticle-injected eyes were at least 2-fold higher than in control eyes, up to PI-30. Eyes injected with nanoparticles expressed transgenic NMP protein in photoreceptor OSs; in addition, ectopic Rds expression in the retinal pigment epithelium (RPE) was detected only with CBA-NMP nanoparticle-injected eyes. ERG of MOP-NMP nanoparticle-injected eyes, compared to saline control eyes, revealed improved rod, but not cone, function. A comparable structural improvement on the MOP-NMP nanoparticles injected eyes as well as structural/functional rescue of mice injected with the CBA-NMP and hIRBP-NMP nanoparticles is ongoing.
Nanoparticle-mediate Rds gene delivery provides partial functional rescue of the rds+/- haploinsufficiency phenotype. Our results provide proof-of-principle for the application of this approach as a potential early therapeutic intervention into retinal degenerative disorders.
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