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The nuclear receptor NR2E3 promotes expression of rod photoreceptor genes while repressing cone genes. Mice lacking NR2E3 (Nr2e3rd7/rd7 referred to here as rd7) are a model for enhanced S-cone syndrome, a disease associated with increased sensitivity to blue light and night blindness. Rd7 retinas have reduced levels of the outer segment (OS) structural protein retinal degeneration slow (RDS). We test the hypothesis that increasing RDS levels would improve the Rd7 phenotype. Transgenic mice over-expressing normal mouse peripherin/RDS (NMP) in rods and cones were crossed onto the rd7 background. Disease phenotypes were assessed in NMP/rd7 eyes and compared to wild-type (WT) and rd7 eyes at postnatal day 30. NMP/rd7 retinas expressed total RDS (transgenic and endogenous) message at WT levels, and NMP protein was correctly localized to the OS. NMP/rd7 retinas have shorter OSs compared to rd7 and WT and significantly reduced number of rosettes. NMP/rd7 mice also exhibited significant deficits in scotopic ERG amplitudes compared to rd7 while photopic amplitudes remained unaffected. Protein levels of rhodopsin, RDS, and the RDS homologue ROM-1 were significantly reduced in the NMP/rd7 retinas compared to rd7. We show that correcting the levels of RDS gene expression does not improve the phenotype of the rd7 suggesting that RDS deficiency is not responsible for the defect in this model. We suggest that the specific rod defect in the NMP/rd7 is likely associated with ongoing problems in the rd7 that are related to the expression of cone genes in rod cells, a characteristic of the model. [ABSTRACT FROM AUTHOR]

Gene therapy is a critical tool for the treatment of monogenic retinal diseases. However, the limited vector capacity of the current benchmark delivery strategy, adeno-associated virus (AAV), makes development of larger capacity alternatives, such as compacted DNA nanoparticles (NPs), critical. Here we conduct a side-by-side comparison of self-complementary AAV and CK30PEG NPs using matched ITR plasmids. We report that although AAVs are more efficient per vector genome (vg) than NPs, NPs can drive gene expression on a comparable scale and longevity to AAV. We show that subretinally injected NPs do not leave the eye while some of the AAV-injected animals exhibited vector DNA and GFP expression in the visual pathways of the brain from PI-60 onward. As a result, these NPs have the potential to become a successful alternative for ocular gene therapy, especially for the multitude of genes too large for AAV vectors. [ABSTRACT FROM AUTHOR]

The neural retinal leucine zipper (Nrl) knockout mouse is a widely used model to study cone photoreceptor development, physiology, and molecular biology in the absence of rods. In the Nrl-/- retina, rods are converted into functional cone-like cells. The Nrl-/- retina is characterized by large undulations of the outer nuclear layer (ONL) commonly known as rosettes. Here we explore the mechanism of rosette development in the Nrl-/- retina. We report that rosettes first appear at postnatal day (P)8, and that the structure of nascent rosettes is morphologically distinct from what is seen in the adult retina. The lumen of these nascent rosettes contains a population of aberrant cells protruding into the subretinal space that induce infolding of the ONL. Morphologically adult rosettes do not contain any cell bodies and are first detected at P15. The cells found in nascent rosettes are photoreceptors in origin but lack inner and outer segments. We show that the adherens junctions between photoreceptors and Müller glia which comprise the retinal outer limiting membrane (OLM) are not uniformly formed in the Nrl-/- retina and thus allow protrusion of a population of developing photoreceptors into the subretinal space where their maturation becomes delayed. These data suggest that the rosettes of the Nrl-/- retina arise due to defects in the OLM and delayed maturation of a subset of photoreceptors, and that rods may play an important role in the proper formation of the OLM. [ABSTRACT FROM AUTHOR]

Previously we have shown that compacted DNA nanoparticles can drive high levels of transgene expression after subretinal injection in the mouse eye. Here we delivered compacted DNA nanoparticles containing a therapeutic gene to the retinas of a mouse model of retinitis pigmentosa. Nanoparticles containing the wild-type retinal degeneration slow (Rds) gene were injected into the subretinal space of rds+/- mice on postnatal day 5. Gene expression was sustained for up to four months at levels up to four times higher than in controls injected with saline or naked DNA. The nanoparticles were taken up into virtually all photoreceptors and mediated significant structural and biochemical rescue of the disease without histological or functional evidence of toxicity. Electroretinogram recordings showed that nanoparticle-mediated gene transfer restored cone function to a near-normal level in contrast to transfer of naked plasmid DNA. Rod function was also improved. These findings demonstrate that compacted DNA nanoparticles represent a viable option for development of gene-based interventions for ocular diseases and obviate major barriers commonly encountered with non-viral based therapies. [ABSTRACT FROM AUTHOR]