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X.-Q. Ding, A. B. Quiambao, J. B. Fitzgerald, M. J. Cooper, S. Conley, M. Naash; Ocular Delivery of Compacted DNA-Nanoparticles Does Not Elicit Toxicity in the Mouse Retina. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1728.
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Subretinal delivery of compacted DNA nanoparticles has been shown to direct effective transfection and expression in both photoreceptors and other retinal cells. This technology therefore provides a useful non-viral approach for gene transfer to treat inherited retinal diseases. Since one of the limitations to successful gene therapy has been vector-associated toxicity, this work evaluates the safety of compacted DNA nanoparticles following subretinal delivery.
Polyethylene glycol-substituted lysine peptide-compacted nanoparticles containing an enhanced green fluorescent protein (EGFP) expression plasmid (pZEOGFP5.1) were sub-retinally injected in adult mice (1 µl at 0.3, 1.0 and 3.0 µg/µl). Mock and saline injections were performed as controls. Retinas were examined for transgene expression and for signs of inflammation at 1, 2, 4 and 7 days post-injection (PI). H&E staining on retinal sections was performed to detect infiltration of polymorphonuclear neutrophils (PMN) and other inflammatory cells. Immunohistochemical analyses were performed to detect expression of macrophage marker F4/80 and myeloid marker myeloperoxidase (MPO). Expression levels of chemokines KC, monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor alpha (TNF-) were evaluated by ELISA and by quantitative RT-PCR.
Subretinal delivery of the nanoparticles induced expression of EGFP in photoreceptors and retinal pigment epithelium cells. Neither infiltration of PMN or lymphocytes nor elevations of F4/80 or MPO were detected in retinas of mice that had been injected with nanoparticles. Levels of KC mRNA were increased 3-4 fold in eyes that had been injected with either nanoparticles or with saline at PI-1, but returned to control level at PI-2. No elevation of KC protein was observed in these mice. Levels of MCP-1 protein were increased 3-4 fold at PI-1 for both nanoparticle and saline injected eyes, but also returned to control levels at PI-2. Thus transient elevations of MCP-1 protein were related to the subretinal injection procedure. No elevations of TNF- mRNA or protein were detected.
These investigations show no signs of local inflammatory responses or cellular toxicity following subretinal injection of compacted DNA nanoparticles, indicating that the retina may be a suitable target for future clinical nanoparticle-based interventions.
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