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Wai Siene Ng, Bing Song, Kate Binley, James E Morgan; Corneal Gene Therapy: Magnetofection. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3339. doi: https://doi.org/.
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Inherited, iatrogenic and metabolic corneal disease could potentially be treated by supplying a functional gene or changing the expression levels of specific genes. Viral-based gene therapy is efficient but restricted by potential inflammatory response and difficulties in restricting the vectors to a region of interest. This study aims to transfect mouse cornea with a non-viral based magnetofection method for the targeted delivery of non-viral genetic material.
Cultured mouse corneas were treated with magnetic nanoparticles tethered to chick beta actin promoter and green fluorescent protein [GFP] plasmids and exposed to a 2Hz oscillating magnetic field for 30 minutes. Corneas were cultured for up to 14 days post-transfection and images were captured by confocal microscopy. Green fluorescent channel intensity and number of cells exhibiting fluorescence were quantified. The experiment was triplicated for each day endpoint and an average of four sample areas were taken from each cornea. Green fluorescence intensity was adjusted for corneal autofluorescence against controls whereby plasmids were cultured with cornea in the absence of a magnetic field.
GFP expression was detected in mouse cornea from as early as day 1 (9.03+/-9.37) and persisted until day 14 at low levels (3.96+/-2.15). Green fluorescence intensity peaked at 17.9 +/-10.2 on day 4. No fluorescent cells were detected in control experiments (no magnetic field). GFP expressing cells were detected in all layers of the cornea; endothelium, stroma and epithelium. The average fluorescent cell density on day 4 was 531.2 cells per mm2. An estimated transfection efficiency of 23.8% was calculated based on the average endothelial cell density of 2234 cells per mm2.
We demonstrate a novel non-viral method for corneal transfection. An important feature is that transfection does not occur in the absence of a magnetic field thereby limiting off target effect. Magnetofection is a promising technique for the treatment of some anterior segment disorders.
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