Use of magnetic nanoparticles and oscillating magnetic field for non-viral gene transfer into mouse cornea.

Abstract

BACKGROUND: Inherited, iatrogenic, and metabolic corneal disease could be potentially treated by supplying a functional gene or changing the expression levels of specific genes. Viral-based gene therapy is efficient, but restricted by evoking immune responses and inflammation. This study aimed to transfect mouse cornea with a non-viral based (oscillating magnetofection) method. METHODS: Cultured mouse corneas were treated with magnetic nanoparticles (MNP) tethered to CAG promoter and green fluorescent protein (GFP) reporter plasmids exposed to a 1 Hz, 2 Hz, and 4 Hz oscillating magnetic field for 30 min and 60 min and in three DNA:MNP ratios (1:2, 1:1, 2:3). Corneas were cultured for up to 3 days and their green fluorescent channel intensity and number of GFP-positive cells were recorded. Transfection efficiency was estimated as the percentage of GFP-positive cells per total cells in a microscopic field. FINDINGS: Control experiments with absent magnetic exposure showed no GFP-positive cells. The optimum condition was recorded at 3:2 DNA:MNP ratio, 1 Hz magnetic oscillation, and 30 min duration of magnetic exposure (mean GFP-positive endothelial cell count 191·7 [SD 54·5], p=0·009; mean green fluorescent intensity 85·3 [SD 48·5]; and average transfection efficiency 23·3% [range 10·6-30·9]). INTERPRETATION: A novel non-viral method of transfecting cornea, magnetofection, is demonstrated and gives proof of principle for its translation into corneal gene therapy. FUNDING: Welsh Clinical Academic Training Scheme.