Abstract
Efficient non-viral plasmid DNA transfection of most stem cells, progenitor cells and primary cell lines currently presents an obstacle for many applications within gene therapy research. From a standpoint of efficiency and cell viability, magnetic nanoparticle-based DNA transfection is a promising gene vectoring technique because it has demonstrated rapid and improved transfection outcomes when compared to alternative non-viral methods. Recently, our research group introduced oscillating magnet arrays that resulted in further improvements to this novel plasmid DNA (pDNA) vectoring technology. Continued improvements to nanomagnetic transfection techniques have focused primarily on magnetic nanoparticle (MNP) functionalization and transfection parameter optimization: cell confluence, growth media, serum starvation, magnet oscillation parameters, etc. Noting that none of these parameters can assist in the nuclear translocation of delivered pDNA following MNP-pDNA complex dissociation in the cell’s cytoplasm, inclusion of a cassette feature for pDNA nuclear translocation is theoretically justified. In this study incorporation of a DNA targeting sequence (DTS) feature in the transfecting plasmid improved transfection efficiency in model neurons, presumably from increased nuclear translocation. This observation became most apparent when comparing the response of the dividing SH-SY5Y precursor cell to the non-dividing and differentiated SH-SY5Y neuroblastoma cells.
Highlights
Gene therapy involves alteration of the amount or type of protein expression in a target cell population for the ultimate purpose of disease treatment
Nanomagnetic transfection starts with the electrostatic association of plasmid DNA and magnetic nanoparticles (MNPs) in aqueous suspension to form pDNA-MNP complexes
The performance of the control transgene reporter vs. a DNA targeting sequence (DTS)-containing plasmid can vary dramatically dependent on the vector implemented
Summary
Gene therapy involves alteration of the amount or type of protein expression in a target cell population for the ultimate purpose of disease treatment. Oscillating array-based magnetic nanoparticle DNA transfection is a gene vectoring technique that is promising because it is capable of outperforming most other non-viral transfection methods, in terms of both transfection efficiency and cell viability [4]. Nanomagnetic transfection starts with the electrostatic association of plasmid DNA (pDNA) and magnetic nanoparticles (MNPs) in aqueous suspension to form pDNA-MNP complexes. These complexes are added to culture plates containing the in vitro target cell population. The addition of a magnet array below the cell culture plate results in sedimentation of the pDNA-MNP complexes, thereby forcing sustained and proximal contact of the transgene vector and target cell [5,6,7]. One-dimensional oscillation of the magnet array produces movement of the pDNA-MNP complexes at the targeted cells’ surface that facilitates endocytosis via mechanical stimulation [4,8]
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