Abstract

Nonviral gene transfer to neurons remains unreliable due to a lack of effective and nontoxic vectors. Here, we achieved effective neuronal gene delivery through salt-free complexation of plasmid DNA and pullulan–spermine, a conjugate prepared from a naturally derived polysaccharide and polyamine. Specifically, at low spermine nitrogen:DNA phosphate (N:P) ratios, complexes formed with ζ-potential and diameter of approximately−40 mV and 350 nm, respectively. Higher N:P ratios increased the ζ-potential to approximately +10 mV. All complexes were stable for at least 1 week and protected DNA from degradation. In vitro transfection of rat sensory neurons occurred at all N:P ratios, but uniquely, efficiency was highest for anionic complexes (anioplexes). Subsequent analyses revealed the inhibition of reporter gene expression by asialofetuin (1 mg/ml) and methyl-beta-cyclodextrin (5 m m), indicating utilization of glycoprotein-specific interactions and lipid rafts for uptake and intracellular trafficking. In marked contrast to a commercial cationic lipid reagent, anioplexes did not exhibit measurable cytotoxicity at up to 20 μg/ml DNA. Additionally, transfection efficiency was maintained in the presence of serum and antibiotics. Based on these favorable properties, we successfully established two transfection methods for cultured adult sensory neurons and tissue explants. Collectively, these data suggest that negatively charged pullulan–spermine/DNA anioplexes could represent an effective gene delivery technology, particularly for neurons.

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call

Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.