Chitosan is an ideal candidate for oral DNA delivery due to its mucoadhesive properties. Chitosan (CS) produced under GMP conditions from fungal source was used to encapsulate a plasmid DNA coding for a reporter gene. Nanoparticles made by complex coacervation of CS and DNA had a size around 200 nm, a positive zeta potential, a high association of DNA and protected the plasmid against nuclease degradation. Their transfection ability was assessed in differentiated intestinal Caco-2 cells. An N/P ratio of 4 and a DNA concentration of 8 microg/ml were the optimal conditions leading to a transfection efficiency similar to the one reached with polyethyleneimine (PEI)-DNA complexes without cytotoxicity. M cells in monolayers influenced DNA uptake up to 8 microg of DNA/ml when complexed with CS. Fungal trimethylchitosan was also tested but the complexes interactions were too strong to induce transfection in vitro. Confocal microscopy studies showed that CS/DNA and PEI/DNA nanoparticles were found at the apical surface of cell monolayers and DNA was co-localized within the nucleus. Quantification seemed to show that more DNA was associated with the cells when incubated with CS nanoparticles and that the presence of M cells slightly influenced DNA uptake when complexed with CS. In conclusion, we developed a new nanocarrier made of fungal CS promising for oral gene delivery and oral DNA vaccination.
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