Effects of incorporation of poly(γ-glutamic acid) in chitosan/DNA complex nanoparticles on cellular uptake and transfection efficiency

Chitosan (CS)/DNA complex nanoparticles (NPs) have been considered as a vector for gene delivery. Although advantageous for DNA packing and protection, CS-based complexes may lead to difficulties in DNA release once arriving at the site of action. In this study, an approach through modifying their internal structure by incorporating a negatively charged poly(γ-glutamic acid) (γ-PGA) in CS/DNA complexes (CS/DNA/γ-PGA NPs) is reported. The analysis of small angle X-ray scattering results revealed that DNA and γ-PGA formed complexes with CS separately to yield two types of domains, leading to the formation of "compounded NPs". With this internal structure, the compounded NPs might disintegrate into a number of even smaller sub-particles after cellular internalization, thus improving the dissociation capacity of CS and DNA. Consequently, after incorporating γ-PGA in CS/DNA complexes, a significant increase in their transfection efficiency was found. Interestingly, in addition to improving the release of DNA intracellularly, the incorporation of γ-PGA in CS/DNA complexes significantly enhanced their cellular uptake. We further demonstrated that besides a non-specific charged-mediated binding to cell membranes, there were specific trypsin-cleavable proteins involved in the internalization of CS/DNA/γ-PGA NPs. The aforementioned results indicated that γ-PGA played multiple important roles in enhancing the cellular uptake and transfection efficiency of CS/DNA/γ-PGA NPs.