Bioactivities of Chitosan and Its Derivatives - Chitosan Based Hydrogels: Functions and Applications - page 113

complexation. Accordingly, cellular uptake was increased 8.5-fold compared to chitosan
polyplexes, resulting in up to 678-fold increased transfection efficiency in NIH/3T3 cells.
Apart from reduction of cytotoxicity, PEGylation led to improved colloidal stability of
polyplexes and significantly increased cellular uptake compared to unmodified trimethyl
chitosan. These improvements resulted in a significant, up to 10-fold increase of transfec-
tion efficiency in NIH/3T3, L929, and MeWo cells compared to trimethyl chitosan, which
not only highlights the importance of investigating polyplex stability under different pH
and ionic strength conditions, but also elucidates correlations between physicochemical
characteristics and biological efficacy of the studied polyplexes.
A self-assembled nanoparticle using a hydrophobically modified glycol chitosan (HGC)
for gene delivery has been prepared [45]. Here a primary amine of glycol chitosan was
modified with 5-cholanic acid to prepare an HGC. The modified chitosan was found to
form DNA nanoparticles spontaneously by a hydrophobic interaction between HGC and
hydrophobized DNA. As the HGC content increased, the encapsulation efficiencies of
DNA increased while the size of HGC nanoparticles decreased. Upon increasing HGC
contents, HGC nanoparticles became less cytotoxic. The increased HGC contents also facil-
itated endocytic uptakes of HGC nanoparticles by COS-1 cells. The HGC nanoparticles
showed increasing in vitro transfection efficiencies in the presence of serum. In vivo results
also showed that the HGC nanoparticles had superior transfection efficiencies compared
to naked DNA and a commercialized transfection agent.
Stearic acid (SA)-grafted chitosan oligosaccharide COSs (COS-SA) could self-aggregate
to form a micelle-like structure in aqueous solution [46]. In particular, SA, an endoge-
nous long-chain saturated fatty acid, was widely accepted for pharmaceutical use. As a
main composition of fat, SA is biocompatible with low cytoxicity. COS-SA has efficient
ability to condense the plasmid DNA to form COS complex nanoparticles, which can
efficiently protect the condensed DNA from enzymatic degradation by DNase I. The
in vitro transfection experiments showed that the optimal transfection efficiency of
COS-SA micelle in A549 cells was higher than that of COS, and comparable with
Lipofectamine™ 2000. The presence of 10% fetal bovine serum increased the transfec-
tion ability of COS-SA. On the other hand, the cytotoxicity of COS-SA was highly lower
than that of Lipofectamine 2000.
Another group used the reverse microemulsion technique as a template to fabricate chi-
tosan-alginate core-shell nanoparticles encapsulated with enhanced green fluorescent pro-
tein-encoded plasmids [47]. These alginate-coated chitosan nanoparticles endocytosed by
NIH 3T3 cells were found to trigger swelling of transport vesicles, which render gene escape
before entering the digestive endolysosomal compartment, and concomitantly promote
gene transfection rate. The results indicate that DNA-encapsulated chitosan-alginate nano-
particles with an average size of 64 nm (N/P ratio of 5) could achieve the level of gene expres-
sion comparable with the one obtained by using polyethyleneimine-DNA complexes.
Chitosan-alginate microcapsules were evaluated as a method of oral delivery of IgY
antibodies. Small and monodispersed nanoparticles with high in vitro transfection capa-
bilities were obtained by the complexation of these two polyelectrolytes. Chitosan (<10 kDa)
presents more advantageous characteristics over the HMW chitosan for clinical applica-
tions, namely increased solubility at physiological pH and improved DNA release.
Consequently, after incorporating γ-polyglycolic acid (γ-PGA) into CS-DNA complexes, a
significant increase in their transfection efficiency was found [48].
An approach for the enhancement of cellular uptake and transfection efficiency of
chitosan-DNA complexes through modifying the internal structure by incorporating a
negatively charged poly(γ-glutamic acid) was reported recently [49]. The aforementioned