Formation of Chitosan-Based Hydrogels Network - Chitosan Based Hydrogels: Functions and Applications - page 185

close to a charge mixing ratio of 1, the size of chitosan/heparin nanoparticles tends to
aggregate [51]. Chitosan/heparin microspheres are prepared using the water-in-oil
emulsification solvent evaporation technique. The stirring speed has a particularly strong
influence on the microsphere size, and microsphere size decreases as the stirring speed
increases [52,53]. Chitosan/heparin PEC microparticles are excellent drug or protein
release carriers.
The chitosan/heparin PEC films or scaffolds are widely used in tissue engineering.
Kratz et al. [54] found that the chitosan/heparin PEC can stimulate wound healing in
human skin. Moreover, the chitosan/heparin complex networks are supposed to exist as
a network to create an appropriate environment for the regeneration of hepatocytes, as
well as to induce growth angiogenesis for the regeneration of livers. They are potential
candidates for liver tissue engineering. For example, the collagen-chitosan-heparin
[55] and alginate-galactosylated chitosan/heparin [56] scaffolds, which are similar to the
liver ECM, play important roles in the regulation of the morphological appearance of
hepatocytes.
4.2.1.5 Chitosan-DNA PECs
Chitosan-DNA PECs microparticles are prepared using a complex coacervation process
under defined conditions. The size of the complexes is of crucial importance for cellular
uptake. Small-sized complexes have the advantage of entering the cells through endocyto-
sis and/or pinocytosis. Illum et al. [57] synthesized chitosan/DNA nanoparticles ranging
from 20 to 500 nm. The chitosan/DNA PECs show excellent stability, and chitosan
microparticles protect DNA during the storage time in freeze-dried form and also from
nuclease degradation in the medium [58].
Incorporating positive-charged arginine (Arg) moieties into chitosan may increase the
charge strength and charge density of nanoparticles and enhance the electrostatic interac-
tion between nanoparticles and the cell membrane. Therefore, the cell uptake mediated by
Arg-chitosan/DNA complexes is improved compared with chitosan/DNA complexes.
The luciferase expression of Arg-chitosan is about 100-fold compared with the expression
levels mediated by chitosan for HeLa cells [59]. The zeta potential of Arg-chitosan/DNA is
about 0.23-12.5 mV, which increases with an increase of the nitrogen/phosphor (N/P) ratio
[60]. Chitosan/DNA PEC nanoparticles administered in the anterior tibialis mice muscles
reveal a high signal corresponding to β-gal gene expression within 48 h. In contrast, the
administration of naked or DNA/Lipofectamine in the anterior tibialis muscle does not
reveal any β-gal gene expression. From these preliminary data, the chitosan/DNA nano-
particles have the potential ability to transfect muscle cells in vivo and lead to protein
synthesis [61].
4.2.2 ionic Cross-linking Network
Because chitosan is a weak polybase with thousands of positive charges, anions with suf-
ficient charge numbers are effective in the ionic cross-linking of chitosan through electro-
static interactions. The processes of ionic cross-linking attract much attention because of
the simplicity, the relatively mild preparation condition procedural, and avoidance of pos-
sible toxicity of the reagents commonly used for chemical cross-linking [62]. Many multi-
valent anions and metal ions are available as cross-linkers to form the chitosan cross-linking
network. The properties of ionically cross-linked chitosan network are influenced by inter-
actions between the multivalent ions cross-linkers and chitosan. Since the interactions