Low molecular weight chitosan nanoparticles for CpG oligodeoxynucleotides delivery: Impact of molecular weight, degree of deacetylation, and mannosylation on intracellular uptake and cytokine induction.

Although synthetic CpG oligodeoxynucleotides (ODNs) have shown substantial potential as immunotherapeutic agents, their effective intracellular delivery remains challenging. In this work, nanoparticles prepared from low-molecular weight (LMW) chitosans were investigated as CpG ODN delivery systems. Chitosan samples with a molecular weight (Mw) of 5 and 15 kDa and degree of deacetylation (DDA) of 50 and 80% were prepared. Additionally, mannosylated chitosans with a substitution degree of 15% were synthesized. The impact of LMW chitosan Mw and DDA on nanoparticle physical properties and the associated immunostimulatory effect in RAW 264.7 cells was studied. Nanoparticles prepared with chitosan of higher DDA and larger Mw exhibited better CpG ODN binding ability and intracellular uptake. Nevertheless, the most efficient immunostimulatory effect was observed while using 50% acetylated and mannosylated samples. The decreased charge density on chitosan backbone resulted in the enhanced intracellular CpG ODN release, which promoted in vitro cytokine secretion. Moreover, mannose ligand grafting promoted nanoparticle uptake through receptor-mediated recognition. Overall, this research suggests that chitosan structural parameters can be modulated to prepare LMW chitosan nanoparticles that first efficiently encapsulate CpG ODN, and then release it in immune cells, thus may be used as an efficient vector for intracellular CpG ODN delivery.

Novel protein-lipid composite nanoparticles with an inner aqueous compartment as delivery systems of hydrophilic nutraceutical compounds.

Food protein and lipid based nanoparticles have attracted recent interest as a means of delivering nutraceuticals. Nanoparticle encapsulation of nutraceuticals faces challenges to overcome for it to be readily applied in the food industry, such as low encapsulation efficiency for hydrophilic compounds and poor stability once in the gastrointestinal tract. This research introduces a new protein-lipid composite nanoparticle with a three-layered structure (a barley protein layer, α-tocopherol layer and phospholipid layer) and an inner aqueous compartment to load hydrophilic nutraceuticals. This delivery system showed efficient encapsulation of vitamin B12 (69%) and controlled release behavior in simulated gastrointestinal media. An in vitro cell evaluation demonstrated that nanoparticles could internalize into Caco-2 cells via energy-dependent endocytosis and significantly increase the uptake and transport efficiency of vitamin B12 in this model. In vivo, the developed vitamin B12 loaded nanoparticle showed increased serum vitamin B12 levels upon oral administration and reduced the methylmalonic acid level more efficiently than the free form in rats. A 14-day in vivo toxicity study showed no evidence of toxicity in rats implying the safety of the developed nanoparticles in long term use. Overall, the results of this study show the great potential of the developed nanoparticles in increasing the absorption of vitamin B12 upon oral administration.