Chitosan Nanoparticles for Plasmid DNA Delivery: Effect of Chitosan Molecular Structure on Formulation and Release Characteristics
ABSTRACT Chitosan can be useful as a nonviral vector for gene delivery. Although there are several reports to form chitosan-pDNA particles, the optimization and effect on transfection remain insufficient. The chitosan-pDNA nanoparticles were formulated using complex coacervation and solvent evaporation techniques. The important parameters for the encapsulation efficiency were investigated, including molecular weight and deacetylation degree of chitosan. We found that encapsulation efficiency of pDNA is directly proportional with deacetylation degree, but there is an inverse proportion with molecular weight of chitosan. DNA-nanoparticles in the size range of 450-820 nm depend on the formulation process. The surface charge of the nanoparticles prepared with complex coacervation method was slightly positive with a zeta potential of +9 to +18 mV; nevertheless, nanoparticles prepared with solvent evaporation method had a zeta potential approximately +30 mV. The pDNA-chitosan nanoparticles prepared by using high deacetylation degree chitosan having 92.7%, 98.0%, and 90.4% encapsulation efficiency protect the encapsulated pDNA from nuclease degradation as shown by electrophoretic mobility analysis. The release of pDNA from the formulation prepared by complex coacervation was completed in 24 hr whereas the formulation prepared by evaporation technique released pDNA in 96 hr, but these release profiles are not statistically significant compared with formulations with similar structure (p > .05). According to the results, we suggest nanoparticles have the potential to be used as a transfer vector in further studies.
- SourceAvailable from: Alireza Tahamtan
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- "Amongst them, chitosan (CS) is a particularly attractive cationic vector derived from polysaccharides. Polysaccharides are the most abundant polymers in nature and CS exhibits desirable characteristics such as biocompatibility and biodegradability that is essential for gene delivery (Bozkir and Saka 2004). CS is also nonimmunogenic and nontoxic. "
ABSTRACT: Chitosan nanoparticles (CS NPs) were prepared as a carrier for Human papillomavirus type 16 HPV-16) E7 gene and their gene transfection ability were evaluated in vitro. The plasmid expressing green fluorescent protein (pEGFP) was used as a reporter gene. Gel electrophoresis demonstrated full binding of CS NPs with the pDNA. The transfection of CS-pEGFP NPs was efficient in CHO cells and the expression of green fluorescent proteins was well observed. The expression of E7 proteins was confirmed under SDS-PAGE and western blot analysis. As a conclusion CS NPs may serve as an effective nonviral carrier for delivery of nucleotides into eukaryotic cells.
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- "A potential alternative to induce oral tolerance in which the antigen is directly expressed within the intestine could be the use of non-viral gene delivery systems encoding for the antigen. Chitosan, a non-toxic biodegradable polycationic polymer with low immunogenicity, was shown to be a useful oral gene carrier   . Chitosan can be complexed with plasmid- DNA, forming nanoparticles which are stable during the gastrointestinal tract, phagocytized in the gut and causes gene expression. "
ABSTRACT: One promising approach for the induction of transplant tolerance is the pre-treatment of transplant recipients with donor MHC-alloantigen. Our study focuses on the oral delivery of MHC-antigen encoding genes via chitosan-DNA nanoparticles to modulate the alloimmune response in order to reduce the development of transplant arteriosclerosis, the hallmark feature of chronic rejection after heart transplantation. Therefore, we performed fully allogeneic mouse abdominal aortic transplantats using C57BL/6 (H2(b)) mice as donors and CBA.J (H2(k)) mice as recipients. Aortic grafts were analyzed by histology and morphometry on day 30 after transplantation, levels of circulating alloantibodies were detected by FACS analysis. Pre-treatment of recipient mice with chitosan-DNA nanoparticles encoding for K(b), one of the MHC-I molecules of the donor, resulted in a significant reduction of intimal proliferation compared to untreated controls. When Ovalbumin was fed instead of K(b) encoding nanoparticles (K(b)-NP) or Balb/c (H2(d)) grafts were used instead of C57BL/6 (H2(b)) grafts as antigen controls, both groups showed no reduction of intimal thickness indicating an antigen-specific mechanism. In addition, analysis of peripheral blood of the transplanted mice showed significant suppression of alloantibody formation in the K(b)-NP fed group compared to all other allogeneic transplanted groups suggesting modulation of the humoral immune response. These results demonstrate the potential of chitosan-DNA nanoparticles to induce K(b)-specific tolerance and to reduce the development of transplant arteriosclerosis.Transplant Immunology 12/2012; 28(1). DOI:10.1016/j.trim.2012.11.007 · 1.83 Impact Factor
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- "Chitosan and its derivatives have been examined extensively for medical and pharmaceutical applications especially in artificial organs, targeted drug delivery, drug transport, protein delivery gene transfer and so on (Lavertu et al., 2006; Pan et al., 2002; Sakai et al., 2001; Wei and Zhang, 2007). Due to the presence of amino groups, CS possess positive charge and can effectively bind negatively charged DNA resulting in nanoparticles of various sizes (Bozkir and Saka, 2004b; Ishii et al., 2001) and also protect DNA from nuclease degradation (MacLaughlin et al., 1998; Mao et al., 2001; Richardson et al., 1999). CS has favorable biocompatibility characteristics as well as the ability to increase membrane permeability, both in vitro and in vivo and can be degraded by lysozyme in serum (Aspden et al., 1996; Takeuchi et al., 2001). "
ABSTRACT: Keywords: CS/TPP nanoparticle pVAOMP DNA In vitro/In vivo gene expression Immunohistochemistry Real-time PCR This work examines the potential use of chitosan tripolyphosphate (CS/TPP) nanoparticles for gene delivery in different tissues of fish through oral route. The porin gene of Vibrio anguillarum was used to construct a DNA vaccine using pcDNA 3.1, a eukaryotic expression vector and the construct was named as pVAOMP. The CS/TPP nanoparticles were synthesized by ionic gelation process and these nanoparticles particles were characterized. The morphology and particle size measurements of the nanoparticles were studied by field emission scanning electron microscopy (FE-SEM) and FTIR (Fourier Transform Infrared Spectra). The pVAOMP was encapsulated by CS/TPP nanoparticles by mixing of equal volume of heated CS/TPP and pVAOMP DNA solutions. The encapsulation efficiency of CS/TPP nanoparticles was found to be 79.9% of DNA binding with CS/TPP nanoparticles. The stability of plasmid DNA was also determined after encapsulation using DNase I and chitosanase. The cytotoxicity of CS/TPP nanoparticles was evaluated by MTT assay using fish cell line. The expression of gene was confirmed by immunohistochemistry, ELISA and real-time PCR analyses. The results indicate that DNA can be easily delivered into fish by feeding with CS/TPP nanoparticles.Aquaculture 07/2012; 358(358-59):14-22. DOI:10.1016/j.aquaculture.2012.06.012 · 1.83 Impact Factor