Abstract
Currently, gold nanoparticles have found applications in engineering and medical sciences, taking advantage from their properties and characteristics. Surface plasmon resonance, for instance, is one of the main features for optical applications and other physical properties, like high density, that represents the key for cellular uptake. Among other applications, in the medical field, some diseases may be treated by using gene therapy, including monogenetic or polygenetic disorders and infections. Gene adding, suppression, or substitution is one of the many options for genetic manipulation. This work explores an alternative non-viral method for gene transfer by using gold nanoparticles functionalized with organic polymers; two routes of synthesis were used: one of them with sodium borohydride as reducing agent and the other one with chitosan oligosaccharide as reducing and stabilizing agent. Gold nanoparticles conjugated with chitosan, acylated chitosan and chitosan oligosaccharide, were used to evaluate transfection efficiency of plasmid DNA into cell culture (HEK-293). Physical and chemical properties of gold nanocomposites were characterized by using UV-Vis Spectroscopy, ξ-potential, and transmission electron microscopy. Furthermore, the interaction between gold nanoparticles and plasmid DNA was demonstrated by using agarose gel electrophoresis. Transfection tests were performed and evaluated by β-galactosidase activity and green fluorescence protein expression. The percentage of transfection obtained with chitosan, acylated chitosan, and chitosan oligosaccharide were of 27%, 33%, and 60% respectively.
Highlights
Gene therapy can be briefly defined as a way of introducing genetic material into cells for different purposes, the treatment of genetic diseases among them [1]
Nanocomposite Synthesis and Characterization Among the three kinds of complexes synthesized in this work, Chitosan oligosaccharide (COS)@n-Gold nanoparticles (AuNPs) deserves special attention since they were synthesized without the use of toxic reagents, Fig. 4 Stability of gold nanoparticle nanocomposites by UV-Vis absorption spectra obtained 150 days after synthesis: chemically synthesized CO-AuNPs (a) and Acylated chitosan (Acyl-CO)-AuNPs (b) and green/ one-pot synthesis for COS@2-AuNPs (c)
The highest cell viability was obtained with COS@1-AuNPs and COS@2AuNPs, with 93.53% and 93.46%, respectively. It is Conclusions Chitosan, acylated chitosan, and chitosan oligosaccharide nanocomposites with gold nanoparticles, featuring positive charges, were synthesized. They presented good stability in colloidal solution; this confirms the viable use of chitosan as a stabilizer and chitosan oligosaccharide as both reducing agent and stabilizer, and positive charges by its amino groups improved affinity with plasmid
Summary
Gene therapy can be briefly defined as a way of introducing genetic material into cells for different purposes, the treatment of genetic diseases among them [1]. Chitosan solubility can be modified changing the distribution of remaining acetyl groups in the chain and the degree of acetylation [13]. Another method to control the physical properties of chitosan is the acylation (Fig. 1b). Bhattarai et al [14] obtained N-acylated chitosan stabilized gold nanoparticles for applications in physiological conditions, exhibiting the advantages of non-acylated chitosan. Another useful modification of chitosan is the reduction of the polymer chain length (Chitosan oligosaccharide, COS) [15]. Synthesis of gold nanoparticles with chitosan oligosaccharide as reducing agent does not include any toxic reagent [20]
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