Preparation and characterisation of CTAB and chitosan modified PLGA nanoparticles containing plasmid DNA and nigella sativa oil for alzheimer disease

Abstract

Background: Alzheimer’s disease (AD) is spreading in both developed and developing countries with no comprehensive treatment. Gene therapy provides new treatment strategies where it can interfere the disease on its genetic levels. Viral gene therapy is currently in clinical trial stage but it is facing problems due to safety issues. Nigella sativa is an herbal and Prophet medicine having numerous medical effects in AD and other diseases. CTAB and chitosan are frequently used in the preparation of nanoparticle for non-viral gene therapy. Objective: To co-encapsulate NSO with pDNA in one gene delivery system based on PLGA nanoparticles for the treatment of AD. Methodology: We fabricated PLGA microspheres loaded with pDNA using double-emulsion solvent-evaporation method and investigated the effect of surfactants and PLGA. N. sativa oil (NSO) was encapsulated in PLGA microparticle and its cell uptake was evaluated using PC-12 cell line. Two approaches were applied in coencapsulating pDNA with NSO in one PLGA nanoparticle system i.e. adsorption on pre-prepared cationic nanoparticles and encapsulation within nanoparticles after being complexed with CTAB. CTAB and chitosan-modified PLGA nanoparticles were fabricated and then incorporated with pDNA. NSO was co-encapsulated with pDNA either by adsorption on chitosan-modified nanoparticles or by encapsulation. NSO encapsulation efficiency was evaluated using FTIR. The neuroregenerative effect of the encapsulated NSO was evaluated on N2a cells. Results and discussion: Tween blend of HLB-16 revealed the highest supercoil preservation index. Span surfactant and carboxyl terminal low molecular weight PLGA, Tween blend of HLB-16, Triton X-100 revealed exhibited high, moderate and low burst release, respectively. NSOloaded PLGA microparticles were taken up mainly by mitotic cells in a similar efficiency to Span-modified microparticles but less than Tween 80-modified microparticles. CTAB-modified PLGA nanoparticles were negatively charged while chitosan-PLGA nanoparticles were positively charged. Chitosan-PLGA nanoparticles were taken up by N2a cells higher than other formulations and exhibited good transfection efficiency after being loaded with pDNA. Meanwhile, CTAB-pDNAPLGA nanoparticles showed the highest transfection efficiency with low burst release. The developed FTIR method exhibited linearity in the range of NSO/PLGA of 5- 150%. Both NSO and pDNA were encapsulated in PLGA nanoparticles. NSO encapsulation efficiency was lower in chitosan-modified nanoparticles. The encapsulated pDNA and NSO exhibited the desirable effects in N2a cells. The encapsulated pDNA exhibited the ex-gene expression of green fluorescent protein whereas the encapsulated NSO exhibited neurite outgrowth effect. Conclusion: NSO was successfully co-encapsulated with pDNA in PLGA nanoparticles as non-viral gene delivery system. The gene expression from the encapsulated pDNA, the encapsulated NSO was able to exhibit neuroregeneration in N2a cells. CTAB-pDNANSO- PLGA nanoparticles revealed the best results with slower pDNA release and higher neurite outgrowth effect.