Acceleration of gene transfection efficiency in neuroblastoma cells through polyethyleneimine/poly(methyl methacrylate) core-shell magnetic nanoparticles.
BACKGROUND
The purpose of this study was to demonstrate the potential of magnetic poly(methyl methacrylate) (PMMA) core/polyethyleneimine (PEI) shell (mag-PEI) nanoparticles, which possess high saturation magnetization for gene delivery. By using mag-PEI nanoparticles as a gene carrier, this study focused on evaluation of transfection efficiency under magnetic induction. The potential role of this newly synthesized nanosphere for therapeutic delivery of the tryptophan hydroxylase-2 (TPH-2) gene was also investigated in cultured neuronal LAN-5 cells.


METHODS
The mag-PEI nanoparticles were prepared by one-step emulsifier-free emulsion polymerization, generating highly loaded and monodispersed magnetic polymeric nanoparticles bearing an amine group. The physicochemical properties of the mag-PEI nanoparticles and DNA-bound mag-PEI nanoparticles were investigated using the gel retardation assay, atomic force microscopy, and zeta size measurements. The gene transfection efficiencies of mag-PEI nanoparticles were evaluated at different transfection times. Confocal laser scanning microscopy confirmed intracellular uptake of the magnetoplex. The optimal conditions for transfection of TPH-2 were selected for therapeutic gene transfection. We isolated the TPH-2 gene from the total RNA of the human medulla oblongata and cloned it into an expression vector. The plasmid containing TPH-2 was subsequently bound onto the surfaces of the mag-PEI nanoparticles via electrostatic interaction. Finally, the mag-PEI nanoparticle magnetoplex was delivered into LAN-5 cells. Reverse-transcriptase polymerase chain reaction was performed to evaluate TPH-2 expression in a quantitative manner.


RESULTS
The study demonstrated the role of newly synthesized high-magnetization mag-PEI nanoparticles for gene transfection in vitro. The expression signals of a model gene, luciferase, and a therapeutic gene, TPH-2, were enhanced under magnetic-assisted transfection. An in vitro study in neuronal cells confirmed that using mag-PEI nanoparticles as a DNA carrier for gene delivery provided high transfection efficiency with low cytotoxicity.


CONCLUSIONS
The mag-PEI nanoparticle is a promising alternative gene transfection reagent due to its ease of use, effectiveness, and low cellular toxicity. The mag-PEI nanoparticle is not only practical for gene transfection in cultured neuronal cells but may also be suitable for transfection in other cells as well.
Center for Excellence in Omics-Nano Medical Technology Development Project, Chulalongkorn University, Bangkok, Thailand.
Tewin
T
Tencomnao
Kewalin
K
Klangthong
Nuttaporn
N
Pimpha
Saowaluk
S
Chaleawlert-Umpon
Somsak
S
Saesoo
Noppawan
N
Woramongkolchai
Nattika
N
Saengkrit
D045744:Cell Line, Tumor; D002470:Cell Survival; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007097:Imines / Q000737:chemistry*; D058185:Magnetite Nanoparticles / Q000737:chemistry*; D008526:Medulla Oblongata / Q000737:chemistry / Q000201:enzymology; D009447:Neuroblastoma / Q000737:chemistry / Q000235:genetics* / Q000628:therapy*; D010316:Particle Size; D011095:Polyethylenes / Q000737:chemistry*; D019904:Polymethyl Methacrylate / Q000737:chemistry*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection / Q000379:methods*; D014365:Tryptophan Hydroxylase / Q000235:genetics / Q000378:metabolism
D007097:Imines; D058185:Magnetite Nanoparticles; D011095:Polyethylenes; C505405:poly(ethylene imine); D019904:Polymethyl Methacrylate; C494784:TPH2 protein, human; D014365:Tryptophan Hydroxylase