Multifunctional magnetic nanocarriers for delivery of siRNA and shRNA plasmid to mammalian cells: Characterization, adsorption and release behaviors

Nucleic acids are promising candidates for treating various diseases. Nucleic acid is negatively charged and hydrophilic; therefore, it is not efficiently taken up by cells. Successful gene therapy requires the development of carriers for efficient delivery of gene-expressing DNA plasmid and small interfering RNA (siRNA) duplex. In this study, we developed MNP-CA-PEI, a citric acid (CA)-modified magnetic nanoparticle (MNP) cross-linked with polyethyleneimine (PEI), using carbonyldiimidazole as the crosslinker. The physical properties of MNP-CA-PEI (particle size, morphologies, surface coating, surface potentials, magnetic hystereses, superparamagnetic be-haviors, and infrared spectra) were systematically characterized by transmission electron microscopy imaging, dynamic light scattering, thermogravimetric analysis, superconducting quantum interference device, and Fourier transform infrared spectroscopy. The adsorption isotherm and kinetics were determined by the Langmuir model, the Freundlich model, a pseudo-first-order equation, and a pseudo-second-order equation. MNP-CA-PEI could form polyelectrolyte complexes with negatively charged nucleic acids, enabling the efficient delivery of nucleic acids into cells. Using MNP-CA-PEI nanoparticles, we magnetically triggered the intracellular delivery of green fluorescence protein (GFP)-expressing DNA plasmid, plasmid-expressing short hairpin RNA (shRNA) against GFP, or siRNA targeting GFP into different cell lines. Nucleic acid/MNP-CA-PEI displayed the enhanced cellular up-take of GFP-expressing DNA plasmid, and it improved the silencing efficiency of shRNA and siRNA, determined by fluorescence imaging. Gene knockdowns mediated by shRNA and siRNA were also confirmed by a quanti-tative real-time polymerase chain reaction. MNP-CA-PEI delivered nucleic acids into cytosol through caveolae-mediated endocytosis. This study introduces a new MNP functionalization that can be used for the magneti-cally driven intracellular delivery of nucleic acids.

Automated summary

Nucleic acids have potential for treating diseases but face challenges in cellular uptake. This study developed a new carrier, MNP-CA-PEI, for efficient delivery of nucleic acids into cells, enhancing gene silencing efficiency. The nanoparticles could be magnetically triggered to deliver DNA plasmids and siRNA, improving gene knockdowns in different cell lines.