Enhancement of magnetofection efficiency using chitosan coated superparamagnetic iron oxide nanoparticles and calf thymus DNA

https://doi.org/10.1016/j.colsurfb.2017.01.028Get rights and content

Highlights

  • Superparamagnetic chitosan-iron oxide nanoparticles (CS-MNPs) have been synthesized and characterized.

  • Cytotoxicity studies show that the DNA-CS-MNPs system is biocompatible.

  • The interaction between CS-MNPs and DNA in physiological buffer (pH 7.4) was studied.

  • The binding process was spontaneous and based on electrostatic interactions a static quenching.

Abstract

Superparamagnetic iron oxide nanoparticles (MNPs) were prepared and coated with chitosan (CS). The chitosan-magnetic iron oxide nanoparticles (CS-MNPs) were characterized using Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), vibrating sample magnetometry (VSM), and the morphology of the particles was studied by transmission electron microscopy (TEM). Our findings show that the magnetic particles were monodisperse (10 nm mean diameter) and exhibited superparamagnetic behavior. The interaction between the particles and calf-thymus DNA (DNA) in physiological buffer was studied with UV–vis, fluorescence and circular dichroism spectroscopy and zeta potential. Spectroscopic studies were indicated DNA conformational changes in the presence of CS-MNPs. Binding and thermodynamic parameters at different temperatures were calculated using the Stern–Volmer, Hill, Scatchard and Van’t Hoff equations. The binding process was spontaneous and interactions were electrostatic with the appropriate binding constant (Kb = 4.52 × 103 M−1, 3.69 × 103 M−1 and 3.02 × 103 M−1 at 300 K, 310 K and 320 K, respectively). Zeta potential measurements of DNA continually increased with the addition of CS-MNPs, supporting our thermodynamic findings. Moreover, CS-MNPs were able to quench the fluorescence of DNA-intercalated ethidium bromide (DNA-EB) by a static quenching mechanism. Cytotoxicity studies show that the DNA-CS-MNP system is biocompatible with a human foreskin fibroblast cell line, HFFF2. Collectively, these results suggest that surface cationic magnetic chitosan-iron oxide nanoparticles can potentially enhance magnetofection efficiency.

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The interaction between biocompatible chitosan-magnetic nanoparticles (CS-MNPs) and calf thymus DNA (DNA) in physiological buffer (pH 7.4) was investigated.

Introduction

Gene therapy is one of several approaches used to treat incurable diseases and genetic disorders. The success of this approach requires the development of innovative gene delivery vectors [1], [2], [3]. Research efforts have focused on the development of safe and effective viral and non-viral vector systems. Although viral vectors have high transduction efficiency, they are plagued with issues such as non-specificity, immunogenicity to target cells, toxicity and enzymatic degradation [2], [4]. Non-viral delivery systems are being pursued to facilitate therapeutic gene transfer in the clinic. These carriers, such as cationic lipids and polymers, typically interact with anionic DNA via charged moieties, condensing the long, string- like DNA molecules into compact, nano-sized particles that are suitable for cellular uptake. A range of cationic lipids and polymers have been engineered to accomplish this function effectively and intelligent carriers are continually being optimized in order to control the intracellular fate of DNA molecules [5], [6].

DNA/polymer complexes with cationic polymers have a number of distinct advantages that make them suitable for gene delivery. These biocompatible complexes are more stable than cationic lipids, have low immunogenicity and minimal cytotoxicity, making these polymers a good alternative to viral- and lipid- mediated delivery methods. Cationic polymers have been used to condense and deliver DNA both in vitro and in vivo. The structural variability and versatility of cationic polymers and the possibility of covalent binding to targeting moieties to mediate gene expression through specific receptors presents interesting possibilities [7], [8], [9], [10], [11].

Chitosan (CS) is an inexpensive, biocompatible material with minimal toxicity and high cationic potential that has been widely used for gene delivery. High molecular weight chitosan (100–400 kDa) can form extremely stable polyplexes with DNA [1], [12], [13]. However, low transfection efficiencies (as compared to viral vectors) and the inability to target specific cells or tissues in vivo remain problematic. In order to overcome these difficulties, magnetofection was developed to enhance the delivery of nucleic acids associated with magnetic nanoparticles [9], [14], [15]. Magnetofection is a novel and highly efficient method of transfecting cells in culture [16].

In the present study, superparamagnetic iron oxide nanoparticles coated with chitosan were prepared and characterized by FT-IR spectroscopy, X-ray diffraction, VSM and TEM. We studied the interaction between CS-MNPs and DNA by multi-spectroscopic methods under physiological conditions. These surface cationic magnetic chitosan-iron oxide nanoparticles can potentially enhance magnetofection efficiency to potentially improve gene delivery systems.

Section snippets

Materials and methods

Ferrous chloride tetrahydrate (FeCl2.4H2O), ferric chloride hexahydrate (FeCl3.6H2O), ammonium hydroxide (NH4OH), sodium hydroxide (NaOH), acetic acid and Tris (hydroxymethyl) amino methane hydrocholoride (Tris-HCl) were obtained from Merck Chemical Co. Ethidium bromide (EB), chitosan (CS) and calf thymus DNA (DNA) were purchased from Sigma Chemical Co. Cell lines used in cytotoxicity studies were obtained from the Cell Bank of the Pasteur Institute (Tehran, Iran). All chemicals were of

Characterization of magnetic nanoparticles and chitosan-MNPs

Magnetic nanoparticles and chitosan-magnetic nanoparticles were prepared as described (Section 2.3.). Fig. S1 shows the XRD pattern of CS-MNPs synthesized by co-precipitation. The Rietveld refinement of the XRD pattern was used to calculate the average grain size from the broadening of the XRD peaks of CS-MNPs. The average grain size of the chitosan coated magnetic nanoparticles is 15 nm, which is in line with previous studies [30], [31]. To confirm the structure of the samples, the FT-IR

Conclusions

We describe the formulation of superparamagnetic chitosan iron oxide nanoparticles for magnetofection and characterized these nanoparticles using Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), vibrating sample magnetometry (VSM) and transmission electron microscopy (TEM). Particle size was calculated by XRD and TEM with similar results. The superparamagnetic properties of CS-MNPs significantly increased the transfection efficiency. Studying DNA-CS-MNPs in human

Acknowledgements

We are grateful for financial support from the Materials and Energy Research Center. We thank Dr. Roya Pedram Fatemi for careful reading of the manuscript.

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