Intracellular Delivery of siRNA by Polycationic Superparamagnetic Nanoparticles

Betzaida Castillo,T Alan Hatton,Valerie Badillo,Gabriel Barletta,Carlos I González,Xaira López,Jose A González Feliciano,Lev Bromberg

doi:10.1155/2012/218940

Abstract

The siRNA transfection efficiency of nanoparticles (NPs), composed of a superparamagnetic iron oxide core modified with polycationic polymers (poly(hexamethylene biguanide) or branched polyethyleneimine), were studied in CHO-K1 and HeLa cell lines. Both NPs demonstrated to be good siRNA transfection vehicles, but unmodified branched polyethyleneimine (25 kD) was superior on both cell lines. However, application of an external magnetic field during transfection (magnetofection) increased the efficiency of the superparamagnetic NPs. Furthermore, our results reveal that these NPs are less toxic towards CHO-K1 cell lines than the unmodified polycationic-branched polyethyleneimine (PEI). In general, the external magnetic field did not alter the cell's viability nor it disrupted the cell membranes, except for the poly(hexamethylene biguanide)-modified NP, where it was observed that in CHO-K1 cells application of the external magnetic field promoted membrane damage. This paper presents new polycationic superparamagnetic NPs as promising transfection vehicles for siRNA and demonstrates the advantages of magnetofection.

Highlights

Small interfering RNA’s are short double-stranded nucleic acids, commonly containing 19–21 residues and 3 dinucleotide overhangs, which are widely used as synthetic reagents to reduce gene expression of target RNA in cells [1] and prevent the synthesis of specific proteins [2]. siRNAs are being developed to target therapeutically important genes involved in cancer, viral infections, autoimmune and neurodegenerative diseases [3]
At low N/P ratios, magnetofection improved the transfection efficiency of poly(ethylene imine) (PEI)-M/SiO2-siRNA complex in CHO-K1 cells over PEI (PEIN/P: 8 = 91%; PEI-M/SiO2 N/P: 3 = 45%; PEIM/SiO2-magnetofection N/P: 3 = 95%—Figure 1(a))
For HeLa cells magnetofection improved the transfection efficiency of PEI-M/SiO2 at low N/P ratios, the transfection efficiency was still less than that observed with PEI, but when N/P ratios higher than 34 were used, magnetofection did improve the transfection efficiency of PEI-M/SiO2 over PEI slightly (PEIN/P: 155 = 94%; PEI-M/SiO2 N/P: 68 = 93%; PEIM/SiO2-magnetofection N/P: 68 = 98%, Figure 1(b))

Summary

Introduction

Small interfering RNA’s (siRNAs) are short double-stranded nucleic acids, commonly containing 19–21 residues and 3 dinucleotide overhangs, which are widely used as synthetic reagents to reduce gene expression of target RNA in cells [1] and prevent the synthesis of specific proteins [2]. siRNAs are being developed to target therapeutically important genes involved in cancer, viral infections, autoimmune and neurodegenerative diseases [3]. SiRNAs are being developed to target therapeutically important genes involved in cancer, viral infections, autoimmune and neurodegenerative diseases [3]. These short doublestranded nucleic acids are unstable within the extracellular environment, they cannot cross cell membranes and due to their small size are readily secreted by the renal system [2, 4]. Several studies have demonstrated that magnetofection can efficiently deliver siRNA to living cells cultivated in vitro [14,15,16], and it appears to be a reliable and gentle method for siRNA and DNA delivery into difficult to transfect cells such as mammalian fibroblasts [17].

Methods

Results

Conclusion