Hydroxyapatite nanoparticles modified by branched polyethylenimine are effective non-viral vectors for siRNA transfection of hepatoma cells in vitro.

Abstract

Small interfering RNA (siRNA) technology is a powerful tool in biomedical research and holds great potential for RNA interference-based therapies for HIV, hepatitis and cancer. However, the absence of a safe and efficient method for the delivery of siRNA has become a bottleneck for their development. Nanocrystallized hydroxyapatite (nHAP) appears to be an optimal candidate non-viral gene vector for several reasons, including its good biocompatibility and ease of production, however, nHAP microemulsions cannot remain monodispersed for long periods of time. Due to their high surface energy, nHAP particles gradually aggregate into large ones that are difficult for the cell to take up. To overcome this we modified nHAP with polyethylenimine (PEI) to generate a compound (MnHAP) with a tight size-distribution of <200 nm. The positive surface potential of MnHAP inhibited particle aggregation and thus made it easier to conjugate more siRNA. The transfection efficiency of MnHAP/fluorescent FAM-labeled siRNA complex was tested using flow cytometry, and the transfected cells were observed using fluorescence microscopy. The cytotoxicity of MnHAP/siRNA complexes to the human liver cancer cell line BEL-7402 was assessed in vitro by a formazan dye assay. Our results show that the in vitro transfection efficiency of MnHAP/siRNA was equivalent to that of the commercially available transfection agent Lipofectamine® 2000, but with decreased cytotoxicity. The MnHAP nanoparticles were also able to deliver siRNA for silencing of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in BEL-7402 cells, which supports that MnHAP might be a promising non-viral vector for biomedical research and gene delivery.