Hepatocyte-targeted psi RNA Delivery Mediated by Galactosylated Poly(Ethylene Glycol)-Graft-Polyethylenimine In Vitro

Gene silencing in liver disease could be achieved by delivering siRNA with nonviral vectors. However, the transfection efficiency of plasmid siRNA ( psiRNA) applied through this approach in hepatocytes is generally low. Based on the fact that the asialoglycoprotein receptors present on hepatocytes can recognize galactose, we synthesized galactosylated poly(ethylene glycol)-graft-polyethylenimine (Gal-PEG-PEI) as a nonviral psiRNA carrier for hepatocyte targeting. Our results indicate that 0.2% (molar percentage) of amine groups of PEI was conjugated with PEG having galactose on its distal end. Increasing the molar ratios of Gal-PEG-PEI to psiRNA in complexation led to a decrease in particle size but an increase in zeta potential of complexes. The transfection efficiency of nanocomplexes, that is, Gal-PEG-PEI/psiRNA, in HepG2 cell line depends on the N/P value, which reflects the molar ratio of Gal-PEG-PEI to psiRNA in the complex. The highest transfection efficiency was 37.34%, which was obtained at N/P 8. At the same N/P value, the transfection efficiency with the nontargeting PEG-PEI/ psiRNA or Lipofectamine 2000/ psiRNA was much lower. The transfection efficiency of Gal-PEG-PEI/ psiRNA dropped to 3.60% from 37.34% after an excessive amount of free galactose was added into the medium for HepG2 cell incubation. By contrast, the similar phenomenon was observed neither when using PEG-PEI or Lipofectamine 2000 as a delivery vector nor in human embryonic kidney 293 cell line lacking ASGR. Real-time PCR analysis and western blot assay demonstrate that the knockdown of HLA-E gene expression by psiRNA/Gal-PEG-PEI (N/P 8) can reach about 60% in HepG2 cells after a 48-h transfection.