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Down‐regulated lysosomal processing improved pegylated lipopolyplex‐mediated gene transfection
Author(s) -
Bai Jing,
Liu Yujie,
Sun Wenqiang,
Chen Jian,
Miller Andrew D.,
Xu Yuhong
Publication year - 2013
Publication title -
the journal of gene medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.689
H-Index - 91
eISSN - 1521-2254
pISSN - 1099-498X
DOI - 10.1002/jgm.2707
Subject(s) - transfection , microbiology and biotechnology , intracellular , chemistry , gene delivery , organelle , cationic liposome , flow cytometry , liposome , cell culture , fluorescence microscope , confocal microscopy , biophysics , biology , biochemistry , gene , fluorescence , genetics , physics , quantum mechanics
Abstract Background Nonviral lipid‐based gene delivery vectors have been shown to possess better stability and a longer circulation time after surface poly(ethylene glycol) PEG modification. However, surface PEGylation may decrease the transfection efficiency dramatically. In the present study, we addressed the hypothesis that down‐regulating lysosomal processing with a clinical available proton pump inhibitor omeprazole might decrease the sequestration of PEGylated Lipid‐Mu‐DNA (LMD) in intracellular organelles, thereby increasing their transfection efficiencies. Methods LMD nanoparticles were prepared by the self‐assembling of cationic liposomes, peptide Mu and plasmid DNA. The characteristics of LMD lipopolyplexes were detected by scanning electron microscopy, photon correlation spectroscopy and DNA gel electrophoresis. They were added to cultured cells with or without omeprazole pretreatment. The detailed cellular uptake and subcellular distribution were followed by flow cytometry and confocal microscopy. Gene transfection efficiencies were evaluated in four cell lines, as well as in xenograft tumor models. Results Lysosome staining revealed that 0.1 mg/ml (290 μ M ) omeprazole raised the pH value of intracellular acidic organelles and induced alterations in the lysosomal compartments. Confocal microscopy showed that more gene materials distributed from intracellular organelles to cytoplasms with omeprazole treatment. A luciferase gene transfection assay showed that omeprazole (probably at a nontoxic concentration) increased PEGylated LMD transfection efficiency significantly in human (NCI‐H1299, HT‐1080 and A375) and mouse (4T1) cell lines, as well as in tumors of H1299 xenograft tumor models ( p < 0.05). Conclusions Omeprazole might be used as a helper to improve gene transfection efficiencies of some PEGylated gene delivery vectors both in vitro and in vivo . Copyright © 2013 John Wiley & Sons, Ltd.