A new cationic liposome for efficient gene delivery with serum into cultured human cells: a quantitative analysis using two independent fluorescent probes.
Cationic liposomes are useful to transfer genes into eukaryotic cells in vitro and in vivo. However, liposomes with good transfection efficiency are often cytotoxic, and also require serum-free conditions for optimal activity. In this report, we describe a new formulation of cationic liposome containing DC-6-14, O,O'-ditetradecanoyl-N-(alpha-trimethylammonioacetyl)diethan olamine chloride, dioleoylphosphatidylethanolamine and cholesterol for gene delivery into cultured human cells. This liposome, dispersed in 5% serum-containing growth medium, efficiently delivered a plasmid DNA for GFP (green fluorescent protein) into more than 80% of the cultured human cell hybrids derived from HeLa cells and normal fibroblasts. Flow cytometric analysis revealed that the efficiency of the GFP gene expression was 40-50% in a tumor-suppressed cell hybrid, while it was greatly reduced in the tumorigenic counterpart. The enhanced GFP expression in tumor-suppressed cell hybrids was quantitatively well correlated with a prolonged presence of the plasmid DNA, which had been labeled with another fluorescent probe, ethidium monoazide, within the cells. These results suggest that a newly developed cationic liposome is useful for gene delivery in serum-containing medium into human cells and the stability of the plasmid DNA inside the cell is a crucial step in this liposome-mediated gene expression. The mechanisms by which cationic liposome mediates gene transfer into eukaryotic cells are also discussed.
Serikawa
T
T
Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Tokyo, Japan.
Suzuki
N
N
Kikuchi
H
H
Tanaka
K
K
Kitagawa
T
T
D002412:Cations; D003470:Culture Media; D004274:DNA, Recombinant / Q000008:administration & dosage / Q000235:genetics; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D015870:Gene Expression; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D006822:Hybrid Cells; D008081:Liposomes / Q000737:chemistry*; D008164:Luminescent Proteins / Q000235:genetics; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D000644:Quaternary Ammonium Compounds; D014162:Transfection
D002412:Cations; D003470:Culture Media; D004274:DNA, Recombinant; D005456:Fluorescent Dyes; D008081:Liposomes; D008164:Luminescent Proteins; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins