The role of dioleoyl phosphatidylethanolamine in cationic liposome mediated gene transfer.

In a reported gene assay, cationic liposomes containing the cationic lipid 3 beta-(N-(N',N'-dimethylaminoethane)carbamoyl)cholesterol (DC-Chol) and a neural phospholipid dioleoylphosphatidylethanolamine (DOPE) showed high transfection activity. DNA/liposome complex which contained low amount of liposomes could bind to the cell surface but failed to transfect the cells. We have designed a two-step protocol to examine this phenomenon in more detail. A431 human cells were incubated on ice (pulse) with DNA complexed to a low level of cationic liposomes. The cells were washed and incubated at 37 degrees C (chase) with or without free cationic liposomes of various composition (helper liposomes). Only liposomes enriched with DOPE showed helper activity; liposomes containing dioleoylphosphatidylcholine (DOPC), a structural analog of DOPE, had no helper activity. The delivery was inhibited by the lysosomotropic agent chloroquine and was optimal if the helper liposome chase was initiated immediately after the pulse. An endocytosis model of DNA delivery by cationic liposomes is proposed in which the principal function of the chase liposomes is to destabilize the endosome membrane and allow the release of DNA into the cytosol. This model is consistent with the known activity of DOPE to assume non-bilayer structures, hence destabilizing the endosome membrane.

[1]  I. Pastan,et al.  Evidence that the penton base of adenovirus is involved in potentiation of toxicity of Pseudomonas exotoxin conjugated to epidermal growth factor , 1984, Molecular and cellular biology.

[2]  L. Huang,et al.  A novel cationic liposome reagent for efficient transfection of mammalian cells. , 1991, Biochemical and biophysical research communications.

[3]  J. German,et al.  Liposomes as agents of DNA transfer. , 1993, Biochimica et biophysica acta.

[4]  M. Cotten,et al.  Adenovirus enhancement of transferrin-polylysine-mediated gene delivery. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[5]  R. Epand,et al.  Effect of cationic cholesterol derivatives on gene transfer and protein kinase C activity. , 1992, Biochimica et biophysica acta.

[6]  R. Kumar,et al.  Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations. , 1994, The Journal of biological chemistry.

[7]  J. Behr,et al.  Efficient gene transfer into mammalian primary endocrine cells with lipopolyamine-coated DNA. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[8]  I. Pastan,et al.  Role of a low-pH environment in adenovirus enhancement of the toxicity of a Pseudomonas exotoxin-epidermal growth factor conjugate , 1984, Journal of virology.

[9]  L. Huang,et al.  Phosphatidylethanolamine liposomes: drug delivery, gene transfer and immunodiagnostic applications. , 1992, Biochimica et biophysica acta.

[10]  I Mellman,et al.  Acidification of the endocytic and exocytic pathways. , 1986, Annual review of biochemistry.

[11]  Leaf Huang,et al.  CATIONIC LIPOSOMES AND POLYMERS FOR GENE TRANSFER , 1993 .

[12]  U. Svensson,et al.  Entry of adenovirus 2 into HeLa cells , 1984, Journal of virology.

[13]  K Mechtler,et al.  The influence of endosome-disruptive peptides on gene transfer using synthetic virus-like gene transfer systems. , 1994, The Journal of biological chemistry.

[14]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[15]  L. Huang,et al.  DNA transfection mediated by cationic liposomes containing lipopolylysine: characterization and mechanism of action. , 1994, Biochimica et biophysica acta.