title: Phase behavior, DNA ordering, and size instability of cationic lipoplexes. Relevance to optimal transfection activity.
abstract: Mechanisms of cationic lipid-based nucleic acid delivery are receiving increasing attention, but despite this the factors that determine high or low activity of lipoplexes are poorly understood. This study is focused on the fine structure of cationic lipid-DNA complexes (lipoplexes) and its relevance to transfection efficiency. Monocationic (N-(1-(2,3-dioleoyloxy)propyl),N,N,N-trimethylammonium chloride, N-(1-(2,3-dimyristyloxypropyl)-N,N-dimethyl-(2-hydroxyethyl)ammonium bromide) and polycationic (2,3-dioleyloxy-N-[2(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanammonium trifluoroacetate) lipid-based assemblies, with or without neutral lipid (1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine, 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine, cholesterol) were used to prepare lipoplexes of different L(+)/DNA(-) charge ratios. Circular dichroism, cryogenic-transmission electron microscopy, and static light scattering were used for lipoplex characterization, whereas expression of human growth hormone or green fluorescent protein was used to quantify transfection efficiency. All monocationic lipids in the presence of inverted hexagonal phase-promoting helper lipids (1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine, cholesterol) induced appearance of Psi(-) DNA, a chiral tertiary DNA structure. The formation of Psi(-) DNA was also dependent on cationic lipid-DNA charge ratio. On the other hand, monocationic lipids either alone or with 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine as helper lipid, or polycationic 2,3-dioleyloxy-N-[2(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanammonium trifluoroacetate-based assemblies, neither of which promotes a lipid-DNA hexagonal phase, did not induce the formation of Psi(-) DNA. Parallel transfection studies reveal that the size and phase instability of the lipoplexes, and not the formation of Psi(-) DNA structure, correlate with optimal transfection.
authors.affiliation: Laboratory of Membrane and Liposome Research, Department of Biochemistry, Hebrew University-Hadassah Medical School, P.O. Box 12272, Jerusalem 91120, Israel.
authors.forename: D
authors.initials: D
authors.lastname: Simberg
authors.forename: D
authors.initials: D
authors.lastname: Danino
authors.forename: Y
authors.initials: Y
authors.lastname: Talmon
authors.forename: A
authors.initials: A
authors.lastname: Minsky
authors.forename: M E
authors.initials: ME
authors.lastname: Ferrari
authors.forename: C J
authors.initials: CJ
authors.lastname: Wheeler
authors.forename: Y
authors.initials: Y
authors.lastname: Barenholz
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002412:Cations; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D020404:Glycerophospholipids / Q000737:chemistry; D049452:Green Fluorescent Proteins; D006801:Humans; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry*; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008854:Microscopy, Electron; D008958:Models, Molecular; D010713:Phosphatidylcholines / Q000737:chemistry; D010714:Phosphatidylethanolamines; D010957:Plasmids / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D013096:Spermine / Q000031:analogs & derivatives* / Q000737:chemistry; D013997:Time Factors; D014162:Transfection; D014466:Ultraviolet Rays
chemical_list: C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; C098223:2,3-dioleyloxy-N-(2-(sperminecarboxamido)ethyl)-N,N-dimethyl-1-propanaminium trifluoroacetate; D002412:Cations; D005229:Fatty Acids, Monounsaturated; D005456:Fluorescent Dyes; D020404:Glycerophospholipids; D008055:Lipids; D008081:Liposomes; D008164:Luminescent Proteins; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins; C087276:(3-dimyristyloxypropyl)(dimethyl)(hydroxyethyl)ammonium; D013096:Spermine; D004247:DNA; C017251:1,2-oleoylphosphatidylcholine; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane