TY - JOUR
T1 - Phase behavior, DNA ordering, and size instability of cationic lipoplexes
T2 - Relevance to optimal transfection activity
AU - Simberg, Dmitri
AU - Danino, Dganit
AU - Talmon, Yeshayahu
AU - Minsky, Abraham
AU - Ferrari, Marilyn E.
AU - Wheeler, Carl J.
AU - Barenholz, Yechezkel
PY - 2001/12/14
Y1 - 2001/12/14
N2 - 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 Ψ- DNA, a chiral tertiary DNA structure. The formation of Ψ- 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 Ψ- DNA. Parallel transfection studies reveal that the size and phase instability of the lipoplexes, and not the formation of Ψ- DNA structure, correlate with optimal transfection.
AB - 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 Ψ- DNA, a chiral tertiary DNA structure. The formation of Ψ- 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 Ψ- DNA. Parallel transfection studies reveal that the size and phase instability of the lipoplexes, and not the formation of Ψ- DNA structure, correlate with optimal transfection.
UR - http://www.scopus.com/inward/record.url?scp=0035861585&partnerID=8YFLogxK
U2 - 10.1074/jbc.M105588200
DO - 10.1074/jbc.M105588200
M3 - 文章
C2 - 11564736
AN - SCOPUS:0035861585
SN - 0021-9258
VL - 276
SP - 47453
EP - 47459
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 50
ER -