Abstract
It has been previously shown that transfection activity of cationic liposome/DNA lipoplexes delivered systemically is drastically inhibited by lipoproteins (Tandia, B. M., Vandenbranden, M., Wattiez, R., Lakhdar, Z., Ruysschaert, J. M., and Elouahabi, A. (2003) Mol Ther. 8, 264-273). In this work, we have compared the binding/uptake and transfection activities of DOTAP (N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride) and diC14-amidine (3-tetradecylamino-N-tert-butyl-N'-tetra-decylpropionamidine)-containing lipoplexes in the presence or absence of purified low density lipoproteins and high density lipoprotein. Binding/uptake of both lipoplexes by the mouse lung endothelial cell line was inhibited to a similar extent in the presence of lipoproteins. In contrast, transfection activity of diC14-amidine-containing lipoplexes was almost completely inhibited (approximately by 95%), whereas approximately 40% transfection activity of DOTAP-containing lipoplexes was preserved in the presence of lipoproteins. Interestingly, the ability of lipoproteins to inhibit the transfection efficiency of lipoplexes was well correlated with their ability to undergo lipid mixing with the cationic lipid bilayer as revealed by fluorescence resonance energy transfer assay. Incubation of lipoplexes with increased doses of lipoproteins resulted in enhanced lipid mixing and reduced transfection activity of the lipoplexes in mouse lung endothelial cells. The role of lipid mixing in transfection was further demonstrated using lipid-mixing inhibitor, lyso-phosphatidylcholine, or activator (dioleoylphosphatidylethanolamine). Incorporation of Lyso-PC into diC14-amidine-containing lipoplexes completely abolished their capacity to undergo lipid mixing with lipoproteins and allowed them to reach a high transfection efficiency in the presence of lipoproteins. On the other hand, the incorporation of dioleoylphosphatidylethanolamine into DOTAP/DNA lipoplex activated lipid mixing with the lipoproteins and was shown to be detrimental toward the transfection activity of these lipoplexes. Taken together, these results indicate that fusion of lipoplexes with lipoproteins is a limiting factor for in vivo transfection.
Highlights
¶ To whom correspondence should be addressed: Laboratory of Structure and Function of Biological Membranes, CP 206/2, Campus PlaineUniversite Libre de Bruxelles, Blvd. du Triomphe, 1050 Brussels, Belgium
Plasma Lipoprotein Particles, LDL and HDL, Inhibit the in Vivo Transfection Efficiency of Lipoplexes—Previous reports have shown that lipoproteins, including LDL and HDL, inhibit the in vitro transfection efficiency of diC14-amidine/DNA lipoplexes in mouse lung endothelial cell line (MLE) [11]
We describe molecular interactions between lipoplexes and plasma lipoproteins that lead to inhibition of the transfection activity of lipoplexes
Summary
Lipoprotein particles including low density lipoproteins (LDL) and high density lipoproteins (HDL) are among the plasma components that bind to the lipoplexes. Such a binding was shown to inhibit the transfection efficiency in cultured cells [11]. Binding/uptake by the cells of both lipoplexes was inhibited to the same extent, the transfection efficiency was affected differently depending on the cationic lipid used. This suggested that other factors were involved. Comparison with transfection data shows a good inverse correlation between lipid mixing and transfection, suggesting that this process is a limiting step for cationic lipid-mediated intravenous gene transfer and expression
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