Physico-Chemical Characteristics of Lipoplexes Influence Cell Uptake Mechanisms and Transfection Efficacy |
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Authors: | Sarah Resina Paul Prevot Alain R Thierry |
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Institution: | 1. Laboratoire de Dynamique des Interactions Membranaires Normales et Pathologiques (DIMNP), Département de Défenses Antivirales et Antitumorales - UMR 5235 - Université de Montpellier II, Montpellier, France.; 2. Modélisation et Ingénierie des Systèmes Complexes Biologiques pour le Diagnostic (SysDiag) – UMR 3145 CNRS/Bio-Rad - Cap Delta, Montpellier, France.;University of Southampron, United Kingdom |
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Abstract: | BackgroundFormulation of DNA/cationic lipid complexes (lipoplexes) designed for nucleic acid delivery mostly results in positively charged particles which are thought to enter cells by endocytosis. We recently developed a lipoplex formulation called Neutraplex that allows preparation of both cationic and anionic stable complexes with similar lipid content and ultrastructure.Methodology/Principal FindingsTo assess whether the global net charge could influence cell uptake and activity of the transported oligonucleotides (ON), we prepared lipoplexes with positive and negative charges and compared: (i) their physicochemical properties by zeta potential analysis and dynamic light scattering, (ii) their cell uptake by fluorescence microscopy and flow cytometry, and (iii) the biological activity of the transported ON using a splicing correction assay. We show that positively or negatively charged lipoplexes enter cells cells using both temperature-dependent and -independent uptake mechanisms. Specifically, positively charged lipoplexes predominantly use a temperature-dependent transport when cells are incubated OptiMEM medium. Anionic lipoplexes favour an energy-independent transport and show higher ON activity than cationic lipoplexes in presence of serum. However, lipoplexes with high positive global net charge and OptiMEM medium give the highest uptake and ON activity levels.ConclusionsThese findings suggest that, in addition to endocytosis, lipoplexes may enter cell via a temperature-independent mechanism, which could be mediated by lipid mixing. Such characteristics might arise from the specific lipoplex ultrastructure and should be taken into consideration when developing lipoplexes designed for in vivo or ex vivo nucleic acid transfer. |
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