title: Modeling cytoplasmic release of encapsulated oligonucleotides from cationic liposomes.
abstract: Transfection activity of antisense oligodeoxynucleotides (ODN)-loaded cationic liposomes is mainly restricted by uptake and ODN release into cytoplasm, which is difficult to evaluate in cell culture studies. Well-designed models of cellular membranes, aim of the present study, might facilitate investigation of such processes. In this investigation, a phosphorothioate ODN was actively encapsulated in a DODAP-containing cationic liposome by ethanol injection with 73% efficiency. ODN release was determined by fluorescence dequenching of FITC-ODN upon incubation of liposomes with early endosomal (EE), late endosomal (LE) and plasma membranes (PM) models. LE provided the highest release (up to 76%) in a temperature-dependent manner. Release by EE (<16%), total PM (<11%) and PM external layer ( approximately 0) were not temperature sensitive. These differences are attributed to lipid charge, chain mobility, critical packing parameter and cholesterol content of the models. Intracellular distribution of FITC-ODN, determined by fluorescence microscopy and flowcytometry in the presence and absence of sodium azide, confirmed that liposomes were internalized mainly via endocytosis; hence inability of our PL models to simulate such active processes. Instead, release of ODN from endosomes into cytoplasm was pH-sensitive and in good agreement with model membrane studies in terms of amount and mechanism.
authors.affiliation: School of Pharmacy, Shaheed Beheshti Medical University, PO Box: 14155-6153, Tehran, Iran.
authors.forename: Ali M
authors.initials: AM
authors.lastname: Tamaddon
authors.forename: Farshad H
authors.initials: FH
authors.lastname: Shirazi
authors.forename: Hamid R
authors.initials: HR
authors.lastname: Moghimi
mesh_terms: D000818:Animals; D002214:Capsules; D002738:Chloroquine / Q000494:pharmacology; D002784:Cholesterol / Q000737:chemistry; D003593:Cytoplasm / Q000378:metabolism*; D004705:Endocytosis / Q000187:drug effects; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D018014:Gene Transfer Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D008055:Lipids / Q000737:chemistry; D008081:Liposomes; D051379:Mice; D008856:Microscopy, Fluorescence; D008985:Monensin / Q000494:pharmacology; D041681:NIH 3T3 Cells; D020319:Oligodeoxyribonucleotides, Antisense / Q000008:administration & dosage / Q000493:pharmacokinetics*; D010710:Phosphates / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry; D019810:Sodium Azide / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D013696:Temperature
chemical_list: D002214:Capsules; D008055:Lipids; D008081:Liposomes; D020319:Oligodeoxyribonucleotides, Antisense; D010710:Phosphates; D010743:Phospholipids; C083120:phosphorodithioic acid; D002738:Chloroquine; D008985:Monensin; D019810:Sodium Azide; D002784:Cholesterol; D016650:Fluorescein-5-isothiocyanate