Cationic lipids derived from glycine betaine promote efficient and non-toxic gene transfection in cultured hepatocytes |
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Authors: | Gilot David Miramon Marie-Laure Benvegnu Thierry Ferrieres Vincent Loreal Olivier Guguen-Guillouzo Christiane Plusquellec Daniel Loyer Pascal |
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Affiliation: | H?pital Pontchaillou, Régulation des Equilibres fonctionnels du foie Normal et Pathologique, INSERM U522, Avenue de la Bataille Flandre/Dunkerque, 35033 Rennes, France. david.gilot@rennes.inserm.fr |
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Abstract: | Background The low efficiency and toxicity of transfection in a primary culture of hepatocytes using cationic lipids remains a limiting step to the study of gene function and the setting up of non‐viral gene therapy. Methods A novel class of cationic lipids (GBs) derived from natural glycine betaine compounds covalently linked to acyl chains by enzymatically hydrolysable peptide and ester bonds, a structure designed to reduce cytotoxicity, was used to improve transfection efficiency in a primary culture of rat hepatocytes. The relationship between lipid structure, lipoplex formulation and transfection efficiency was studied using six GBs (12‐14‐16, 22‐24‐26) varying in their spacer and acyl chains. Results GB12, characterized by short [(CH2)10] acyl chains and spacer, allowed plasmid uptake in all cells and reporter gene expression in up to 40% of hepatocytes with a low cytotoxicity, a much higher efficiency compared with transfections using other reagents including Fugene6? and Lipofectin?. We also showed that numerous cells accumulated high amounts of plasmids demonstrating that GB12 promoted a very efficient DNA transfer through plasma membrane leading to an increase in nuclear plasmid translocation, allowing a much higher gene expression. Moreover, GB12‐transfected hepatocytes survived to injection in normal livers and were found to express the LacZ reporter gene. Conclusions The non‐toxic GB12 formulation is a powerful vehicle for plasmid delivery in cultured hepatocytes with relevance in liver gene therapy. Copyright © 2002 John Wiley & Sons, Ltd. |
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Keywords: | hepatocytes cationic lipids gene transfer gene therapy ex vivo |
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