Abstract
In the last two decades, cationic liposomes have been investigated as vehicles for nucleic acids [plasmid DNA (pDNA) and small interfering RNA (siRNA)] delivery in vitro and in vivo. The formation of cationic liposomes–nucleic acids complexes, termed lipoplexes, depends on a number of experimental variables. The quality of the nucleic acid and the cationic liposome as well as the selection of diluents for diluting the concentrated stocks strongly affect the resulting lipoplexes and their efficiency of gene-expression or gene-silencing effect following transfection. In addition, the molar ratio of cationic lipid nitrogen (N) to siRNA or pDNA phosphate (P) (N/P ratio) influences the final characteristics of the lipoplexes, such as size, surface zeta potential, and reproducibility, thereby reflecting their efficiency following transfection. The methods presented in this chapter could be helpful to obtain reliable and reproducible lipoplexes and experimental results.
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References
Templeton NS, Lasic DD, Frederik PM, Strey HH, Roberts DD, Pavlakis GN (1997) Improved DNA: liposome complexes for increased systemic delivery and gene expression. Nat Biotechnol 15:647–652
Elbashir SM, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411:494–498
Dass CR, Choong PF (2006) Selective gene delivery for cancer therapy using cationic liposomes: in vivo proof of applicability. J Control Release 113:155–163
Zhang S, Zhao B, Jiang H, Wang B, Ma B (2007) Cationic lipids and polymers mediated vectors for delivery of siRNA. J Control Release 123:1–10
Garcia-Chaumont C, Seksek O, Grzybowska J, Borowski E, Bolard J (2000) Delivery systems for antisense oligonucleotides. Pharmacol Ther 87:255–277
Lasic DD, Templeton NS (1996) Liposome in gene therapy. Adv Drug Deliv Rev 20:221–266
Lima MCP, Simoes S, Pires P, Faneca H, Duzgunes N (2001) Cationic lipid-DNA complexes in gene delivery: from biophysics to biological applications. Adv Drug Deliv Rev 47:277–294
Felgner JH, Kumar R, Sridhar CN, Wheeler CJ, Tsai YJ, Border R et al (1994) Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations. J Biol Chem 269:2550–2561
Sternberg B, Hong K, Zheng W, Papahadjopoulos D (1998) Ultrastructural characterization of cationic liposome-DNA complexes showing enhanced stability in serum and high transfection activity in vivo. Biochim Biophys Acta 1375:23–35
Li CX, Parker A, Menocal E, Xiang S, Borodyansky L, Fruehauf JH (2006) Delivery of RNA interference. Cell Cycle 5:2103–2109
Spagnou S, Miller AD, Keller M (2004) Lipidic carriers of siRNA: differences in the formulation, cellular uptake, and delivery with plasmid DNA. Biochemistry 43:13348–13356
Li W, Ishida T, Tachibana R, Almofti MR, Wang X, Kiwada H (2004) Cell type-specific gene expression, mediated by TFL-3, a cationic liposomal vector, is controlled by a post-transcription process of delivered plasmid DNA. Int J Pharm 276:67–74
Almofti MR, Harashima H, Shinohara Y, Almofti A, Li W, Kiwada H (2003) Lipoplex size determines lipofection efficiency with or without serum. Mol Membr Biol 20:35–43
Hirsch-Lerner D, Zhang M, Eliyahu H, Ferrari ME, Wheeler CJ, Barenholz Y (2005) Effect of “helper lipid” on lipoplex electrostatics. Biochim Biophys Acta 1714:71–84
Hofland HEJ, Shephard L, Sullivan SM (1996) Formation of stable cationic lipid/DNA complexes for gene transfer. Proc Natl Acad Sci USA 93:7305–7309
Boussif O, Lezoualc’h F, Zanta MA, Mergny MD, Scherman D, Demeneix B, Behr J-P (1995) A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: Polyethylenimide. Proc Natl Acad Sci USA 92:7297–7301
Doyle SR, Chan CK (2007) Differential intracellular distribution of DNA complexed with polyethylenimine (PEI) and PEI-polyarginine PTD influences exogenous gene expression within live COS-7 cells. Genet Vaccines Ther 5:11
Lleres D, Weibel JM, Heissler D, Zuber G, Duportail G, Mely Y (2004) Dependence of the cellular internalization and transfection efficiency on the structure and physicochemical properties of cationic detergent/DNA/liposomes. J Gene Med 6:415–428
Zuidam NJ, Hirsch-Lerner D, Margulies S, Barenholz Y (1999) Lamellarity of cationic liposomes and mode of preparation of lipoplexes affect transfection efficiency. Biochim Biophys Acta 1419:207–220
Acknowledgments
The research was supported in part by the Health and Labour Sciences Research Grants for Research on Advanced Medical Technology from The Ministry of Health, Labour and Welfare of Japan. We thank the Japan Association for the Advancement of Medical Equipment for supporting the Postdoctoral Fellowship for Dr. Jose Mario Barichello.
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Barichello, J.M., Ishida, T., Kiwada, H. (2010). Complexation of siRNA and pDNA with Cationic Liposomes: The Important Aspects in Lipoplex Preparation. In: Weissig, V. (eds) Liposomes. Methods in Molecular Biology, vol 605. Humana Press. https://doi.org/10.1007/978-1-60327-360-2_32
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DOI: https://doi.org/10.1007/978-1-60327-360-2_32
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