Abstract
Purpose. The purpose of this study was to evaluate protamine-mediated gene transfection by liposomes with a novel cationic cholesterol derivative (I) compared to those with DC-Chol or DOTMA (Lipofectin).
Methods. Plasmid pGL3 DNA was complexed to the cationic liposomes with the derivative (I), DC-Chol, or DOTMA in SFM101(Nissui) at room temperature for 15 min, and thereafter the complex was incubated with target cells (NIH3T3) for 4 h at 37°C. The cells then were washed and cultured for another 40 h in the growth medium at 37°C before luciferase assay.
Results. The transfection efficiency by the liposomes with the derivative (I) was much higher than that by the liposomes with DC-Chol or DOTMA. In addition, its transfection efficiency was enhanced greatly by the addition of protamine. Atomic force microscopy showed clearly how the size of the DNA-liposome complex was changed by protamine. Furthermore, fluorescence microscopic images showed that Cy5-labeled antisense DNAs were transferred quicker into the nucleus of the target cells by the liposomes with the derivative I in the presence of protamine.
Conclusion. Although there exist several possible mechanisms, such as improved protection of DNA intracellularly by derivative (I), one possibility is that the DNA-protamine-liposome complex with the derivative (I) promoted gene transfection more significantly into the nucleus of the target cells using the nuclear localization signal of protamine.
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Noguchi, A., Hirashima, N. & Nakanishi, M. Cationic Cholesterol Promotes Gene Transfection Using the Nuclear Localization Signal in Protamine. Pharm Res 19, 933–938 (2002). https://doi.org/10.1023/A:1016449902541
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DOI: https://doi.org/10.1023/A:1016449902541