Eulsoon Park1, Hong-Baek Cho2, Koichi Takimoto3. 1. Department of Bioengineering, Nagaoka University of Technology, Nagaoka, Niigata, Japan. 2. Extreme Energy-Density Research Institute, Nagaoka University of Technology, Nagaoka, Niigata, Japan. 3. Department of Bioengineering, Nagaoka University of Technology, Nagaoka, Niigata, Japan. Electronic address: koichi@vos.nagaokaut.ac.jp.
Abstract
BACKGROUND AIMS: Adipose-derived stem cells have the ability to turn into several clinically important cell types. However, it is difficult to transfect these cells with the use of conventional cationic lipid-based reagents. Polyethylenimine (PEI) is considered to be an inexpensive and effective tool for delivery of nucleic acids into mammalian cells. METHODS: We used a linear PEI conjugated with the nuclear localization signal (NLS) peptide of Simian vacuolating virus 40 large T antigen (PEI-NLS) for transfection of plasmid DNA into adipose-derived cells. We also tested if transfection of cells in suspension might improve the degree and duration of exogenous gene expression. RESULTS: Transfection of cells in suspension with the use of a PEI conjugated with an NLS peptide resulted in high levels of reporter gene expression for an extended period of time in clonal 3T3-L1 preadipocytes and native human adipose-derived stem cells. The reporter gene expression increased for 3 days after the addition of the PEI-NLS peptide-DNA mixture in cell suspension and remained significant for at least 7 days. Cell density did not influence the level of reporter gene expression. Thus, the suspension method with the use of an NLS peptide-conjugated PEI leads to a robust and sustained expression of exogenous genes in adipose-derived cells. CONCLUSIONS: The devised transfection method may be useful for reprogramming of adipose-derived stem cells and cell-based therapy.
BACKGROUND AIMS: Adipose-derived stem cells have the ability to turn into several clinically important cell types. However, it is difficult to transfect these cells with the use of conventional cationic lipid-based reagents. Polyethylenimine (PEI) is considered to be an inexpensive and effective tool for delivery of nucleic acids into mammalian cells. METHODS: We used a linear PEI conjugated with the nuclear localization signal (NLS) peptide of Simian vacuolating virus 40 large T antigen (PEI-NLS) for transfection of plasmid DNA into adipose-derived cells. We also tested if transfection of cells in suspension might improve the degree and duration of exogenous gene expression. RESULTS: Transfection of cells in suspension with the use of a PEI conjugated with an NLS peptide resulted in high levels of reporter gene expression for an extended period of time in clonal 3T3-L1 preadipocytes and native human adipose-derived stem cells. The reporter gene expression increased for 3 days after the addition of the PEI-NLS peptide-DNA mixture in cell suspension and remained significant for at least 7 days. Cell density did not influence the level of reporter gene expression. Thus, the suspension method with the use of an NLS peptide-conjugated PEI leads to a robust and sustained expression of exogenous genes in adipose-derived cells. CONCLUSIONS: The devised transfection method may be useful for reprogramming of adipose-derived stem cells and cell-based therapy.
Authors: Savannah E Est-Witte; Ashley L Farris; Stephany Y Tzeng; Daphne L Hutton; Dennis H Gong; Kaitlyn G Calabresi; Warren L Grayson; Jordan J Green Journal: Acta Biomater Date: 2020-07-02 Impact factor: 10.633