Nuclear localization signal peptides enhance transfection efficiency of chitosan/DNA complexes.
The purpose of this study was to investigate the potential of nuclear localization signal (NLS) "KPKKKRKV" to mediate the in vitro transfection efficiency of chitosan (CS)/DNA complexes, aiming at its use in gene therapy applications. In the preparation of CS/DNA complexes containing NLS, peptide with NLS was directly incorporated without covalent conjugation to pDNA or chitosan. The gene transfer efficiency of CS/DNA complexes with and without NLS was evaluated in the human cervical carcinoma cell line (Hela cells). The CS/DNA complex containing NLS increased transfection efficiencies in a NLS-dose dependent manner on the Hela cells, compared to the control (CS/DNA complex or NLS). The highest transfection efficiency was significantly observed in CS/DNA complex at the weight ratio of 8 with 120 microg NLS and was 74-fold higher than that in the cells transfected with CS/DNA complex. Cytotoxicity of the NLS/CS/DNA complexes increased as the amount of the peptide increased, however, over 80% average cell viability was observed for complexes at the effective concentration of the peptide for transfections. Therefore, the NLS is expected to be a potent transfection enhancing agent without a covalent conjugation to pDNA or chitosan. Our findings suggest that the high gene expression with the negligible cytotoxicity can be achieved by adding the NLS peptide to chitosan/DNA complexes at an optimal ratio.
Opanasopit
Praneet
P
Nanotechnology for Drug/Gene Delivery Systems Group, Faculty of Pharmacy, Silpakorn University, Nakhon Pathom 73000, Thailand.
Rojanarata
Theerasak
T
Apirakaramwong
Auayporn
A
Ngawhirunpat
Tanasait
T
Ruktanonchai
Uracha
U
D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D048271:Chitosan / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D006801:Humans; D010316:Particle Size; D021382:Protein Sorting Signals; D014162:Transfection / Q000379:methods*
D021382:Protein Sorting Signals; D004247:DNA; D048271:Chitosan