Characterization and evaluation of the efficacy of cationic complex mediated plasmid DNA delivery in human embryonic palatal mesenchyme cells.
The purpose of this study was to develop and test a non-viral gene delivery system that can be employed to deliver genes of interest into a pre-osteoblastic cell line. Human embryonic palatal mesenchymal (HEPM 1486) cells were transfected with vector-plasmid DNA (pDNA) complexes. We explored calcium phosphate and polyethylenimine (PEI) as non-viral vectors and compared their respective in vitro transfection efficacies. Plasmid DNA encoding luciferase protein (LUC) was complexed with PEI (with differing N:P ratios) and calcium phosphate (with differing Ca:P ratios), using established protocols. The complexes prepared were then characterized for size and surface charge, using a Malvern Zetasizer Nano-ZS. The transfection efficiency and cytotoxicity of the prepared complexes were evaluated in HEPM cells. The PEI-pDNA complexes over the whole range of N:P ratios were found to be < 160 nm in size, while the calcium phosphate-pDNA complexes were relatively bigger. The PEI-pDNA complexes prepared at a N:P ratio of 10 were found to have maximum transfection efficiency at 4 h of treatment, with minimal cytotoxicity. The highest transfection efficiency obtained with calcium phosphate-pDNA complexes (Ca:P 200) was nearly 12-fold lower than that obtained with PEI-pDNA complexes (N:P 10). Following this, transgene expression in the HEPM cells treated with complexes prepared at a N:P ratio of 10 was further examined, using pDNA coding for enhanced green fluorescent protein (EGFP-N1) or therapeutically relevant platelet-derived growth factor B (PDGF-B). In conclusion, PEI was a more effective vector for delivering genes of interest to pre-osteoblasts than calcium phosphate. Copyright © 2014 John Wiley & Sons, Ltd.
Division of Pharmaceutics and Translational Therapeutics, College of Pharmacy, University of Iowa, IA, USA.
Sheetal
S
D'Mello
Division of Pharmaceutics and Translational Therapeutics, College of Pharmacy, University of Iowa, IA, USA.
Aliasger K
AK
Salem
Department of Periodontics, College of Dentistry, University of Iowa, IA, USA.
Liu
L
Hong
Department of Periodontics, College of Dentistry, University of Iowa, IA, USA.
Satheesh
S
Elangovan
D002130:Calcium Phosphates / Q000737:chemistry*; D002460:Cell Line; D004622:Embryo, Mammalian / Q000166:cytology*; D005822:Genetic Vectors; D006801:Humans; D008648:Mesoderm / Q000166:cytology*; D010957:Plasmids; D014162:Transfection / Q000379:methods*
D002130:Calcium Phosphates; C020243:calcium phosphate
HEPM cells; branched polyethylenimine; calcium phosphate; cytotoxicity; gene delivery; non-viral vectors; plasmid DNA; transfection efficiency