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Keywords:

  • branched polyethylenimine;
  • calcium phosphate;
  • non-viral vectors;
  • gene delivery;
  • plasmid DNA;
  • HEPM cells;
  • transfection efficiency;
  • cytotoxicity

Abstract

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.