ISSN : 0975-9492


Open Access

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Title : Increased Transfection Efficiency of Lipid- Coated Methacrylate-Based Gene Delivery Systems
Authors : Noura H. Abd Ellah, Leeanne Taylor, Neil Ayres, Mona M. Elmahdy, Gihan N. Fetih, El-Sayed A. Ibrahim, Giovanni M. Pauletti
Keywords : lipopolyplex, DOTAP, DOPE, pHPMA-b-pDMAEMA copolymer
Issue Date : November 2014
Abstract :
Purpose: To develop a safe, lipid-based non-viral gene delivery system that achieves high transfection efficiency in the presence of serum proteins.
Methods: Polyplexes with the pAcGFP1-C1 plasmid were formed in phosphate buffered saline, pH 7.4 (PBS) using the novel poly[N-(2-hydroxypropyl)methacrylamide]-poly(N,N-dimethylaminoethylmethacrylate) diblock copolymer (pHPMA-b-pDMAEMA) at N/P=4. Cationic-Liposomes were prepared from a dried lipid film comprised of equimolar 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). Lipopolyplexes were fabricated at lipid/DNA weight ratios up to 40. Particle size distribution and zeta potential of lipopolyplexes were determined by dynamic light scattering. HeLa cells viability in the presence and absence of lipopolyplexes was quantified using the CellTiter-Glo® luminescent assay. HeLa cell transfection efficiency in the presence and absence of FBS was visually assessed by confocal microscopy and quantitatively compared to the TurboFect™ control.
Results: pHPMA-b-pDMAEMA exhibited a high condensation capacity of 1 µg of pDNA per 0.513 µg of polymer (N/P=1). Lipid-coating of polyplexes at lipid/DNA weight ratios up to 40 resulted in particle sizes <200 nm and zeta potential >+25 mV. Exposure to FBS significantly increased mean particle size to >300 nm, reduced zeta potential to -10 mV, and augmented polydispersity. Lipid coating of polyplexes only decreased HeLa cell viability at lipid/DNA ratios >20. HeLa transfection with lipopolyplexes was most effective at lipid/DNA = 20 and was significantly greater in the presence of FBS than measured for lipid-free polyplexes.
Conclusion: Lipid coating of pHPMA-b-pDMAEMA/DNA polyplexes with an equimolar DOTAP/DOPE mixture at a lipid/DNA ratio = 20 effectively enhances in vitro transfection efficiency of HeLa cells in the presence of serum proteins.
Page(s) : 812-819
ISSN : 0975-9492
Source : Vol. 5, No.11