THE EFFECT OF PHYSICO-CHEMICAL PROPERTIES OF PROTEIN-DNA NANOPARTICLES ON THE TRASNFECTION EFFICIENCY OF CULTURED HELA AND MACROPHAGE CELLS

THE EFFECT OF PHYSICO-CHEMICAL PROPERTIES OF PROTEIN-DNA NANOPARTICLES ON THE TRASNFECTION EFFICIENCY OF CULTURED HELA AND MACROPHAGE CELLS

OTTENGY, S.; ALVES, R. F.; TORRE, L. G. DE LA; AZZONI, A. R.; SILVA, D. C.

Artigo:

A major concern in DNA vaccination and gene therapy protocols using non-viral vectors is the low efficiency of gene delivery. This is because during the traffic to the target cells nuclei, the DNA vectors must overcome a series of physical, diffusional and enzymatic barriers. The objective of this work is the development of novel delivery particles formed by the combination of proteins, plasmid DNA (pDNA), and lipids. We propose that these particles, also called “artificial viruses”, will rely on the high efficiency of cell internalization of the lipids and then, exploit the cell machinery for intracellular trafficking to the interior of the cells nuclei. In this work, protamine-pDNA and protamine-pDNA-lipid nanoparticles were characterized by gel retardation assay, confirming the ability of the protamine to interact and condense pDNA. Zeta potential and dynamic light scattering studies indicated the presence of positive particles with size in the range of 100 to 700 nm, depending on the incubation pH and time during nanoparticle formation. Transfection studies indicated that addition of protamine to pDNA (pVAX1GFP) and lipids increased the transfection efficiency of mammalian cells (HeLa), comparing to naked DNA. The next step of this study will be the transfection of macrophage cells IC21, hopefully achieving high transfection efficiencies, and adding new information on how the physico-chemical properties of the nanoparticles correlate with transfection efficiency of mammalian cells.

A major concern in DNA vaccination and gene therapy protocols using non-viral vectors is the low efficiency of gene delivery. This is because during the traffic to the target cells nuclei, the DNA vectors must overcome a series of physical, diffusional and enzymatic barriers. The objective of this work is the development of novel delivery particles formed by the combination of proteins, plasmid DNA (pDNA), and lipids. We propose that these particles, also called “artificial viruses”, will rely on the high efficiency of cell internalization of the lipids and then, exploit the cell machinery for intracellular trafficking to the interior of the cells nuclei. In this work, protamine-pDNA and protamine-pDNA-lipid nanoparticles were characterized by gel retardation assay, confirming the ability of the protamine to interact and condense pDNA. Zeta potential and dynamic light scattering studies indicated the presence of positive particles with size in the range of 100 to 700 nm, depending on the incubation pH and time during nanoparticle formation. Transfection studies indicated that addition of protamine to pDNA (pVAX1GFP) and lipids increased the transfection efficiency of mammalian cells (HeLa), comparing to naked DNA. The next step of this study will be the transfection of macrophage cells IC21, hopefully achieving high transfection efficiencies, and adding new information on how the physico-chemical properties of the nanoparticles correlate with transfection efficiency of mammalian cells.

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DOI: 10.5151/chemeng-cobeq2014-1434-19328-182228

Referências bibliográficas

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• [10] 5. AKNOWLEDGMENTS The authors would like to thank FAPESP, CAPES and CNPq for the financial support.

Como citar:

OTTENGY, S.; ALVES, R. F.; TORRE, L. G. DE LA; AZZONI, A. R.; Dennys Correia da Silva; "THE EFFECT OF PHYSICO-CHEMICAL PROPERTIES OF PROTEIN-DNA NANOPARTICLES ON THE TRASNFECTION EFFICIENCY OF CULTURED HELA AND MACROPHAGE CELLS", p-2049-2056. In: Anais do XX Congresso Brasileiro de Engenharia Química - COBEQ 2014 [= Blucher Chemical Engineering Proceedings, v.1, n.2]. São Paulo: Blucher, 2015.
ISSN 23591757, DOI 10.5151/chemeng-cobeq2014-1434-19328-182228