Abstract
Gene delivery is a widespread strategy in current experimental medicine. In this work, we report a method for low-toxic intracellular DNA vector delivery and post transfection localisation of this vector in mouse embryonic fibroblast cell lines. The surface of modified ferumoxide nanoparticles conjugated with Rhoda-mine B isothiocyanate (FeNV-Rh) was modified with linear polyethyleneimine and medium molecular weight chitosan to increase Accelerated Sensor of Action Potentials DNA vector adhesion. The size of the FeNV-Rh/DNA transfection complex was studied using dynamic light scattering (DLS) and scanning electron microscopy (SEM) techniques. The transfection complex internalisation of plasmid expression and FeNV-Rh, and stability of rhodamine fluorescence in intracellular space were observed at time periods 6, 12, 24 and 48 h post transfection. Results showed high transfection complex intracellular biocompatibility—cell viability after Rh-MNP labelling was higher than 97% 24 h after transfection, and higher than 95% after the next 24 h. Selective FeNV-Rh localisation in the lysosomes was quantified. More than 82% of nanoparticles were localised in the lysosomes 12 h post transfection and 94% of lysosomes had a significant and long-term deposit of nanoparticles. DNA vector expression was visible in >65% of the cells and precise protein localisation on the cell membrane was confirmed using confocal microscopy.
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References
Dobson J (2006) Gene therapy progress and prospects: magnetic nanoparticle-based gene delivery. Gene Ther 13:283–287. https://doi.org/10.1038/sj.gt.3302720.
Bregar VB, Lojk J, Šuštar V, et al (2013) Visualization of internalization of functionalized cobalt ferrite nanoparticles and their intracellular fate. Int J Nanomedicine 8:919–931.
Durán MC, Willenbrock S, Barchanski A, et al (2011) Comparison of nanoparticle-mediated transfection methods for DNA expression plasmids. J Nanobiotechnology 9:47.
Duan J, Yu Y, Yu Y, et al (2014) Silica nanoparticles enhance autophagic activity, disturb endothelial cell homeostasis and impair angiogenesis. Part Fibre Toxicol 11:50.
Popov A, R. Popova N, Selezneva I, et al (2016) Cerium oxide nanoparticles stimulate proliferation of primary mouse embryonic fibroblasts in vitroSt-Pierre F, Marshall JD, Yang Y, et al (2014) High-fidelity optical reporting of neuronal electrical activity with an ultrafast fluorescent voltage sensor. Nat Neurosci 17:884–9.
Lee EEL, Bezanilla F (2017) Biophysical Characterization of Genetically Encoded Voltage Sensor ASAP1: Dynamic Range Improvement. Biophys J 113:2178–2181.
Cmiel V, Skopalik J, et al (2017) Rhodamine bound maghemite as a long-term dual imaging nanoprobe of adipose tissue-derived mesenchymal stromal cells. Eur Biophys J 46:433–444.
Acknowledgements
Research described in this paper was financed by Czech Ministry of Education in frame of National Sustainability Program under grant LO1401. For research, infrastructure of the SIX Center was used.
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Svoboda, O. et al. (2019). DNA Intracellular Delivery into 3T3 Cell Line Using Fluorescence Magnetic Ferumoxide Nanoparticles. In: Lhotska, L., Sukupova, L., Lacković, I., Ibbott, G. (eds) World Congress on Medical Physics and Biomedical Engineering 2018. IFMBE Proceedings, vol 68/3. Springer, Singapore. https://doi.org/10.1007/978-981-10-9023-3_27
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DOI: https://doi.org/10.1007/978-981-10-9023-3_27
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