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Magnetic Nanoparticle and Magnetic Field Assisted siRNA Delivery In Vitro

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RNA Interference

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1218))

Abstract

This chapter describes how to design and conduct experiments to deliver siRNA to adherent cell cultures in vitro by magnetic force-assisted transfection using self-assembled complexes of small interfering RNA (siRNA) and cationic lipids or polymers that are associated with magnetic nanoparticles (MNPs). These magnetic complexes are targeted to the cell surface by the application of a gradient magnetic field. A further development of the magnetic drug-targeting concept is combining it with an ultrasound-triggered delivery using magnetic microbubbles as a carrier for gene or drug delivery. For this purpose, selected MNPs, phospholipids, and siRNAs are assembled in the presence of perfluorocarbon gas into flexible formulations of magnetic lipospheres (microbubbles). Methods are described how to accomplish the synthesis of magnetic nanoparticles for magnetofection and how to test the association of siRNA with the magnetic components of the transfection vector. A simple method is described to evaluate magnetic responsiveness of the magnetic siRNA transfection complexes and estimate the complex loading with magnetic nanoparticles. Procedures are provided for the preparation of magnetic lipoplexes and polyplexes of siRNA as well as magnetic microbubbles for magnetofection and downregulation of the target gene expression analysis with account for the toxicity determined using an MTT-based respiration activity test. A modification of the magnetic transfection triplexes with INF-7, fusogenic peptide, is described resulting in reporter gene silencing improvement in HeLa, Caco-2, and ARPE-19 cells. The methods described can also be useful for screening vector compositions and novel magnetic nanoparticle preparations for optimized siRNA transfection by magnetofection in any cell type.

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Acknowledgments

We gratefully acknowledge the support from the German Research Foundation through the DFG Research Unit FOR917 (Project PL 281/3-1), from the German Federal Ministry of Education and Research through grants ZIM-KOOP “STEP-MAG,” Nanobiotechnology grants 13N8186 and 13N8538, and from the Excellence Cluster “Nanosystems Initiative Munich.” The authors would like to thank Dr. Bob Scholte for transduction of the H441 cells with eGFP and luciferase using lentiviral vectors and Dr. Christian Bergemann (chemicell GmbH, Berlin) for providing MNPs for the preparation of the MAALs. This work was supported by the European Union through the FP6-LIFESCIHEALTH Project “Improved precision of nucleic acid based therapy of cystic fibrosis” under contract no. 005213.

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Authors and Affiliations

  1. Institute of Experimental Oncology, Klinikum rechts der Isar der Technischen Universität München, Ismaninger Strasse 22, Munich, 81675, Germany

    Olga Mykhaylyk, Yolanda Sanchez-Antequera, Dialechti Vlaskou, Edelburga Hammerschmid & Martina Anton

  2. National Atomic Energy Commission, Av General Paz 1499, Buenos Aires, Argentina

    Maria Belen Cerda

  3. Faculty of Pharmaceutical Sciences, The University of British Columbia, 2405 Wesbrook Mall, Vancouver, BC, Canada, V6T 1Z3

    Mehrdad Bokharaei

  4. Institute of Experimental Oncology, Klinikum rechts der Isar der Technischen Universität München, Ismaninger Strasse 22, Munich, 81675, Germany

    Christian Plank

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  1. Olga Mykhaylyk
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Correspondence to Olga Mykhaylyk .

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  1. Institute for Cancer Research, Oslo University Hospital, Montebello, Oslo, Norway

    Mouldy Sioud

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Mykhaylyk, O. et al. (2015). Magnetic Nanoparticle and Magnetic Field Assisted siRNA Delivery In Vitro. In: Sioud, M. (eds) RNA Interference. Methods in Molecular Biology, vol 1218. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1538-5_5

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  • DOI: https://doi.org/10.1007/978-1-4939-1538-5_5

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