title: Ultrasound-mediated gene delivery of naked plasmid DNA in skeletal muscles: a case for bolus injections.
abstract: Localized gene delivery has many potential clinical applications. However, the nucleic acids (e.g. pDNA and siRNA) are incapable of passively crossing the endothelium, cell membranes and other biological barriers which must be crossed to reach their intracellular targets. A possible solution is the use of ultrasound to burst circulating microbubbles inducing transient permeabilization of surrounding tissues which mediates nucleic acid extravasation and cellular uptake. In this study we report on an optimization of the ultrasound gene delivery technique. Naked pDNA (200 μg) encoding luciferase and SonoVue® microbubbles were co-injected intravenously in mice. The hindlimb skeletal muscles were exposed to ultrasound from a non-focused transducer (1 MHz, 1.25 MPa, PRI 30s) and injection protocols and total amounts as well as ultrasound parameters were systemically varied. Gene expression was quantified relative to a control using a bioluminescence camera system at day 7 after sonication. Bioluminescence ratios in sonicated/control muscles of up to 101× were obtained. In conclusion, we were able to specifically deliver genetic material to the selected skeletal muscles and overall, the use of bolus injections and high microbubble numbers resulted in increased gene expression reflected by stronger bioluminescence signals. Based on our data, bolus injections seem to be required in order to achieve transient highly concentrated levels of nucleic acids and microbubbles at the tissue of interest which upon ultrasound exposure should lead to increased levels of gene delivery. Thus, ultrasound mediated gene delivery is a promising technique for the clinical translation of localized drug delivery.
authors: Biomedical NMR, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands; Center for Imaging Research and Education (CIRE), Eindhoven, The Netherlands.
authors: Pedro Gomes
authors: PG
authors: Sanches
authors: Nijmegen Center for Mitochondrial Disorders, Radboud University Medical Centre, Nijmegen, The Netherlands.
authors: Mareike
authors: M
authors: Mühlmeister
authors: Philips Research North America, Briarcliff Manor, United States.
authors: Ralf
authors: R
authors: Seip
authors: Department of Radiology, Leiden University Medical Center, Leiden, The Netherlands.
authors: Eric
authors: E
authors: Kaijzel
authors: Department of Radiology, Leiden University Medical Center, Leiden, The Netherlands.
authors: Clemens
authors: C
authors: Löwik
authors: Department of Minimally Invasive Healthcare, Philips Research Eindhoven, Eindhoven, The Netherlands.
authors: Marcel
authors: M
authors: Böhmer
authors: Department of Cardiology, Isar Heart Center, Munich, Germany; Department of Nuclear Medicine Technical University of Munich, Munich, Germany.
authors: Klaus
authors: K
authors: Tiemann
authors: Biomedical NMR, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands; Center for Imaging Research and Education (CIRE), Eindhoven, The Netherlands; Department of Minimally Invasive Healthcare, Philips Research Eindhoven, Eindhoven, The Netherlands. Electronic address: h.gruell@tue.nl.
authors: Holger
authors: H
authors: Grüll
mesh_terms: D000818:Animals; D004247:DNA / Q000008:administration & dosage*; D005260:Female; D018014:Gene Transfer Techniques; D007262:Infusions, Intravenous; D007267:Injections; D008156:Luciferases / Q000235:genetics; D049449:Luminescence; D051379:Mice; D045423:Microbubbles; D018482:Muscle, Skeletal / Q000378:metabolism* / Q000473:pathology; D010957:Plasmids; D013010:Sonication
chemical_list: D004247:DNA; D008156:Luciferases
keywords: Gene delivery; Luciferase; Microbubbles; Skeletal muscle; Ultrasound