Magnetic nanoparticles assisted modulation of the vascular cGMP system
Zusammenfassung der Projektergebnisse
Targeted and localized gene- and cell-delivery to the vascular walls is a highly attractive tool for the treatment of vascular disorders. However, practical application of gene therapy of the vascular system is hampered by non-permissive conditions that arise due to blood flow (in vivo) as well as hypothermia and time constraints (ex vivo). Additionally, application of gene transfer reagents through the blood stream easily results in low gene transfer efficiency and a high degree of unspecific targeting. We could show that associating lentiviral vectors with magnetic nanoparticles enables specific delivery of genetic material to the target tissue under the application of a magnetic field, even under physiological flow conditions, low temperatures, and short incubation periods. Furthermore, we were able to specifically position MNP-loaded and LV-transduced endothelial cells both ex vivo and in vivo, highlighting the practical therapeutic potential of this technique. Additionally, we screened an array of differently coated MNPs in order to optimize LV/MNP assembly, identifying parameters maximizing binding capacity, transduction efficiency, and magnetic responsiveness. Transduction with LV/MNP complexes under non-permissive conditions were tested in different endothelial cell lines, reaching transduction efficiencies equal or better than traditional overnight transduction in all tested cell types. The MNPs and MNP/LV complexes were also tested for cytotoxicity; here we could see no significant decrease in cell viability with any of the MNP types or concentrations tested. Finally, we investigated the role of the vasodilator-stimulated phosphoprotein (VASP) and its role in cyclic GMP signaling in vascular smooth muscle cells. Previous work in brown adipose tissue had revealed negative feedback loop, where VASP inhibited the expression of soluble guanylate cyclase through the small GTPase Rac-1. While VASP-deficient aortas did not display any cGMP-related phenotype, overexpression of a constitutively active Rac-1 mutant significantly increased sGCß1 expression in vitro. With the aim of further investigating the role of Rac-1 activity in smooth muscle cells ex vivo, we adapted our flow loop perfusion system to allow for tuned disruption of LV-MMB complexes in intact or denuded aortas. Due to the dense matrix of the internal elastic laminae, this approach proved to be insufficient, and we were unable to achieve any transgene expression of the smooth muscle cell layer under the tested conditions.
Projektbezogene Publikationen (Auswahl)
- Combined targeting of lentiviral vectors and positioning of transduced cells by magnetic nanoparticles. PNAS. 2009;106 (1)
Hofmann A, Wenzel D, Becher UM, Freitag D, Klein AM, Eberbeck D, Schulte M, Zimmermann K, Bergemann C, Gleich B, Roell W, Weyh T, Trahms L, Nickenig G, Fleischmann BK, Pfeifer A
(Siehe online unter https://doi.org/10.1073/pnas.0803746106) - Identification of magnetic nanoparticles for combined positioning and lentiviral transduction of endothelial cells. Pharm Res. 2012 May;29(5):1242-54
Wenzel D, Rieck S, Vosen S, Mykhaylyk O, Trueck C, Eberbeck D, Trahms L, Zimmermann K, Pfeifer A, Fleischmann BK
(Siehe online unter https://doi.org/10.1007/s11095-011-0657-5) - Magnetic nanoparticles for biomedical applications. Pharm Res. 2012 May;29(5):1161-4
Pfeifer A, Zimmermann K, Plank C
(Siehe online unter https://doi.org/10.1007/s11095-012-0736-2) - Optimization of Magnetic Nanoparticle-Assisted Lentiviral Gene Transfer. Pharm Res. 2012 May;29(5):1255-69
Trueck C, Zimmermann K, Mykhaylyk O, Anton M, Vosen S, Wenzel D, Fleischmann BK, Pfeifer A
(Siehe online unter https://doi.org/10.1007/s11095-011-0660-x) - Targeted endothelial gene delivery by ultrasonic destruction of magnetic microbubbles carrying lentiviral vectors. Pharm Res. 2012 May;29(5):1282-94
Mannell H, Pircher J, Räthel T, Schilberg K, Zimmermann K, Pfeifer A, Mykhaylyk O, Gleich B, Pohl U, Krötz F
(Siehe online unter https://doi.org/10.1007/s11095-012-0678-8) - Vascular Repair by Circumferential Cell Therapy Using Magnetic Nanoparticles and Tailored Magnets. ACS Nano. 2016 Jan 26;10(1):369-76
Vosen S, Rieck S, Heidsieck A, Mykhaylyk O, Zimmermann K, Bloch W, Eberbeck D, Plank C, Gleich B, Pfeifer A, Fleischmann BK, Wenzel D
(Siehe online unter https://doi.org/10.1021/acsnano.5b04996)
