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Intravital vascular delivery of genetic constructs is limited by low transfection efficiencies and systemic side effects. The magnetofection technique is a new approach for site-directed delivery of oligodesoxynucleotides (ODN) and pDNA coupled to superparamagnetic nanoparticles (“magnetofectins”, MF) using an external magnet, thereby increasing efficiency and reducing toxicity, as shown in. primary human endothelial cells (EC). To exploit these advantages for targeted intravascular ODN transfer in vivo, we injected Cy3-fluorescence-labeled ODN (Cy3-ODN) either “locally” i.e. injection and magnetofection into the femoral artery (n=4) or “systemically” i.e. via the carotid artery and magnetofection of dorsal skin vessels (dorsal skinfold chamber model) of Black6 mice (n=24). Following local magnetofection of Cy3-ODN using a cylindrical permanent magnet (0,4T, 10mm diameter), the vessel yielded high fluorescence as assessed by ex vivo confocal microscopy, in contrast to experiments performed without a magnetic field. Systemic injection followed by exposure of the skinfold chamber to the same permanent magnet (15min) resulted in markedly lower fluorescence within arteriolar and venolar walls, as measured by intravital microscopy. Accumulation of ODN at the target tissue when using an electromagnet (0,9T) was enhanced but did not reach the efficiency obtained after local magnetofection. Assuming that the lower efficiency was due to scavenging of the injected ODN at other sites, ODNs were applied in lipid-based microbubbles (2-4µm). These were observed to circulate for up to 1h. Targeted release of MFs from microbubbles site was achieved by external ultrasound application to the dorsal skin. This lead to a fluorescence intensity in target vessels similar to that observed after local application. To confirm functionality of MF delivery into ECs, we transferred pDNA (pDS-red-N1) encoding for a Red-Fluorescent-Protein. Unlike in experiments using Cy3-ODN, no fluorescence was seen directly after magnetofection, whereas 24h later there was marked red fluorescence in the vessel wall of arterioles and venoles in the target region indicating functionally efficient delivery of the genetic material. These results demonstrate that efficiency of systemic gene delivery can be considerably enhanced when packing the MFs in microbubbles and targeting them by magnetofection. This novel method may be helpful in investigating gene function in vascular pathophysiology.
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J. Vasc. Biol. 42, Sup:2 (2005) pp86-87