Leong-Poi Howard, Kuliszewski Michael A, Lekas Michael, Sibbald Matthew, Teichert-Kuliszewska Krystyna, Klibanov Alexander L, Stewart Duncan J, Lindner Jonathan R
Division of Cardiology, Keenan Research Centre in the Li Ka Shing Knowledge Institute, St Michael's Hospital, Toronto, Ontario, Canada.
Circ Res. 2007 Aug 3;101(3):295-303. doi: 10.1161/CIRCRESAHA.107.148676. Epub 2007 Jun 21.
Current methods of gene delivery for therapeutic angiogenesis are invasive, requiring either intraarterial or intramuscular administration. A noninvasive method of gene delivery has been developed using ultrasound-mediated destruction of intravenously administered DNA-bearing carrier microbubbles during their microcirculatory transit. Here we show that chronic ischemia could be markedly improved by ultrasound-mediated destruction of microbubbles bearing vascular endothelial growth factor-165 (VEGF(165)) plasmid DNA. Using a model of severe chronic hindlimb ischemia in rats, we demonstrated that ultrasound mediated VEGF(165)/green fluorescent protein (GFP) plasmid delivery resulted in a significant improvement in microvascular blood flow by contrast-enhanced ultrasound, and an increased vessel density by fluorescent microangiography, with minimal changes in control groups. The improvement in tissue perfusion was attributed predominantly to increases in noncapillary blood volume or arteriogenesis, with perfusion peaking at 14 days after delivery, followed by a partial regression of neovascularization at 6 weeks. Transfection was localized predominantly to the vascular endothelium of arterioles in treated ischemic muscle. RT-PCR confirmed the presence of VEGF(165)/GFP mRNA within treated ischemic muscle, being highest at day 3 postdelivery, and subsequently decreasing, becoming almost undetectable by 6 weeks. We found a modulation of endogenous growth factor expression in VEGF-treated ischemic muscle, consistent with a biologic effect of ultrasound mediated gene delivery. The results of our study demonstrate the utility of ultrasonic destruction of plasmid-bearing microbubbles to induce therapeutic arteriogenesis in the setting of severe chronic ischemia.
目前用于治疗性血管生成的基因递送方法具有侵入性，需要动脉内或肌肉内给药。现已开发出一种非侵入性基因递送方法，即在静脉注射携带DNA的载体微泡进行微循环转运期间，利用超声介导对其进行破坏。在此我们表明，通过超声介导破坏携带血管内皮生长因子165（VEGF(165)）质粒DNA的微泡，可显著改善慢性缺血状况。利用大鼠严重慢性后肢缺血模型，我们证明超声介导的VEGF(165)/绿色荧光蛋白（GFP）质粒递送通过超声造影增强显著改善了微血管血流，并通过荧光微血管造影增加了血管密度，而对照组变化极小。组织灌注的改善主要归因于非毛细血管血容量增加或动脉生成，灌注在递送后14天达到峰值，随后在6周时新生血管形成部分消退。转染主要定位于治疗的缺血肌肉中小动脉的血管内皮。RT-PCR证实治疗的缺血肌肉中存在VEGF(165)/GFP mRNA，在递送后第3天最高，随后下降，到6周时几乎检测不到。我们发现VEGF治疗的缺血肌肉中内源性生长因子表达受到调节，这与超声介导基因递送的生物学效应一致。我们的研究结果证明了超声破坏携带质粒的微泡在严重慢性缺血情况下诱导治疗性动脉生成的效用。
