Tsurumi Y, Takeshita S, Chen D, Kearney M, Rossow S T, Passeri J, Horowitz J R, Symes J F, Isner J M
Department of Medicine (Cardiology), St Elizabeth's Medical Center of Boston, Tufts University School of Medicine, Boston, Mass 02135, USA.
Circulation. 1996 Dec 15;94(12):3281-90. doi: 10.1161/01.cir.94.12.3281.
Striated muscle has been shown to be capable of taking up and expressing foreign genes transferred in the form of naked plasmid DNA, although typically with a low level of gene expression. In the case of genes that encode secreted proteins, however, low transfection efficiency may not preclude bio-activity of the secreted gene product. Accordingly, we investigated the hypothesis that intramuscular (IM) gene therapy with naked plasmid DNA encoding vascular endothelial growth factor (VEGF) could augment collateral development and tissue perfusion in an animal model of hindlimb ischemia.
Ten days after ischemia was induced in one rabbit hindlimb, 500 micrograms of phVEGF165, or the reporter gene LacZ, was injected IM into the ischemic hindlimb muscles. Thirty days later, angiographically recognizable collateral vessels and histologically identifiable capillaries were increased in VEGF transfectants compared with controls. This augmented vascularity improved perfusion to the ischemic limb, documented by a superior calf blood pressure ratio for phVEGF165 (0.85 +/- 0.05) versus controls (0.64 +/- 0.05, P < .01), improved blood flow in the ischemic limb (measured with an intra-arterial Doppler wire) at rest (phVEGF165 = 21.3 +/- 3.9 mL/min, control = 14.6 +/- 1.6 mL/min, P < .01) and after a vasodilator (phVEGF165 = 54.2 +/- 12.0 mL/min, control = 37.3 +/- 8.9 mL/min, P < .01) and increased microspheres in the adductor (phVEGF165 = 4.3 +/- 1.6 mL.min-1.100 g of tissue-1, control = 2.9 +/- 1.2 mL.min-1.100 g of tissue-1, P < .05) and gastrocnemius (phVEGF165 = 3.9 +/- 1.0 mL.min-1.100 g of tissue-1, control = 2.8 +/- 1.4 mL.min-1.100 g of tissue-1, P < .05) muscles of the ischemic limb.
Ischemic skeletal muscle represents a promising target for gene therapy with naked plasmid DNA. IM transfection of genes encoding angiogenic cytokines, particularly those that are naturally secreted by intact cells, may constitute an alternative treatment strategy for patients with extensive peripheral vascular disease in whom the use of intravascular catheter-based gene transfer is compromised and/or prohibited.
尽管通常基因表达水平较低，但已证明横纹肌能够摄取并表达以裸质粒DNA形式转入的外源基因。然而，对于编码分泌蛋白的基因而言，低转染效率可能并不妨碍分泌的基因产物的生物活性。因此，我们研究了以下假说：在动物后肢缺血模型中，用编码血管内皮生长因子（VEGF）的裸质粒DNA进行肌内（IM）基因治疗可促进侧支循环发育和组织灌注。
在一只兔的一侧后肢诱导缺血10天后，将500微克的phVEGF165或报告基因LacZ肌内注射到缺血后肢肌肉中。30天后，与对照组相比，VEGF转染组血管造影可识别的侧支血管和组织学可识别的毛细血管增多。这种血管增多改善了对缺血肢体的灌注，phVEGF165组的小腿血压比值（0.85±0.05）优于对照组（0.64±0.05，P<0.01），静息时缺血肢体的血流（用动脉内多普勒导丝测量）（phVEGF165=21.3±3.9 mL/min，对照组=14.6±1.6 mL/min，P<0.01）以及使用血管扩张剂后的血流（phVEGF165=54.2±12.0 mL/min，对照组=37.3±8.9 mL/min，P<0.01）均有所改善，并且缺血肢体的内收肌（phVEGF165=4.3±1.6 mL·min-1·100 g组织-1，对照组=2.9±1.2 mL·min-1·100 g组织-1，P<0.05）和腓肠肌（phVEGF165=3.9±1.0 mL·min-1·100 g组织-①，对照组=2.8±1.4 mL·min-1·100 g组织-1，P<0.05）中的微球数量增加。
缺血骨骼肌是裸质粒DNA基因治疗的一个有前景的靶点。对编码血管生成细胞因子的基因进行肌内转染，尤其是那些由完整细胞自然分泌的细胞因子基因，可能为患有广泛外周血管疾病且血管内导管介导的基因转移受限和/或被禁止的患者提供一种替代治疗策略。
