肌肉电针介导血管内皮生长因子基因治疗猴闭塞性血管病的实验研究Experimental study on treatment of limb ischemia with electro-acupuncture-mediated VEGF gene
汤健,周爱儒,鲁东成,高炜,霍勇,彭旭,朱国英
摘要(Abstract)：
目的 利用一种新的基因转移方法探讨血管内皮生长因子（ＶＥＧＦ）基因治疗猴闭塞性血管病的可行性。方法构建重组ＶＥＧＦ基因的真核表达载体。转基因前３天肌肉内注入肌肉再生剂了哌卡因（ｂｕｐｉｖａｃａｉｎｅ），通过电针向血管闭塞症猴（ｎ＝６）骨骼肌内转移重组ＶＥＧＦ基因（１６００μｇ／只），应用血管造影、局部血管染色及超声多普勒等方法观察ＶＥＧＦ基因导入猴体内后的生物效应，并利用临床自动生化分析仪、病理切片和Ｂ超进行转基因的安全性研究。结果转ｐｃＤ２／ＶＥＧＦ基因２８天后，血管造影可见明显的新生血管和侧支循环形成，超声多普勒发现血流明显恢复，３个月后更显著，血管染色记数两组间差异有显著性（ＰｃＤ２／ＶＥＧＦ组４０．３±９．４３／ｍｍ２；ｐｃＤ２组２５．９±９．００／ｍｍ２，Ｐ＜０．０１）。血液生化测定、主要脏器病理切片和Ｂ超检查未发现异常。结论利用电针向肌肉内转移ＶＥＧＦ基因可促进局部组织新生血管形成和侧支循环建立，促进血流恢复，且安全、简便，为闭塞性血管病的基因治疗提供一种新的基因转移方法。
关键词(KeyWords)： 电针;血管内皮生长因子;基因疗法;侧支循环
基金项目(Foundation): 国家八六三项目
作者(Author): 汤健,周爱儒,鲁东成,高炜,霍勇,彭旭,朱国英
参考文献(References)：
- 1 European Working Group on Critical Ischemia. SecondEuropean consensus document on chronic critical leg ischemia. Circulation, 1991, 84 (supple 4): 4-26.
- 2 Tischer E, Mitchell R, Dominic JW, et al. The humangene for vascular endothelial growth factor: multipleprotein forms are encoded through alternative exon splicing. J Biol Chem, 1991,266: 11947-11954.
- 3 Thomas KA. Vascular endothelial growth factor, a potent and selective angiogenic agent. J Biol Chem, 1996,271: 603-606.
- 4周爱儒,郑卫,邢德智,等.分子搭桥术-VEGF基因治疗闭塞性血管病的实验研究.中华医学杂志,1996,76:662- 666.
- 5 Isner JM, Pieczek A, Schainfeld R, et al. Clinical evidence of therapeutic angiogenesis in patients with criticaltimb ischemia after intra-muscular phVEGF165 genetransfer. Lancet, 1996, 348: 370-374.
- 6卢圣栋,主编.现代分子生物学实验技术.北京:高等教育 出版社,第1版,1993,111-118.
- 7 Evic Van, Belle Bernhard, Witzenbihler, et al. Potentiated angiogenic effect of scatter factor/hepatocytegrowth factor via induction of vascular endothelialgrowth factor. Circulation, 1998, 97: 381-390.
- 8 Banal S, Jaklitsch MT, Shou M, et al. Angiogenic-in-duced enhancement of collateral blood flow to ischemicmiocardium by vascular endothelial growth factor indogs. Circulation, 1994, 89: 2183-2190.
- 9张曼,周爱儒,汤健,等.血管内皮生长因子在基因治疗兔髂动脉内膜损伤的实验研究.中华医学杂志,1998,26: 32-35.
- 10 Isner JM, Walsh K, Sywmes JF, et al. Clinical protocol: Arterial gene transfer for therapeutic angiogenesis inpatients with peripheral artery disease. Human GeneTherapy, 1996, 7: 959-988.
- 11 Wolff JA, Dowty ME, Jiao SS, et al. Expression ofnaked plasmids by cultured myotubes and entry of plasmids into tubules and caveolae of mammalian skeletalmuscle. Science, 1992, 103: 1249-1257.
- 12 Yukio Tsurumi, Satoshi Takeshita, Dongfen Chen, etal. Direct intramuscular gene transfer of naked DNA encoding vascular endothelial growth factor augments collateral development and tissue perfusion. Circulation,1996, 94: 3281-3290.