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人前胰岛素原基因裸质粒DNA肌肉注射可显著降低链脲佐菌素诱发糖尿病小鼠的血糖水平

王留义, 孙威, 陈明哲, 王宪

北京大学第三医院血管医学研究所.北京 100083

摘要

通过基因治疗的方法补充胰岛素已用于实验性治疗胰岛素依赖型糖尿病(IDDM)。该研究构建了含有重组人前胰岛素原基因的裸质粒DNA载体(pCMV-IN),将其肌肉注射入链脲佐菌素(STZ)诱发的糖尿病C57小鼠体内,并辅以电穿孔方法,以获得在体胰岛素转基因治疗。该质粒载体表达的胰岛素mRNA,可通过RT-PCR方法在转基因局部的骨骼肌组织中检测到。在接受pCMV-IN注射的糖尿病小鼠中,血浆胰岛素水平显著升高,达到了未注射STZ的正常对照小鼠的水平,且胰岛素的表达可持续至少35d。pCMV-IN质粒注射转基因治疗显著降低了糖尿病小鼠在d7至d35的血糖水平,其下降幅度约6mmol/L;转基因治疗也显著降低了严重糖尿病小鼠的死亡率,其第6周时的死亡率由100%降为37%。结果表明,直接肌肉注射含人前胰岛素原基因裸质粒可获得胰岛素的有效表达,显著降低糖尿病小鼠的血糖水平并降低严重糖尿病小鼠的死亡率。裸质粒注射胰岛素转基因治疗有望成为IDDM的一种有效治疗手段。

关键词： 基因治疗; 胰岛素; 糖尿病

Intramuscular injection of naked plasmid DNA encoding human preproinsulin gene in streptozotocin-diabetes mice results in a significant reduction of blood glucose level

Wang Liuyi, Sun Wei, Chen Mingzhi, Wang Xian

Institute of Vascular Medicine,Third Hospital,Peking University.Beijing 100083

Abstract

The insulin complement with gene therapy has been used as an experimental treatment for insulin dependent diabetes (IDDM). In the present study, we constructed naked plasmid DNA vector encoding recombinant human preproinsulin gene (pCMV-IN), and injected the plasmids (100 #mu#g/mouse) intramuscularly combined with electroporation, to achieve the in vivo transfer of insulin gene in streptozotocin (STZ)-induced diabetic C57 mice. The expression of vector-derived insulin mRNA was detected with RT-PCR in transfected local skeletal muscles. The plasma insulin was elevated significantly in pCMV-IN injected diabetic C57 mice, which was complemented to the level similar to the intact normal control. The protein expression lasted for at least 35 days after the plasmid injection. Gene therapy with pCMV-IN plasmids considerably decreased the blood glucose level in STZ-induced diabetic mice from d 7 to d 35 by about 6 mmol/L. The gene therapy also reduced the mortality of severe diabetic mice significantly from 100% to 37% at the 6th week. Our results indicate that the direct intramuscular injection of naked plasmids encoding human preproinsulin gene achieves the effective expression of insulin. The restoration of insulin decreases blood glucose and increases the survival in severe diabetic mice. The gene therapy might be provided as practical therapeutic approach to IDDM.

Key words: Plasmid;Insulin;Diabetes

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引用本文：

王留义, 孙威, 陈明哲, 王宪. 人前胰岛素原基因裸质粒DNA肌肉注射可显著降低链脲佐菌素诱发糖尿病小鼠的血糖水平[J]. 生理学报 2003; 55 (6): .

Wang Liuyi, Sun Wei, Chen Mingzhi, Wang Xian. Intramuscular injection of naked plasmid DNA encoding human preproinsulin gene in streptozotocin-diabetes mice results in a significant reduction of blood glucose level. Acta Physiol Sin 2003; 55 (6): (in Chinese with English abstract).