人血管抑素克隆表达及抑制血管生成的研究

Cloning and Expression of Recombinant Human Angiostatin and Its Antiangiogenic Activity

目的 研究并获得重组人血管抑素 (angiostatin) ,探讨其临床应用价值。方法 采用逆转录 -聚合酶链式反应 (RT PCR )方法 ,从人胚胎肝细胞中扩增出人全长血管抑素cDNA片段。用原核细胞表达载体构建 pBV 2 2 0 /angiostatin重组质粒 ,并将其转化大肠杆菌DH 5α进行表达 ,初步纯化。应用鸡胚尿囊膜血管生成实验及脐静脉内皮细胞增殖实验检测其活性。结果 经SDS PAGE分析 ,表达产物相对分子量约 3 8kD ,薄层扫描显示表达产物可达细菌总蛋白的 3 0 %。活性实验显示 ,重组蛋白能够抑制血管形成。结论 重组人Angiostatin在大肠杆菌中实现高效表达 。

Objective To express recombinant human angiostatin for further clinical application.Methods One amplification angiostatin cDNA fragment was obtained from the human embryo hepatic cells by RT-PCR method and cloned into prokaryotic to express vector pBV220 and transform into E.coli DH5α.The activities of angiostatin protein were examined by the endothelial cell inhibition assay and the chicken-embryo chorioallantoic membrane assay.Results SDS-PAGE analysis showed that expressed product was about Mr 38 kD and about 30% of bacterial total protein.Conclusion Recombinant human angiostatin can be expressed efficiently with biological active in E.coli DH5α.