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目的:研究腺病毒介导的血管内皮因子受体(KDR)启动子-胸苷激酶系统对血管内皮细胞的旁观者效应。方法:采用新型腺病毒载体AdEasy系统,构建由KDR启动子调控的、腺病毒介导的、单纯疱疹病毒胸苷酸激酶基因系统(AdK- DR-HSV-tk),在人胚胎肾细胞系293细胞中包装、扩增后,体外感染表达KDR的人脐静脉血管内皮细胞(HUVEC),建立HUVEC/tk转化细胞株。观察丙氧鸟苷((GCV)对HUVEC、HUVEC／tk细胞的存活率及旁观者效应。结果:从病毒滴度及PCR检查,证实tk基因随病毒的感染已成功地导入293细胞及HUVEC细胞。HUVEC／tk细胞对丙氧鸟苷的敏感性显著高于亲代HUVEC细胞。HUVEC／tk及HUVEC+HUVEC／tk混合细胞还随丙氧鸟苷浓度的增加受到明显的抑制。HUVEC+HUVEC／tk混合细胞的存活率,随HUVEC／tk细胞比例的增加而降低,说明腺病毒介导的KDR启动子-单纯疱疹病毒胸苷激酶／丙氧鸟苷系统不但能杀伤tk+细胞,也能杀伤未导入tk的细胞,存在明显的旁观者效应。结论:腺病毒介导的KDR启动子单纯疱疹病毒胸苷激酶／丙氧鸟苷系统对血管内皮细胞具有明显的杀伤及旁观者效应。
AIM: To study the bystander effect of adenovirus-mediated vascular endothelial factor receptor (KDR) promoter-thymidine kinase system on vascular endothelial cells. Methods: A novel adenoviral vector AdEasy system was used to construct adenovirus-mediated, herpes simplex virus thymidylate kinase gene system (AdK-DR-HSV-tk) regulated by KDR promoter. Cells were packaged, expanded, and in vitro transfected into KDR-expressing human umbilical vein endothelial cells (HUVECs) to establish HUVEC / tk transformed cell lines. To observe the survival rate and bystander effect of GCV on HUVEC and HUVEC / tk cells.Results: The tk gene was successfully transfected into 293 cells and HUVEC cells by virus titer and PCR .The sensitivity of HUVEC / tk cells to ganciclovir was significantly higher than that of the parental HUVECs.HUVEC / tk and HUVEC + HUVEC / tk mixed cells were also significantly inhibited with the increase of ganciclovir concentration.HUVEC + HUVEC / tk Mixed cell survival rate, with the proportion of HUVEC / tk cells decreased, indicating that the adenovirus mediated KDR promoter - herpes simplex virus thymidine kinase / ganosine system not only kill tk + cells, but also can not kill the import tk cells, there is a significant bystander effect.Conclusion: Adenovirus-mediated KDR promoter herpes simplex virus thymidine kinase / ganoside system has obvious killing and bystander effect on vascular endothelial cells.