Ad－HGF基因修饰的胎盘间充质干细胞治疗兔肢体缺血的实验研究

摘要

目的：研究重组腺病毒介导肝细胞生长因子（ adenoviral vector mediated human hepatocyte growth factor，Ad－HGF）修饰的胎盘源间充质干细胞（ placenta－derived mesenchymal stem cells，PMSC）在兔肢体缺血模型中促新生血管生成的作用。方法无菌取足月产妇胎盘胎儿面中心区域胎盘组织，胶原酶消化法分离干细胞，体外培养传代，感染Ad－HGF 48 h后收集细胞用于治疗兔肢体缺血实验。左侧治疗组后肢缺血肌肉组织内多点注射总细胞数5×106／ml Ad－HGF修饰的PMSC，右侧对照组后肢缺血肌肉组织内注射生理盐水。结果治疗后第14天腹主动脉穿刺行数字减影血管造影（ digital subtraction angiography，DSA）检查，可见左侧肢体缺血治疗后微血管生成数明显多于未治疗右侧肢体，血管形成能力明显增强。苏木精伊红染色法（ HE）后置光镜下观察显示，治疗组毛细血管密度明显大于未治疗组（P＜0．05）。实时荧光定量聚合酶链反应（quantitative polymerase chain reaction，qPCR）检测后肢肌肉组织中血管内皮生长因子（ vascular endothelial growth factor， VEGF）、碱性成纤维细胞生长因子（ basic fibroblast growth factor，bFGF）、HGF的表达明显升高（P＜0．05）。结论经Ad－HGF基因修饰的PMSC能明显促进血管新生，加快局部缺血组织血流循环的重建，是治疗肢体慢性缺血的有效手段。%Objective To evaluate the therapeutic effect of hepatocyte growth factor(HGF) gene modified placenta-derived mesenchymal stem cells( PMSCs) on limb ischemia in a rabbit model.Methods The placental tissue was digested with enzyme, cultured and passaged.The PMSCs were characterized by surface marker expression.These cells were infected with adenoviral( Ad)-HGF and intramuscular injected for treatment of limb ischemia in a rabbit model.The blood supply of the limb was detected by digital subtraction angiography ( DSA ) and the vessel number was evaluated in histopathological HE staining.Results The results showed that Ad-HGF gene transduction increased the vascular endothelial growth factor ( VEGF ) , basic fibroblast growth factor, bFGF ( bFGF ) and HGF expression in PMSCs. Transplantation of HGF-transduced PMSCs resulted in the increase in vessel density and improvement of blood supply in the rabbit limb ischemia model.Conclusion The therapeutic effect of HGF gene engineered PMSCs on ischemia by enhancing angiogenesis in a rabbit model is evaluated.Transplantation of PMSCs with HGF gene therapy may be a promising strategy for the treatment of ischemia diseases.