骨髓间充质干细胞自体移植促进缺血肢体血管生成的实验研究

Experimental Study of Implantation of Autologous Bone Marrow Mesenchymal Stem Cells to Promote Angiogenesis in Ischemic Limbs

目的探索骨髓间充质干细胞在血管生成中的作用,为临床建立一种简便安全的下肢动脉缺血性疾病的治疗途径。方法24只新西兰兔分为实验组(12只)和对照组(12只),分离培养实验组兔骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs),Brdu标记,建立兔后肢缺血动物模型,将取于自体的骨髓间充质干细胞制成悬液注射于实验组兔缺血部位,对照组注射等量生理盐水。2周后行实验组和对照组兔股动脉的二维及彩色多普勒超声检测,观察移植前后的血管内径,血流峰值速度,血流加速时间等;取缺血部位肌肉组织,免疫荧光染色观察移植细胞分布及血管生成情况。结果细胞移植后2周,实验组的股动脉内径,血流峰值速度,加速时间均大于对照组,有显著性差异(P<0.01);免疫荧光染色结果显示移植部位有移植细胞存在,移植组血管生成情况明显优于未移植组。结论骨髓间充质干细胞具有促进血管生成的作用,自体移植可望成为一种简单的、有效的治疗下肢缺血的方法。

Objective To investigate the mechanism of bone marrow mesenchymal stem cells in angiogenesis and to find an easier and safer method in treatment of ischemic arterial disease of the lower extremity. Methods Bone marrow mesenchymal stem cells were isolated and cultured from New Zealand rabbits and these cells were marked with Brdu. Autologous bone marrow mesenchymal stem cells were injected into the ischemic hind limb muscles of ischemic hind limb animal model （ n = 12） with same volume of normal saline as control group （ n = 12）. Two weeks after the implantation of autologous bone marrow mesenchymal stem cells, 2D and color Doppler ultrasound detection were used on rabbit arteria cruralis of experimental group and control group to observe the inner diameter of blood vessel, the peak value velocity and the acceleration time of blood flow. Obtain muscles from ischemic tissue then observe the disposition of transplant cells and the state of angiogenesis using immunofluorescence staining. Results The results of Doppler ultrasound detection show that the inner diameter of blood vessel, the peak value velocity and the acceleration time of blood flow of experimental group rabbits were larger than those of control group obviously（ P 〈 0.01）. The result of immunofluorescence staining shows that transplant cells exist within transplant part and state of angiogenesis in experimental group was superior to control group obviously. Conclusion Bone marrow mesenchymal stem cells have the effect to promote angiogenesis, implantation of autologous bone marrow mesenchymal stem cells was a simple and effective method to treat ischemic hind limb.