人脐血内皮祖细胞体内促进血管重建的实验研究

目的：通过观察人脐血内皮祖细胞（EPC）在裸鼠体内缺氧状态下分化成内皮细胞的现象，初步研究其促进血管重建的作用。方法：根据EPC表面标记CD133，采用免疫磁珠法分离培养EPC。设计EPC裸鼠成瘤实验，观察瘤体内血管增生情况；将荧光标记的EPC注射于裸鼠腹部皮瓣中，观察EPC参与皮瓣血管重建情况；将EPC接种于聚羟基乙酸（PGA）中，移植于裸鼠皮下，观察EPC参与血管重建。结果：原代EPC贴壁后呈梭形，从第7天开始数量明显增加，呈克隆状生长。透射电镜观察到成熟内皮细胞最具特征性的细胞器——weibel-Palade小体。裸鼠成瘤实验：抗人vWF免疫荧光显示大量EPC参与肿瘤血管生成；裸鼠皮瓣实验：荧光标记EPC参与裸鼠皮瓣血管生成；PGA移植实验：抗人vWF免疫酶组织化学染色显示EPC参与血管生成。结论：EPC参与血管重建，具有促进血管新生，加速缺血组织血管化的作用。

Differentiation and expansion of endothelial progenitor cells from human umbilical cord blood

Objective To study EPC biological characteristics and its neovascularization in athymic mice. Methods CD133+ cells were enriched from human umbilical cord blood by immunomagnetic sorting, and cultured with EGM-2MV media. The mixture of EPCs and human stomach cell line GC7901 were injected into athymic mice to observe the tumor growth and vascularization in tumor. The EPC labeled with PKH26 (Fluorescent Cell Linker) were injected into the over-length flap models made on athymic mice, or seeded into dermal substitute constructed by PGA. Observing the EPCs trace and their participating in the flap vascularization were done with fluorescent microscope. Results The culture cells showed a typical spindle-shaped morphology at 1 week and cluster areas of cobblestone-like cells after 2 weeks. The cells after 10 days culture have typical Weibel-Palade body observed under transmission electron microscope. Tumor forming experiment in athymic mice showed many human vWF antigen-positive endothelial cells involved in the tumor vascularization. Immunohistological and label fluorescent analyses showed vWF antigen-positive cells and labeled cells constructing blood vessels in flap and in PGA dermal substitute. Conclusion EPC might contribute to angiogenesis, and speed up ischemic tissue vascularization.