骨髓内皮祖细胞促带蒂皮瓣血管化的实验研究

Experimental study on skin flap angiogenesis promotion using bone marrow derived endothelial progenitor cells

目的研究大鼠骨髓来源的内皮祖细胞(EPC)对皮瓣移植后血管化的影响,探讨提高移植皮瓣存活率的方法。方法将包含双侧腹壁下动脉的带蒂皮瓣内右侧血管保留,建立皮瓣局部缺血的Wistar大鼠动物模型。冲洗大鼠骨髓腔,密度梯度离心获得单个核细胞;通过fibronectin包被的培养皿,差速分离快速黏附细胞;鉴定细胞CD34、CD133及VEGFR-2表型,并将目的细胞注射于移植后皮瓣不同部位。对皮瓣进行大体观察,计算局部毛细血管密度。以皮瓣移植后不做任何处理或仅注射普通培养液的Wistar大鼠作为对照组。结果原代EPC快速黏附细胞与体外培养7 d的EPC表达CD34、CD133和VEGFR-2表型,两者阳性表达率无统计学差异(P>0.05);皮瓣内注射EPC部位的存活面积以及毛细血管密度显著高于对照部位(P<0.05)。结论富集的EPC皮瓣内注射显著提高了移植皮瓣的存活率。

Objective To investigate the implanted skin flap angiogenesis affected by rat derived endothelial progenitor cells （EPCs） , and to explore the method for improving skin flap survival rate. Methods Wistar rat skin flap model with topical ischemia was established by cutting off left side inferior epigastric artery. EPCs were harvested by flushing bone marrow cavity and were isolated by density gradient centrifugation. The isolated cells were further purified and enriched by different adhension speed to fibronectin coating dish. Flow cytometer was applied to identify the phenotype of CD34, CD133 and VEGFR-2 of the isolated cells. Cells with different adhension speed were injected into different skin flap sites. Gross observation and capillary vessel density evaluation was conducted. Wistar rats with no treatment or rountine culture fluid injection after skin flap implant were served as controls. Results Both the primary EPCs and EPCs cultured for 7 d in vitro expressed the phenotype of CD34, CD133 and VEGFR-2 （ P 〉 O. 05）. The survival area and capillary density were more favorable in the EPCs-injection sites than the controls（P 〈 0. 05）. Conclusion Injection of enriched EPCs can significantly improve the survival rate of implanted skin flap.