兔主动脉内皮细胞制备方法的优化

An optimized method for the preparation of aortic endothelial cells from rabbits

目的：为构建骨组织工程血管化提供优质种子细胞。方法采用血管内膜外翻和胶原酶消化法分离培养兔主动脉内皮细胞并传代；采用倒置显微镜、透射电镜观察和免疫荧光法鉴定。结果培养的血管内皮细胞在第3-5天为对数生长期，倒置显微镜下可见内皮细胞呈紧密排列、短梭多角形的“鹅卵石”形态；透射电镜可见内皮细胞特征性Weible-Palade小体；经免疫荧光法鉴定证明该细胞CD31抗原阳性。结论采用血管内膜外翻等方法制备的兔血管内皮细胞的成活率高，且纯度高、数量多、具有良好的生物学活性。该法可为骨组织工程血管化的研究提供种子细胞。

Objective To provide high-quality seed cells for the vascularization of bone tissue engineering. Methods Processing includes intimal eversion and collagenase digestion, endothelial cells （ ECs） harvested from rabbit aorta were isolated,cultured and passaged. ECs were identified by inverted microscope, transmission electron microscopy and immunofluorescence. Results The cultured vascular endothelial cells entered the logarithmic phase on the third to fifth day. The cells displayed the typical endothelial phenotype of closely opposed polygonal cells with very clear and well-defined cell border （ cobble-stone appearance） in inverted microscope. ECs identitied as Weibel-Palade bodies in transmission electron microscopy and showed CD31 （ ＋） in immunofluorescence. Conclusions It is ob-tained that a good cellular activity with a high purity using a new and easy method. The cells display a high cellular yield and proliferation. These cells can be used for the vascularization of bone tissue engineering.