人脐静脉内皮细胞的分离及原代培养体会

Experience on the isolation and cultivation of the human umbilical vein endothelial cells in vitro

目的:建立一种操作简便、且能够获得数量较多及活力良好的人脐静脉内皮细胞的分离、培养方法。方法:采用0.1%IV型胶原酶灌注消化分离人脐静脉内皮细胞,将其置于含有20%胎牛血清、内皮细胞生长因子及肝素的M199培养基内培养,倒置显微镜观察内皮细胞生长情况,免疫荧光方法鉴定内皮细胞的Ⅷ因子相关抗原。结果:细胞呈梭形或椭圆形生长,单层铺路石状排列或呈漩涡状排列。免疫荧光检测结果显示细胞的胞浆内有大量绿色荧光颗粒,Ⅷ因子相关抗原呈阳性反应。结论:实验建立的人脐静脉内皮细胞分离和培养方法简单、可靠,且能获得足够数量和高纯度的血管内皮细胞,为进一步研究人体大血管内皮细胞的功能奠定了较好的基础。

Objective：To establish the methods of isolating and culturing human umbilical vein endothelial cells （HUVECs） in vitro. Methods： HUVECs were acquired by type IV collagenase solution into the lumen of umbilical veins and then cultured in M199 with 20% FBS, ECCS and heparin. They were identified by cell morphology and Ⅷ factor immunofluorescence. Results： HUVECs were presented as Cobblestone arrangement, and the antigens of Ⅷ factor were positive according to the immunofluoresccnce. Conclusion： A large number of highly purified endothelial cells could be acquired by digestion of collagenase, which can successfully establish the model for research of ECs.