翻转消化法和灌注消化法分离培养兔耳缘静脉内皮细胞的比较  

Comparison of Techniques Irrigating and Reversing Rabbit Ear Vein to Digest, Isolate and Culture Endothelial Cells

作　　者：陈娟[1] 王廷华[2] 黄桂琴[2] 易伟斌[1] 徐岩泽[1] 罗哲文[1] 胡竹林[1] 

机构地区：[1]云南省红十字会医院眼科中心,昆明650031 [2]昆明医学院神经科学研究所

出　　处：《四川大学学报（医学版）》2007年第3期501-503,521,共4页Journal of Sichuan University(Medical Sciences)

基　　金：国家自然科学基金(批准号30360108)资助

摘　　要：目的比较翻转消化法和灌注消化法对培养血管内皮细胞生长的影响,以探讨培养血管内皮细胞的最佳方法。方法采用胰蛋白酶用翻转消化和灌注消化分别获取和培养兔耳缘静脉内皮细胞,并记录各组细胞生长情况,取第3代细胞用第Ⅷ因子相关抗原抗体鉴定所获得的细胞。结果两种消化方法首次获得的血管内皮细胞数量差别不大,但培养5d、10d后,灌注法组的细胞生长情况要明显优于翻转法组的细胞(P<0.01)。免疫组织化学检测细胞胞浆内Ⅷ因子相关抗原呈阳性。结论采用胰蛋白酶灌注法获取的内皮细胞生长情况要比翻转法获取的细胞好。Objeetive To establish the better method for isolating and culturing the endothelial cells （ECs）. Methods The ＂irrigative digestion＂ and ＂reverse interiorly-exteriorly digestion＂ methods were performed for digesting, isolating, collecting and culturing the ear vein endothelial cells of rabbit. The trypan blue stain was used to test the cell activity. The third-passage cell was identified by factor Ⅷ related antigen. The differences between the methods in cell number, activity and purity were compared to get an optimal method. Results The number of ECs deriving from the ＂irrigative digestion＂ method had no significant difference from that deriving from the ＂reverse interiorly-exteriorly＂ method when cells were isolated from rabbit ear vein originally. However, after cultured for 5 or 10 days, the vein endothelial cells from the ＂irrigative digestion＂ method showed the growth status superior to those from the ＂reverse interiorly-exteriorly＂ method. The cultured cells had a cobble stone appearance with a strict monolayer growth, it could be observed under inverted microscope. The factor Ⅷ related antigen was tested by immunohistochemistry, it supported that the cultured cells was ECs. Conclusion The ＂irrigative digestion＂ method is the better choice to isolate endothelial cells from small vessel.

关 键 词：耳缘静脉 血管内皮细胞 台盼蓝 翻转消化法 灌注消化法 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]