三种分离大鼠主动脉内皮细胞方法的比较研究

目的:建立一种有效的、重复性好的大鼠主动脉内皮细胞(RAEC)分离方法。方法:采用外翻胶原酶消化法、瞬间热处理植环法和植块法分离RAEC,并进行培养、纯化和传代,观察细胞生长和形态,用免疫组织化学法检验Von Villebrand Factor/Ⅷ因子相关抗原进行内皮细胞鉴定。结果:采用植块法分离RAEC,5~7天后可见细胞贴壁生长,14天呈单层铺路石样分布,Ⅷ因子相关抗原阳性细胞(胞浆呈棕黄色)比率为52.3±10.6%,其他两种方法分离后未见上皮细胞贴壁生长。结论:比较外翻胶原酶消化法、瞬间热处理植环法,植块法分离RAEC能培养获得较多内皮细胞,且方法较简便,细胞污染和损伤较少,重复性好。

Compared Study for Three Methods to Isolate Endothelial Cells from Rat Aorta

Objective: To develop a effective method with good reproducibility to isolate primary rat aortic endothelial cells(RAECs).Method: Enzyme digestion, planting loop method with moment heat treatment and planting clump method were used to isolate the RAECs. Cultured,purified and subcultured RAECs,then observed the cell growth and morphology and identified the RAECs by immunohistochemical method. Results: Using planting clump method, RAECs could emigrate from explants for 5~7 d and reached monolayer cobblestone-like for 14 d. RAECs were identified by immunohistochemical method, the positive reaction is showed in the cytoplasm, the rate of positive cells were 52.3 ±10.6%.Conclusion: Compared with enzyme digestion and planting loop method with moment heat treatment, planting clump method has more convenience and better reproducibility.