大鼠骨髓内皮祖细胞的分离培养及其生物学特性

目的：探讨大鼠骨髓内皮祖细胞的分离培养方法及其生物学特性。方法：采用二次贴壁法体外扩增培养大鼠骨髓内皮祖细胞,观测细胞增殖能力;免疫化学检测细胞CD133、CD34、Flk1、CD31的表达;三维培养细胞并观察其体外成血管能力。结果：细胞培养第4天集落样生长,呈圆形或梭形;第10天细胞长满瓶底,呈鹅卵石样外观。原代血管内皮祖细胞CD133、CD34、Flk1均表达阳性;第2代细胞CD34、CD31、Flk1均表达阳性,CD133表达阴性。三维培养第2代血管内皮祖细胞在胶原内有＂出芽式＂生长现象及管腔样结构形成。结论：体外扩增法可以从骨髓中分离培养出内皮祖细胞,三维培养发现其具有体外成血管能力。

Isolating culture and characteristics of rat-marrow-derived endothelial progenitor cells

Objective：To investigate the isolating culture and characteristics of endothelial progenitor cells （EPCs） derived from rat bone marrow. Methods：The EPCs of rat bone marrow were cultured by second adherent amplification in vitro. The capacity of the cells＇ proliferation was observed daily. These cells were performed immunofluorescence staining to detect the expression of CD133, CD34, Flkland CD31. The capability of folming blood vessels was observed through three-dimensional cultured cells. Results： Cell colonyforming units（CFUs） appeared at the 4th day. At the center of CFUs, cells were round, but around the CFUs, cells were fusiform shaped. Up to the lOth day,cells covered the bottom of the bottle like cobblestones. The proto-passage EPCs expressed CD133, CD34 and Flkl. And the 2nd passage cells expressed CD34, CD31 and Flkl. The 2nd passage EPCs by three-dimensional culture manifested sprouted EPCs and formed tube-like structures in collagen. Condusions：EPCs can be gained from bone marrow through proliferation of cells ex vivo and these cells has the capability of forming blood vessel.