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目的:研究人脐血CD34~+细胞群体中内皮祖细胞在体外分化为内皮细胞的过程中,干细胞标志以及内皮细胞表型随时间的变化。方法:将免疫磁珠细胞分选法(MACS)得到的CD34~+细胞体外培养于纤连蛋白和无纤连蛋白处理的培养皿中,以免疫细胞化学鉴定贴壁细胞的内皮标志Flk-1和vWF,并以流式细胞仪分析其干细胞标志AC133。结果:贴壁细胞的内皮标志Flk-1和vWF是逐步出现的,d3时有27.0％贴壁细胞表达Flk-1,vWF不表达,d7时已100％表达vWF和Flk-1。纤连蛋白促进贴壁细胞内皮标志Flk-1和vWF的表达,d3时的表达百分率分别为34.0％和47.0％,d7时Flk-1和vWF的表达均为100％。在培养过程中,AC133阳性细胞的比例迅速下降,但纤连蛋白对AC133的表达无显著影响。结论:在内皮祖细胞分化的过程中,干细胞标志迅速消失,向内皮细胞分化,内皮细胞的表型是逐步出现的。纤连蛋白促进内皮祖细胞的分化。
OBJECTIVE: To study the changes of endothelial cell phenotypes and endothelial cell phenotypes during the differentiation of endothelial progenitor cells into endothelial cells in human cord blood CD34 + cell population in vitro. Methods: The CD34 + cells obtained from MACS were cultured in fibronectin and fibronectin-treated dishes in vitro. Immunocytochemistry was used to identify the endothelial marker Flk-1 And vWF, and analyzed their stem cell marker AC133 by flow cytometry. RESULTS: Flk-1 and vWF, the endothelial markers of adherent cells, were gradually developed. Flk-1 was expressed in 27.0% of adherent cells on day d3, while vWF was not expressed on day d7. Fibronectin promoted the expression of Flk-1 and vWF, which were 34.0% and 47.0% on d3 and 100% on d7. During the culture, the proportion of AC133-positive cells decreased rapidly, but fibronectin had no significant effect on the expression of AC133. CONCLUSIONS: During the differentiation of endothelial progenitor cells, the stem cell marker disappears rapidly and differentiates into endothelial cells. The phenotype of endothelial cells is gradually emerging. Fibronectin promotes differentiation of endothelial progenitor cells.