体外诱导人脂肪干细胞向内皮细胞分化的研究

目的摘要目的诱导人脂肪间充质干细胞（hADAS cells）成内皮细胞，探索最佳诱导条件，为血管组织工程种子细胞的来源及其临床应用提供基础。方法分为内皮细胞生长因子（VEGF）50ng／ml和20ng／ml、碱性成纤维细胞生长因子（b-FGF）10ng／ml两组诱导脂肪干细胞成内皮细胞，流式细胞仪检测其表面抗原CD34、CD31表达，荧光显微镜下观察其吞噬乙酰化低密度脂蛋白（DiI—ac—LDL）和结合植物凝集素（FITC—Lectin）的能力，细胞免疫荧光法检测Ⅷ因子的表达。结果流式证实间充质干细胞在诱导第4和8天CD31、CD34表达逐渐增加；可吞噬DiI—ac—LDL，与Letin结合；细胞免疫荧光法可检测到Ⅷ因子表达。结论VEGF、b-FGF可诱导脂肪干细胞成内皮样细胞，对VEGF有很大的依赖性，成为血管组织工程种子细胞的选择之一。

Characteristics of Human adipose tissue-derived stem cells and the ability of differentiate into endothelial cells in vitro

Objective The aim of this study was to explore a approach to isolate and culcure human adipose-derived adherent stromal cell （hADAS） from adipose tissue and investigate suitable conditions for hADAS to differentiate into ceils of the endothelial lineage in vitro in ordor to use abdominal wall fat as a source of autologous stem cells for vascular tissue engineering. Methods Cultured hADAS were detected the cell surface antigens by flow cytometry. Cultured hADAS were divided into two groups：low concentrations group which added VEGF20ng/ml and b-FGF10ng/ml; high concentrations group which added VEGF50ng/ ml and b-FGF10ng/ml . The different positive rate of endothelial-specific markers CD34,CD31 were detected by flow cytometry. The positive rate of high concentrations group was higher than the other after four days. The positive rates of two groups had no obvious difference after eight days. The ability of phagocytosis DiI-ac- LDL and combine with FITC-Letin was observed under a fluorescence microscope. Ⅷ factor was detected by immunofluorescence. Results Flow cytometry showed that ： stromal cell-associated markers （ CD44, CD29） increased significantly with successive passages; CD34,CD31 were negative ; differentiated hADAS were positive for the markers CD34 ,CD31. Differentiated hADAS can phagocytosis DiI-ac-LDL and combine with FITC-Letin . Immunofluorescence analysis showed typical expression of the Ⅷ factor. Conclusions The proliferation ability of hADAS were very strong. They can be differentiated into endothelial-like ceils in the presence of VEGF ,b-FGF, have characteristics of endothelial ceils and represent an important source of cells for vascular tissue engineering. Low concentration of VEGF, and culture for a long time may be a suitable inducing conditions.