人脂肪干细胞向内皮细胞的定向分化

目的 为解决血管组织工程种子细胞来源不足问题,分离和培养人脂肪干细胞(hADSCs),并在体外定向诱导hADSCs分化为内皮细胞.方法 利用胶原酶消化法和贴壁筛选法从人脂肪组织中分离、培养及扩增hADSCs;应用纤维粘连蛋白(FN)与富含多种生长因子的内皮细胞支持液EGM2-MV及高浓度血管内皮细胞生长因子(VEGFi6s)50 ng/ml,协同定向诱导hADSCs分化为内皮细胞,然后对其进行形态、表型及功能鉴定.结果 体外分离、培养出高度同源性的hADSCs,hADSCs定向内皮分化后免疫组化及RT-PCR结果显示,内皮特异性标志物CD31、CD34及血管内皮细胞生长因子受体(KDR)阳性表达,透射电镜下观察到内皮特异性结构怀布尔-帕拉德(Weibel-Palade)小体,内皮细胞功能性检测结果显示诱导后hADSCs能够吞噬Dil标记的乙酰化的低密度脂蛋白(Dil-Ac-LDL)及在基质胶(Matrigel)上形成小管样结构,成管数(18.9±1.203)、分支数(31.2±1.304)、结点数(16.9±1.183),与HUVEC组比较差异均无统计学意义(P＞0.05).结论 在体外可成功诱导hADSCs分化为内皮细胞.

Abstract：

Objective To investigate the possibility of directional differentiation of human adipose stem cells (hADSCs) into endothelial cells (EC), so as to provide seed cells for tissue engineered vessels.Methods hADSCs were isolated from human adipose tissue by collagenase digestion, cultured and amplified by adherence to flasks. Then hADSCs were directionally induced to differentiate into EC by a combination of fibronectin ( FN ), endothelial cells support liquid ( EGM2-MV ) containing various growth factors and high concentration of VEGF165 (50 ng/ml). Then, the cells morphology, phenotype and function were identified. Results Highly homologous hADSCs were obtained, and then hADSCs were directionally differentiated into EC. CD31 and CD34, the specific markers for EC, and vascular endothelial growth factor receptor (KDR) were positive by immunohistochemical staining and RT-PCR. In addition,unique Weibel-Palade bodies in EC were observed under transmission electron microscope. Functionally,hADSCs could swallow Dil-Ac-LDL and form tube-like structures in matrigel after endothelial differentiation. Conclusions hADSCs can be successfully induced to differentiate into endothelial cells in vitro.

Directional differentiation of human adipose-derived stem cells into endothelial cells

Objective To investigate the possibility of directional differentiation of human adipose stem cells (hADSCs) into endothelial cells (EC), so as to provide seed cells for tissue engineered vessels.Methods hADSCs were isolated from human adipose tissue by collagenase digestion, cultured and amplified by adherence to flasks. Then hADSCs were directionally induced to differentiate into EC by a combination of fibronectin ( FN ), endothelial cells support liquid ( EGM2-MV ) containing various growth factors and high concentration of VEGF165 (50 ng/ml). Then, the cells morphology, phenotype and function were identified. Results Highly homologous hADSCs were obtained, and then hADSCs were directionally differentiated into EC. CD31 and CD34, the specific markers for EC, and vascular endothelial growth factor receptor (KDR) were positive by immunohistochemical staining and RT-PCR. In addition,unique Weibel-Palade bodies in EC were observed under transmission electron microscope. Functionally,hADSCs could swallow Dil-Ac-LDL and form tube-like structures in matrigel after endothelial differentiation. Conclusions hADSCs can be successfully induced to differentiate into endothelial cells in vitro.