【摘要】 背景:种子细胞的数量和质量是制约血管组织工程研究的重要瓶颈,而如何获取干细胞源内皮细胞是解决该瓶颈问题的关键。目的:探讨添加血管内皮生长因子联合低氧环境是否能够协同高效诱导胚胎干细胞向内皮细胞定向分化。方法:采用无血清培养基m Te SR1维持培养人胚胎干细胞H9。通过添加血管内皮生长因子(50μg/L)联合低氧条件(体积分数为5%O2)诱导H9细胞向内皮细胞分化。采用免疫荧光染色技术、定量RT-PCR以及低密度脂蛋白摄取实验对人胚胎干细胞源内皮细胞进行表型和功能评价。结果与结论:添加血管内皮生长因子联合低氧培养条件可促进H9细胞向内皮细胞方向分化。该胚胎干细胞源内皮细胞不但高水平表达内皮细胞的标志基因(kdr,pecam)和标记蛋白D31,而且还能够摄取低密度脂蛋白,形成类似微血管结构。提示血管内皮生长因子与低氧在诱导干细胞向内皮细胞分化过程中存在协同效应,可高效获得具有理想表型和功能活性的干细胞源内皮细胞。更多还原

【Abstract】 BACKGROUND: The quantity and quality of seed cells is a critical bottleneck of the development of vascular tissue engineering. To address this issue, stem cell-derived endothelial cells have been a hot spot in this field due to their potential in providing the ideal seed cells. OBJECTIVE: To elucidate the effect of vascular endothelial growth factor(VEGF) supplementation combined with hypoxic culture condition on the lineage-specific differentiation of embryonic stem cells into endothelial cells. METHODS: Serum-free medium m Te SR1 was applied to cultivate H9 cells in vitro. A conditioned medium containing 50 μg/L vascular endothelial growth factor was utilized to induce H9 cells to differentiate into endothelial cells under the hypoxic culture condition(5% O2). The cell under normal condition(5% CO2) with or without vascular endothelial growth factor served as controls. The phenotype and function of human embryonic stem cells-derived endothelial cells were assayed by immunofluorescence staining, quantitative RT-PCR, and low-density lipoprotein uptake experiment. RESULTS AND CONCLUSION: Compared with the control group, the H9 cells were induced to be differentiated into endothelial-like cells more efficiently when they were cultivated under a conditioned medium with vascular endothelial growth factor supplementation under the hypoxic condition. These differentiated cells not only expressed some important surface markers of endothelia cells, including kdr, pecam, but also took in low-density lipoprotein to form microvessle-like structures. This culture system supports a synergy effect of vascular endothelial growth factor and hypoxic environment that can efficiently promotes the lineage-specific differentiationof embryonic stem cells into endothelial cells with good phenotype and functionality.更多还原