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目的:探讨人胎盘来源的间充质干细胞(placenta derived mesenchymal stem cells,PDMSCs)在体外分化为血管内皮细胞的潜能。方法:利用组织块种植法体外分离培养PDMSCs,流式细胞术鉴定其表面抗原,应用含50μg/LVEGF和10μg/L bFGF的诱导分化液定向诱导PDMSCs向内皮细胞分化。免疫细胞化学检测分化过程中不同时点内皮特异性标志的表达变化。透射电镜和体外血管生成实验分别检测内皮特异性结构和内皮细胞功能。结果:PDMSCs表面抗原CD105和CD166阳性,CD31、CD34和CD45阴性。诱导分化的内皮细胞形态呈鹅卵石样,早期内皮标志Flk-1/KDR在4d表达最强;成熟的内皮标志CD31、vWF、CD144/VE-cadherin在内皮细胞分化过程中呈现时间依赖性表达(0d、4d、8d和12d)。透射电镜下可见内皮特异性结构:Weibel-Palade小体。细胞接种在细胞外基质凝胶中可形成毛细血管样结构。结论:胎盘中可分离出大量PDMSCs,并且PDMSCs在体外可分化为具有功能特性的内皮细胞,因此胎盘组织可能成为血管组织工程和再生医学的理想种子细胞来源。
Objective: To investigate the potential of human placenta derived mesenchymal stem cells (PDMSCs) differentiating into vascular endothelial cells in vitro. Methods: PDMSCs were isolated and cultured by tissue explants in vitro. The surface antigens were identified by flow cytometry. Differentiation of PDMSCs into endothelial cells was induced by differentiation medium containing 50μg / LVEGF and 10μg / L bFGF. Immunocytochemistry was used to detect the expression changes of endothelial specific markers at different time points during differentiation. Transmission electron microscopy and in vitro angiogenesis assay were used to detect endothelial specific structure and endothelial cell function respectively. Results: PDMSCs were positive for CD105 and CD166 but negative for CD31, CD34 and CD45. The differentiated endothelial cells were in the shape of pebbles, and the early endothelial marker Flk-1 / KDR was the strongest at 4 days. The mature endothelial markers CD31, vWF and CD144 / VE-cadherin showed time-dependent expression during endothelial cell differentiation , 4d, 8d and 12d). Transmission electron microscopy shows endothelial specific structure: Weibel-Palade body. Cells seeded in the extracellular matrix gel can form capillary-like structures. CONCLUSIONS: A large number of PDMSCs can be isolated from the placenta and PDMSCs can differentiate into endothelial cells with functional characteristics in vitro. Therefore, placenta tissue may be an ideal seed cell source for vascular tissue engineering and regenerative medicine.