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目的探讨做为微环境的滋养层细胞在体外大量扩增晚期内皮祖细胞(EPC)中的作用。方法预铺辐射后的滋养层细胞,而后植入分离的外周血单个核细胞,计数培养21 d后获得的晚期EPC克隆数目,并检测目的细胞免疫表型,用Matrigel鉴定其体外血管形成能力。采用动态显微镜追踪单个干细胞在滋养层细胞上的分化过程。结果培养21 d时,在预铺晚期EPC和人脐静脉内皮细胞(HUVEC)为滋养层的培养环境下,每100ml外周血单个核细胞分别获得(40.0±3.9)和(39.3±3.1)个晚期EPC克隆,均明显高于无滋养层条件下的(2.0±1.3)个(P均<0.05)。目的细胞可表达CD31、CD34、eNOS、FLt-1、P1H12、Sendo、VEcadherin和CD117,并在体外与Matrigel形成管状结构。动态追踪示单个干细胞的扩增是在有滋养层细胞参与下完成不对称分裂的。结论通过提供滋养层细胞方法可以大量扩增晚期EPC,滋养层细胞可能参与了早阶段内皮干细胞的不对称分裂扩增。
Objective To investigate the role of trophoblast cells as a microenvironment for large-scale amplification of advanced endothelial progenitor cells (EPCs) in vitro. Methods The irradiated trophoblast cells were pre-plated. Then the isolated peripheral blood mononuclear cells were implanted. The number of late EPCs obtained after 21 days of culture was counted. The immunophenotypes of the cells were detected, and their in vitro vascularization ability was identified by Matrigel. Dynamic microscopy was used to track the differentiation of individual stem cells on trophoblast cells. Results At 21 days of culture, perilymph of peripheral blood mononuclear cells (40.0 ± 3.9) and (39.3 ± 3.1), respectively, were obtained in the culture of pre-plated EPCs and human umbilical vein endothelial cells (HUVECs) EPCs were significantly higher than those in the absence of trophoblast (2.0 ± 1.3) (all P <0.05). The target cells express CD31, CD34, eNOS, FLt-1, P1H12, Sendo, VE cadherin and CD117, and form tubular structures with Matrigel in vitro. Dynamic tracking shows that the expansion of individual stem cells completes asymmetric division with the involvement of trophoblast cells. Conclusion Trophoblast cells can provide a large number of advanced EPCs by providing trophoblast cells. Trophoblast cells may be involved in the asymmetric division and expansion of early stage endothelial stem cells.