Wang Xiuli, Wei Guofeng, Yu Weiting, Zhao Yunshan, Yu Xingju, Ma Xiaojun
Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, P.R. China.
Biotechnol Prog. 2006 May-Jun;22(3):811-8. doi: 10.1021/bp060018z.
Embryonic stem (ES) cells are of significant interest either as an in vitro model recapitulating early embryonic development or as a renewable source of therapeutically useful cells. ES cells aggregation is important for embryoid bodies (EBs) formation and the subsequent generation of ES cell derivatives. This study was conducted to describe scalable production of EBs by the rotary cell culture system (RCCS, STLV type) and estimate the feasibility of constructing engineered cardiac tissue (ECT). In comparison with suspension culture in a Petri dish, the efficiency of the dynamic process was analyzed with respect to the yield of EB formation and their cardiomyocyte differentiation. Cardiomyocyte differentiation was evaluated by immunohistochemical analysis. After the elementary enrichment by gradient percoll, ES cell-derived cardiomyocytes were applied to construct ECT. Cell gross morphology, spatial distribution, and ultrastructure were evaluated by using histological analysis, confocal laser scanning microscopy, and transmission electron microscopy. Results showed that EB efficiencies in STLV were nearly 1.5-2.0 times higher than that of liquid suspension cultures, and cardiomyocyte differentiation of EBs progressed in a normal course after the dynamic cultivation in STLV. Additionally, the differentiated cultures could be enriched elementarily by gradient percoll. Once cast into the collagen strand, cells grew well and became more matured in Petri dishes. Synchronous contraction of the cell cluster was observed on the surface of the ECT, and cell connection was also established. It was the first report to have beating ES-derived cardiomyocytes on a 3-D collagen scaffold, which might provide a promising model for physiological and pharmacological studies and tissue replacement therapy.
胚胎干细胞（ES细胞）作为一种概括早期胚胎发育的体外模型或作为治疗有用细胞的可再生来源，备受关注。ES细胞聚集对于胚状体（EBs）形成以及随后ES细胞衍生物的产生很重要。本研究旨在描述通过旋转细胞培养系统（RCCS，STLV型）可扩展生产EBs，并评估构建工程化心脏组织（ECT）的可行性。与在培养皿中进行悬浮培养相比，针对EB形成的产量及其心肌细胞分化，分析了动态过程的效率。通过免疫组织化学分析评估心肌细胞分化。在通过梯度Percoll进行初步富集后，将ES细胞来源的心肌细胞应用于构建ECT。使用组织学分析、共聚焦激光扫描显微镜和透射电子显微镜评估细胞的总体形态、空间分布和超微结构。结果表明，STLV中的EB效率比液体悬浮培养高出近1.5 - 2.0倍，并且在STLV中进行动态培养后，EBs的心肌细胞分化正常进行。此外，分化的培养物可通过梯度Percoll进行初步富集。一旦铸入胶原纤维束中，细胞在培养皿中生长良好并变得更加成熟。在ECT表面观察到细胞簇的同步收缩，并且也建立了细胞连接。这是首次报道在三维胶原支架上有跳动的ES来源心肌细胞，这可能为生理和药理研究以及组织替代治疗提供一个有前景的模型。
