Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
Cell Transplant. 2010;19(4):471-86. doi: 10.3727/096368909X484707. Epub 2009 Dec 18.
During the last decade, much progress has been made in developing protocols for the differentiation of human embryonic stem cells (hESCs) into a neural phenotype. The appropriate agent for cell therapy is neural precursors (NPs). Here, we demonstrate the derivation of highly enriched and expandable populations of proliferating NPs from the CCTL14 line of hESCs. These NPs could differentiate in vitro into functionally active neurons, as confirmed by immunohistochemical staining and electrophysiological analysis. Neural cells differentiated in vitro from hESCs exhibit broad cellular heterogeneity with respect to developmental stage and lineage specification. To analyze the population of the derived NPs, we used fluorescence-activated cell sorting (FACS) and characterized the expression of several pluripotent and neural markers, such as Nanog, SSEA-4, SSEA-1, TRA-1-60, CD24, CD133, CD56 (NCAM), beta-III-tubulin, NF70, nestin, CD271 (NGFR), CD29, CD73, and CD105 during long-term propagation. The analyzed cells were used for transplantation into the injured rodent brain; the tumorigenicity of the transplanted cells was apparently eliminated following long-term culture. These results complete the characterization of the CCTL14 line of hESCs and provide a framework for developing cell selection strategies for neural cell-based therapies.
在过去的十年中，在开发将人类胚胎干细胞（hESC）分化为神经表型的方案方面取得了很大进展。细胞治疗的合适制剂是神经前体细胞（NPs）。在这里，我们展示了从 hESC 的 CCTL14 系中衍生出高度富集和可扩增的增殖性 NPs 群体。这些 NPs 可以在体外分化为功能活跃的神经元，这通过免疫组织化学染色和电生理分析得到证实。体外从 hESC 分化的神经细胞在发育阶段和谱系特化方面表现出广泛的细胞异质性。为了分析衍生的 NPs 群体，我们使用了荧光激活细胞分选（FACS），并表征了几种多能性和神经标记物的表达，例如 Nanog、SSEA-4、SSEA-1、TRA-1-60、CD24、CD133、CD56（NCAM）、β-III-微管蛋白、NF70、巢蛋白、CD271（NGFR）、CD29、CD73 和 CD105，在长期传代过程中。分析的细胞被用于移植到受损的啮齿动物大脑中；在长期培养后，移植细胞的致瘤性显然被消除。这些结果完成了 hESC 的 CCTL14 系的表征，并为开发基于神经细胞的治疗的细胞选择策略提供了框架。
