采用高通量方法筛选用于人类胚胎干细胞体外长期培养的温敏型水凝胶

Identification of Thermo-Responsive Hydrogels for Long-Term Culture of Human Embryonic Stem Cells by Using High-Throughput Approach

人类胚胎干细胞(hESC)的体外培养通常需要附着在蛋白质培养基质或饲养细胞层上,并主要依靠细胞的酶解离方法传代。采用动物来源的蛋白酶处理如此敏感的细胞会损伤膜蛋白、增加污染风险,这显然是不可取的,应尽量避免使用。在此,我们汇报了采用高通量方法筛选出一类以2-(二乙基氨基)乙基丙烯酸酯为主要单体的温敏型水凝胶。该类材料不仅具有在小幅度降温时改变性能以使其表面生长的干细胞脱粘附的功能,从而取代细胞传代中使用的生物酶,而且能取代蛋白质基质使hESC可以在体外进行长期培养(>20代),并保持完整的干细胞特征。这些具有可控化学结构的、可取代具动物来源生物基质的合成材料代表了一种新型的体系。它将是一种能使干细胞的培养达到可控、高效和安全的材料,会大大促进干细胞的深入研究和在临床医学中的应用。

Human embryonic stem cell （hESC） cultures typically rely on protein matrices or feeder cells to support attachment and growth with enzymatic cell dissociation methods for passaging. Clearly the stripping of membrane proteins from such sensitive cells by proteases is undesirable and best avoided. Here we report the identification of a family of thermoresponsive synthetic hydrogels based on 2-- （diethylamino） ethyl acrylates which provide enzyme--free cell detachment at passaging in a chemically defined medium,with robust preservation of undifferentiated hESC properties for multiple lines over extended culture （ 20 passages）. These chemically defined alternatives to biological substrates represent a novel approach to improving the definition, efficacy and safety of hESC culture systems for research and translational appli- cations,