Watanabe Kiichi, Ueno Morio, Kamiya Daisuke, Nishiyama Ayaka, Matsumura Michiru, Wataya Takafumi, Takahashi Jun B, Nishikawa Satomi, Nishikawa Shin-ichi, Muguruma Keiko, Sasai Yoshiki
Organogenesis and Neurogenesis Group, Center for Developmental Biology, RIKEN, Kobe 650-0047, Japan.
Nat Biotechnol. 2007 Jun;25(6):681-6. doi: 10.1038/nbt1310. Epub 2007 May 27.
Poor survival of human embryonic stem (hES) cells after cell dissociation is an obstacle to research, hindering manipulations such as subcloning. Here we show that application of a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, to hES cells markedly diminishes dissociation-induced apoptosis, increases cloning efficiency (from approximately 1% to approximately 27%) and facilitates subcloning after gene transfer. Furthermore, dissociated hES cells treated with Y-27632 are protected from apoptosis even in serum-free suspension (SFEB) culture and form floating aggregates. We demonstrate that the protective ability of Y-27632 enables SFEB-cultured hES cells to survive and differentiate into Bf1(+) cortical and basal telencephalic progenitors, as do SFEB-cultured mouse ES cells.
人胚胎干细胞（hES细胞）在细胞解离后存活率低是研究的一个障碍，阻碍了诸如亚克隆等操作。在此我们表明，将选择性Rho相关激酶（ROCK）抑制剂Y-27632应用于hES细胞可显著减少解离诱导的细胞凋亡，提高克隆效率（从约1%提高到约27%），并促进基因转移后的亚克隆。此外，用Y-27632处理的解离hES细胞即使在无血清悬浮（SFEB）培养中也能免受细胞凋亡影响，并形成漂浮聚集体。我们证明，Y-27632的保护能力使SFEB培养的hES细胞能够存活并分化为Bf1(+)皮质和基底前脑祖细胞，SFEB培养的小鼠ES细胞也是如此。
