Xiong Wen, Gao Yan, Cheng Xun, Martin Charles, Wu Dongmei, Yao Shuyuan, Kim Min-Ju, Liu Yang
Research and Development, Stemgent.
J Vis Exp. 2009 Nov 18(33):1550. doi: 10.3791/1550.
Mouse embryonic stem (ES) cells are conventionally cultured with Leukemia Inhibitory Factor (LIF) to maintain self-renewal.(1) However, LIF is expensive and activation of the LIF/JAK/STAT3 pathway is not absolutely required to maintain the self-renewal state.(2) The SC1 small molecule may be an economical alternative to LIF. SC1 functions through dual inhibition of Ras-GAP and ERK1.(3) Illustration of its mechanism of action makes it a useful tool to study the fundamental molecular mechanism of self-renewal. Here we demonstrate the procedure for culturing mouse ES cells in the presence of SC1 and show that they are able to maintain self-renewal in the absence of LIF. Cells cultured with SC1 showed similar morphology compared to cells maintained with LIF. Both exhibited typical mouse ES morphology after five passages. Expression of typical pluripotency markers (Oct4, Sox2, Nanog, and SSEA1) was observed after five passages in the presence of SC1. Furthermore, SC1 caused no overt toxicity on mouse ES cells.
传统上，小鼠胚胎干细胞（ES细胞）通过白血病抑制因子（LIF）进行培养以维持自我更新。（1）然而，LIF价格昂贵，并且维持自我更新状态并非绝对需要激活LIF/JAK/STAT3信号通路。（2）SC1小分子可能是LIF的一种经济替代物。SC1通过双重抑制Ras-GAP和ERK1发挥作用。（3）对其作用机制的阐释使其成为研究自我更新基本分子机制的有用工具。在此，我们展示了在SC1存在的情况下培养小鼠ES细胞的方法，并表明它们能够在没有LIF的情况下维持自我更新。与用LIF培养的细胞相比，用SC1培养的细胞表现出相似的形态。传代五次后，两者均呈现典型的小鼠ES细胞形态。在存在SC1的情况下传代五次后，观察到典型的多能性标志物（Oct4、Sox2、Nanog和SSEA1）的表达。此外，SC1对小鼠ES细胞没有明显毒性。
