Laboratory of Stem Cell Engineering, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan.
Stem Cells Dev. 2012 May 20;21(8):1287-98. doi: 10.1089/scd.2011.0283. Epub 2011 Oct 18.
Conventional human induced pluripotent stem cells (hiPSCs), reprogrammed from somatic cells by induced expression of Oct4, Sox2, Klf4, and c-Myc, are phenotypically different from mouse embryonic stem cells (ESCs). In mice, culture in N2B27 serum-free 2i media (mitogen-activated protein kinase/extracellular signal-regulated kinase and glycogen synthase kinase 3 inhibitors; PD0325901 and CHIR99021) plus leukemia inhibitory factor (LIF) (2i+LIF medium) enriches for germline competent ESCs. Here, we demonstrate that flat-shaped hiPSC colonies can be reprogrammed into bowl-shaped multi-potent stem cells (2i-hiPSCs) by using 2i+LIF medium. Mechanical dissociation of 2i-hiPSC colonies enables stable maintenance for >20 passages. Importantly, gene expression profiling demonstrated that 2i-hiPSCs more closely resemble primitive neural stem cells (PNSCs). Notably, this 2i-induced phenotype was generated from conventional hiPSCs, but not human ESCs (hESCs), thus correlating with the observation of neuroectodermal SOX1-positive colonies in conventional hiPSCs, but not hESCs in 2i+LIF medium. Thus, 2i-hiPSCs, which are nonteratoma forming PNSCs, may represent a safe source of cells for neural research and regenerative medicine.
传统的人诱导多能干细胞（hiPSCs），通过诱导表达 Oct4、Sox2、Klf4 和 c-Myc 从体细胞重新编程，表型上与小鼠胚胎干细胞（ESCs）不同。在小鼠中，在无血清 N2B27 2i 培养基（丝裂原活化蛋白激酶/细胞外信号调节激酶和糖原合酶激酶 3 抑制剂；PD0325901 和 CHIR99021）中培养，并添加白血病抑制因子（LIF）（2i+LIF 培养基）可富集生殖系相容的 ESCs。在这里，我们证明扁平形状的 hiPSC 集落可以通过使用 2i+LIF 培养基重编程为碗状多能干细胞（2i-hiPSCs）。2i-hiPSC 集落的机械解离能够稳定维持超过 20 代。重要的是，基因表达谱分析表明 2i-hiPSCs 更类似于原始神经干细胞（PNSCs）。值得注意的是，这种 2i 诱导的表型是由传统的 hiPSCs 产生的，而不是人类胚胎干细胞（hESCs），因此与传统 hiPSCs 中神经外胚层 SOX1 阳性集落的观察结果一致，但在 2i+LIF 培养基中 hESCs 中没有观察到。因此，2i-hiPSCs 是不形成畸胎瘤的 PNSCs，可能代表神经研究和再生医学的安全细胞来源。
