Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
Proc Natl Acad Sci U S A. 2012 Feb 14;109(7):2527-32. doi: 10.1073/pnas.1121003109. Epub 2012 Jan 30.
We recently showed that defined sets of transcription factors are sufficient to convert mouse and human fibroblasts directly into cells resembling functional neurons, referred to as "induced neuronal" (iN) cells. For some applications however, it would be desirable to convert fibroblasts into proliferative neural precursor cells (NPCs) instead of neurons. We hypothesized that NPC-like cells may be induced using the same principal approach used for generating iN cells. Toward this goal, we infected mouse embryonic fibroblasts derived from Sox2-EGFP mice with a set of 11 transcription factors highly expressed in NPCs. Twenty-four days after transgene induction, Sox2-EGFP(+) colonies emerged that expressed NPC-specific genes and differentiated into neuronal and astrocytic cells. Using stepwise elimination, we found that Sox2 and FoxG1 are capable of generating clonal self-renewing, bipotent induced NPCs that gave rise to astrocytes and functional neurons. When we added the Pou and Homeobox domain-containing transcription factor Brn2 to Sox2 and FoxG1, we were able to induce tripotent NPCs that could be differentiated not only into neurons and astrocytes but also into oligodendrocytes. The transcription factors FoxG1 and Brn2 alone also were capable of inducing NPC-like cells; however, these cells generated less mature neurons, although they did produce astrocytes and even oligodendrocytes capable of integration into dysmyelinated Shiverer brain. Our data demonstrate that direct lineage reprogramming using target cell-type-specific transcription factors can be used to induce NPC-like cells that potentially could be used for autologous cell transplantation-based therapies in the brain or spinal cord.
我们最近表明，定义的转录因子集足以将小鼠和人类成纤维细胞直接转化为类似功能神经元的细胞，称为“诱导神经元”（iN）细胞。然而，对于某些应用，将成纤维细胞转化为增殖性神经前体细胞（NPC）而不是神经元可能是理想的。我们假设可以使用与生成 iN 细胞相同的主要方法来诱导 NPC 样细胞。为此，我们用一组在 NPC 中高表达的 11 种转录因子感染来自 Sox2-EGFP 小鼠的胚胎成纤维细胞。转染后 24 天，出现 Sox2-EGFP(+)集落，表达 NPC 特异性基因，并分化为神经元和星形胶质细胞。通过逐步消除，我们发现 Sox2 和 FoxG1 能够产生克隆自我更新、双潜能诱导 NPC，分化为星形胶质细胞和功能性神经元。当我们将 Pou 和 Homeobox 结构域转录因子 Brn2 添加到 Sox2 和 FoxG1 中时，我们能够诱导三潜能 NPC，不仅可以分化为神经元和星形胶质细胞，还可以分化为少突胶质细胞。转录因子 FoxG1 和 Brn2 本身也能够诱导 NPC 样细胞；然而，这些细胞产生的成熟神经元较少，尽管它们确实产生了星形胶质细胞，甚至能够整合到发育不良的 Shiverer 大脑中的少突胶质细胞。我们的数据表明，使用靶细胞类型特异性转录因子的直接谱系重编程可用于诱导 NPC 样细胞，这些细胞可能用于大脑或脊髓的基于自体细胞移植的治疗。
