Stable Expression of Neurogenin 1 Induces LGR5, a Novel Stem Cell Marker, in an Immortalized Human Neural Stem Cell Line HB1.F3
Stable Expression of Neurogenin 1 Induces LGR5, a Novel Stem Cell Marker, in an Immortalized Human Neural Stem Cell Line HB1.F3
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DOI：
10.1007/s10571-009-9466-3
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发表时间：
2010-04
影响因子：
4
通讯作者：
J. Satoh;S. Obayashi;H. Tabunoki;Taeko Wakana;Seung U. Kim
中科院分区：
文献类型：
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作者：
J. Satoh;S. Obayashi;H. Tabunoki;Taeko Wakana;Seung U. Kim
神经干细胞（NSC）具有自我更新和多能性，是治疗脊髓损伤、中风和神经退行性疾病的理想细胞来源。为了有效地诱导来自NSC的神经元谱系细胞用于神经元替代治疗，我们应该明确参与人类NSC分化的时间和地点特异性调控的内在遗传程序。最近，我们建立了一个永生化的人NSC克隆HB1。F3提供无限的NSC来源，适用于细胞治疗的基因操作。为了研究神经原生性碱性螺旋-环-螺旋（bHLH）转录因子Ngn1 （neurogenin 1）在人NSC分化中的作用，我们建立了一个稳定过表达Ngn1的F3克隆。全基因组基因表达谱分析发现，在ngn1过表达的F3细胞（F3- ngn1）与野生型F3细胞（F3- wt）中，有250个基因上调，338个基因下调。值得注意的是，一种新的干细胞标记物——富含亮氨酸重复序列的G蛋白偶联受体5 （LGR5）显示F3-Ngn1的表达增加了167倍，尽管Ngn1的短暂过表达并未诱导LGR5的上调，这表明LGR5不是Ngn1的直接转录靶点。KeyMolnet是一个生物信息学工具，用于分析综合知识库上的分子关系，表明差异表达基因的分子网络涉及多种转录因子调节的网络的复杂相互作用。基因本体（Gene ontology， GO）的发育和形态发生方面由上调基因富集，而细胞外基质和粘附方面则由下调基因富集。这些结果表明，单个基因Ngn1在F3细胞中的稳定表达不仅可以诱导神经源性的变化，还可以诱导多种功能的变化，这些变化可能通过复杂基因调控网络的重组影响人类NSC的分化。
Neural stem cells (NSC) with self-renewal and multipotent properties serve as an ideal cell source for transplantation to treat spinal cord injury, stroke, and neurodegenerative diseases. To efficiently induce neuronal lineage cells from NSC for neuron replacement therapy, we should clarify the intrinsic genetic programs involved in a time- and place-specific regulation of human NSC differentiation. Recently, we established an immortalized human NSC clone HB1.F3 to provide an unlimited NSC source applicable to genetic manipulation for cell-based therapy. To investigate a role of neurogenin 1 (Ngn1), a proneural basic helix-loop-helix (bHLH) transcription factor, in human NSC differentiation, we established a clone derived from F3 stably overexpressing Ngn1. Genome-wide gene expression profiling identified 250 upregulated genes and 338 downregulated genes in Ngn1-overexpressing F3 cells (F3-Ngn1) versus wild-type F3 cells (F3-WT). Notably, leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a novel stem cell marker, showed an 167-fold increase in F3-Ngn1, although transient overexpression of Ngn1 did not induce upregulation of LGR5, suggesting that LGR5 is not a direct transcriptional target of Ngn1. KeyMolnet, a bioinformatics tool for analyzing molecular relations on a comprehensive knowledgebase, suggests that the molecular network of differentially expressed genes involves the complex interaction of networks regulated by multiple transcription factors. Gene ontology (GO) terms of development and morphogenesis are enriched in upregulated genes, while those of extracellular matrix and adhesion are enriched in downregulated genes. These results suggest that stable expression of a single gene Ngn1 in F3 cells induces not simply neurogenic but multifunctional changes that potentially affect the differentiation of human NSC via a reorganization of complex gene regulatory networks.