Large-scale generation of highly enriched neural stem-cell-derived oligodendroglial cultures: maturation-dependent differences in insulin-like growth factor-mediated signal transduction
Large-scale generation of highly enriched neural stem-cell-derived oligodendroglial cultures: maturation-dependent differences in insulin-like growth factor-mediated signal transduction
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DOI：
10.1111/j.1471-4159.2006.04171.x
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发表时间：
2007-02-01
影响因子：
4.7
通讯作者：
Back, Stephen A.
中科院分区：
文献类型：
--
作者：
Broughton, Sarah K.;Chen, Hanqin;Back, Stephen A.
多能神经干细胞（NSCs）在体内外均可定向分化为少突胶质细胞（OL）。我们利用这一特性开发了一种基于大鼠神经球（NS）/寡球（OS）的培养系统，以产生大量高度富集的晚期OL祖细胞（preOL）和成熟OL（MatOL）。CNS神经母细胞瘤细胞系B104衍生的条件培养基促进从解离的OS产生几乎纯的preOL群体。随后用睫状神经营养因子（CNTF）和3，3 '，5'-三碘-L-甲状腺原氨酸（T-3）培养preOL产生几乎纯的MatOL群体。通过免疫细胞化学、定量RT-PCR和基因表达谱分析证实了OL谱系特异性，这表明前OL和MatOL之间存在很大差异。胰岛素样生长因子（IGFs）是OL生存所需的有效神经保护剂。我们用这个系统来系统地定义在OL分化过程中IGF信号的成熟依赖性变化。与OS和MatOLs相比，NS和preOLs中IGF-I和胰岛素受体、胰岛素受体底物-1（IRS-1）和IRS-2、蛋白激酶B（PKB）/Akt和Janus激酶（JNK）的表达水平较高。Erk表达从NS到OS显著增加，在承诺到前OL时仅部分下降，并且在MatOL中，返回到与NS相似的低水平。在NSC分化过程中，通常增殖性Erk途径的IGF激活逐渐获得，而通常促存活、抗凋亡PI 3 K/PKB途径的IGF激活在每个发育阶段都是稳定的。
Multipotent neural stem cells (NSCs) are competent for commitment to the oligodendrocyte (OL) lineage both in vitro and in vivo. We exploited this property to develop a rat neurospheres (NS)/oligospheres (OS)-based culture system to generate large numbers of highly enriched late OL progenitors (preOLs) and mature OLs (MatOLs). CNS neuroblastoma cell line B104-derived conditioned medium promoted the generation of nearly pure populations of preOLs from dissociated OS. The subsequent culture of preOLs with ciliary neurotrophic factor (CNTF) and 3,3',5'-triiodo-L-thyronine (T-3) generated nearly pure populations of MatOLs. OL lineage specificity was confirmed by immunocytochemistry, quantitative RT-PCR and gene expression profiling, which demonstrated large differences between preOLs and MatOLs. The insulin-like growth factors (IGFs) are potent neuro-protective agents required for OL survival. We used this system to systematically define maturation-dependent changes in IGF signaling during the course of OL differentiation. The IGF-I and insulin receptors, insulin receptor substrate-1 (IRS-1) and IRS-2, protein kinase B (PKB)/Akt and Janus kinase (JNK) were expressed at higher levels in NS and preOLs compared with OS and MatOLs. Erk expression increased markedly from NS to OS, decreased only partially upon commitment to preOLs, and, in MatOLs, returned to a low level similar to NS. IGF activation of the generally proliferative Erk pathway was gradually acquired during NSC differentiation, whereas IGF activation of the generally pro-survival, anti-apoptotic PI3K/PKB pathway was consistently robust at each developmental stage.