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神经干细胞向少突胶质前体细胞的定向分化诱导

富赛里¹, 胡建国¹, 李莹¹, 尹岚¹, 金建强¹, 徐晓明^1,2, 陆佩华^1,*

¹上海第二医科大学神经生物学实验室, 200025；²美国肯塔基脊髓损伤研究中心， 美国路易威尔大学神经外科系， 肯塔基 州 40292，美国

摘要

本研究采用神经胶质瘤细胞株(B104 neuroblatoma cells, B104 cells)培养上清(B104CM)和碱性成纤维细胞生长因子(basic fibroblast growth factor, bFGF), 将冷冻复苏的大鼠胚胎脊髓神经干细胞(neural stem cells, NSCs)定向诱导为少突胶质前体细胞 (oligodendrocyte precusor cells, OPCs)。形态学和免疫组化的结果显示, 诱导后 95% 以上的细胞具有双极或多极突起的典型 OPCs 形态,并表达 A2B5 和血小板源生长因子受体 -α (platelet derived growth factor receptor-α, PDGFR-α)等 OPCs 标志, 所有PDGFR-α 阳性的 OPCs 均不表达β-Tublin Ⅲ，其中仅少量细胞表达胶质原纤维酸性蛋白 (glia fibrillary acidic protein, GFAP)。在 B104CM 和 bFGF 共存的培养条件下，悬浮培养的 OPCs 可大量增殖形成少突胶质细胞球，该细胞球可通过传代继续扩增，且扩增的 OPCs 仍能维持其特有的形态和自我增殖的特性。撤去 bFGF 和 B104CM 后, OPCs 能进一步分化为成熟的少突胶质细胞 (oligodendrocytes, OLs) 或Ⅱ型星形胶质细胞。实验表明，诱导NSCs 产生的OPCs 在形态、增殖以及分化格局等方面均与已报道 的存在于胚胎脑区的O-2A 前体细胞相类似。该培养系统可为实验性细胞移植的研究提供丰富的细胞来源。

关键词： 神经干细胞; 少突胶质前体细胞; B104 细胞; 碱性成纤维细胞生长因子

Induction of rat neural stem cells into oligodendrocyte precursor cells

FU Sai-Li¹, HU Jian-Guo¹, LI Ying¹, YIN Lan¹, JIN Jian-Qiang¹, XU Xiao-Ming^1,2, LU Pei-Hua^1,*

¹Department of Neurobiology, Shanghai Second Medical University, Shanghai 200025, China；²Kentucky Spinal Cord Injury Research Center, Department of Neurological Surgery, University of Louisville, KY 40292, USA

Abstract

We have previously established a culture method to isolate and cultivate neural stem cells (NSCs) derived from the rat embryonic brain and spinal cord. In the present study, we demonstrate that the spinal cord-derived NSCs can be induced to differentiate into oligodendrocyte precursor cells (OPCs) with a combined treatment composed of (1) conditioned medium collected from B104 neuroblastoma cells (B104CM) and (2) basic fibroblast growth factor (bFGF, 10 ng/ml). After induction, over 95% of the cells displayed bipolar or tri-polar morphology and expressed A2B5 and platelet derived growth factor receptor-α (PDGFR-α), markers that are specific for OPCs. Among PDGFR-α positive OPCs, only a few cells expressed glia fibrillary acidic protein (GFAP) and none expressed β-tubulin Ⅲ. In the presence of B104CM and bFGF, OPCs proliferated rapidly, formed spheres, expanded for multiple passages, and maintained their phenotypic properties. Upon withdrawal of B104CM and bFGF, these cells differentiated into either O4/GlaC-positive oligodendrocytes (OLs) or GFAP- and A2B5-positive type-2 astrocytes. Our results indicate that NSCs can be induced to differentiate into OPCs that possess properties of self-renewal and differentiation into oligodendrocytes and type-2 astrocytes, a property similar to that of O-2A progenitor cells. The OPCs can be maintained in an undifferentiated state over multiple divisions as long as both B104CM and bFGF are present in the medium. Thus, large quantity of OPCs can be obtained through this method for potential therapeutical interventions for various neurological degenerative diseases.

Key words: neural stem cells; oligodendrocyte precusor cells; B104 neuroblastoma cells; basic fibroblast growth factor

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通讯作者：陆佩华　　E-mail: neuron@shsmu.edu.cn

引用本文：

富赛里, 胡建国, 李莹, 尹岚, 金建强, 徐晓明, 陆佩华. 神经干细胞向少突胶质前体细胞的定向分化诱导[J]. 生理学报 2005; 57 (2): 132-138.

FU Sai-Li, HU Jian-Guo, LI Ying, YIN Lan, JIN Jian-Qiang, XU Xiao-Ming, LU Pei-Hua. Induction of rat neural stem cells into oligodendrocyte precursor cells. Acta Physiol Sin 2005; 57 (2): 132-138