Simultaneous prospective purification of adult subventricular zone neural stem cells and their progeny
Simultaneous prospective purification of adult subventricular zone neural stem cells and their progeny
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DOI：
10.1073/pnas.0810407106
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发表时间：
2009-04-14
影响因子：
11.1
通讯作者：
Doetsch, Fiona
中科院分区：
文献类型：
--
作者：
Pastrana, Erika;Cheng, Li-Chun;Doetsch, Fiona
前瞻性分离成体神经干细胞及其后代的能力对于研究其生物学和治疗潜力至关重要。成年哺乳动物神经源性龛中的干细胞是星形胶质细胞的一个子集。该领域的一个主要限制是无法区分干细胞星形胶质细胞和小生境星形胶质细胞。在这里，我们表明，表皮生长因子受体（EGFR）阳性室下区（SVZ）星形胶质细胞是激活的干细胞，被消除的抗有丝分裂治疗。我们开发了一种简单的策略，通过使用流式细胞术同时纯化成人SVZ干细胞谱系的不同阶段的细胞。该方法在GFAP：：GFP转基因小鼠中结合使用荧光EGF配体、CD 24和GFP表达，并允许同时纯化活化的干细胞星形胶质细胞（GFP+EGFR+ CD 24（-））、小生境星形胶质细胞（GFP+EGFR（-）CD 24（-））、转运扩增细胞（GFP-EGFR（+）CD 24（-））和成神经细胞（GFP（-）EGFR（-）CD 24（低））。三分之一的EGFR（+）星形胶质细胞在体外产生神经球，比未分选的细胞富集20倍。重要的是，这些细胞构成了SVZ星形胶质细胞中的神经球形成群体。这种方法将是非常实用的SVZ干细胞谱系的未来功能和分子研究。
The ability to prospectively isolate adult neural stem cells and their progeny is crucial to study their biology and therapeutic potential. Stem cells in adult mammalian neurogenic niches are a subset of astrocytes. A major limitation in the field has been the inability to distinguish stem cell astrocytes from niche astrocytes. Here, we show that epidermal growth factor receptor (EGFR)-positive subventricular-zone (SVZ) astrocytes are activated stem cells that are eliminated by antimitotic treatment. We developed a simple strategy to simultaneously purify cells at different stages of the adult SVZ stem cell lineage by using FACS. This method combines the use of fluorescent EGF ligand, CD24, and GFP expression in GFAP:: GFP transgenic mice and allows the simultaneous purification of activated stem cell astrocytes (GFP+EGFR+CD24(-)), niche astrocytes (GFP+EGFR(-)CD24(-)), transit amplifying cells (GFP-EGFR(+)CD24(-)), and neuroblasts (GFP(-)EGFR(-)CD24(low)). One in three EGFR(+) astrocytes gives rise to neurospheres in vitro, a 20-fold enrichment over unsorted cells. Importantly, these cells constitute the neurosphere-forming population among SVZ astrocytes. This approach will be of great utility for future functional and molecular studies of the SVZ stem cell lineage.