Identification and characterization of early glial progenitors using a transgenic selection strategy
Identification and characterization of early glial progenitors using a transgenic selection strategy
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DOI：
10.1523/jneurosci.19-02-00759.1999
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发表时间：
1999-01-15
影响因子：
5.3
通讯作者：
Hudson, LD
中科院分区：
文献类型：
--
作者：
Chandross, KJ;Cohen, RI;Hudson, LD
为了定义少突胶质细胞 (OL) 和雪旺细胞 (SC) 的时空发育并同时选择，产生了在鼠 2'3'-环核苷酸控制下表达细菌 β-半乳糖苷酶 (β-gal) 和新霉素磷酸转移酶融合蛋白 (β geo) 的转基因小鼠 3'-磷酸二酯酶 (muCNP) 启动子 I 和 II。在胚胎第 12.5 天，在菱脑和脊髓的腹侧区域以及神经嵴中检测到转基因 β-gal 活性。当菱脑细胞在 G418 存在的情况下培养时，存活的细胞分化为 OL，这表明在发育过程中该大脑区域提供了 OL 祖细胞的来源之一。出生后，强大的 P-gal 活性局限于整个大脑的 OL，星形胶质细胞、神经元、小胶质细胞或单核细胞中不存在。在坐骨神经中，β-半乳糖活性仅局限于 SC。出生后第 10 天脑或坐骨神经的培养物在 G418 存在下生长，在接触抗生素的 8-9 天内，所有存活细胞中 99% 为 β-gal 阳性 OL 或 SC。这些研究表明，muCNP-β geo 转基因小鼠可用于识别神经胶质细胞谱系早期阶段及其整个发育过程中的 OL 和 SC。这种新方法明确地确定了体内鉴定的β-gal阳性细胞是神经胶质祖细胞，定义为它们能够在抗生素选择中存活并在体外分化为OL或SC。此外，该实验范式有助于快速有效地选择小鼠 OL 和 SC 的纯群体，并进一步强调了细胞特异性启动子在不同细胞类型纯化中的使用。
To define the spatiotemporal development of and simultaneously select for oligodendrocytes (OLs) and Schwann cells (SCs), transgenic mice were generated that expressed a bacterial beta-galactosidase (beta-gal) and neomycin phosphotransferase fusion protein (beta geo) under the control of murine 2'3'-cyclic nucleotide 3'-phosphodiesterase (muCNP) promoters I and II. Transgenic beta-gal activity was detected at embryonic day 12.5 in the ventral region of the rhombencephalon and spinal cord and in the neural crest. When cells from the rhombencephalon were cultured in the presence of G418, surviving cells differentiated into OLs, indicating that during development this brain region provides one source of OL progenitors. Postnatally, robust P-gal activity was localized to OLs throughout the brain and was absent from astrocytes, neurons, and microglia or monocytes. In the sciatic nerve beta-gal activity was localized exclusively to SCs. Cultures from postnatal day 10 brain or sciatic nerve were grown in the presence of G418, and within 8-9 d exposure to antibiotic, 99% of all surviving cells were beta-gal-positive OLs or SCs. These studies demonstrate that the muCNP-beta geo transgenic mice are useful for identifying OLs and SCs beginning at early stages of the glial cell lineage and throughout their development. This novel approach definitively establishes that the beta-gal-positive cells identified in vivo are glial progenitors, as defined by their ability to survive antibiotic selection and differentiate into OLs or SCs in vitro. Moreover, this experimental paradigm facilitates the rapid and efficient selection of pure populations of mouse OLs and SCs and further underscores the use of cell-specific promoters in the purification of distinct cell types.