A procedure for culturing astrocytes from white matter and the application of the siRNA technique for silencing the expression of their specific marker, S100A4
A procedure for culturing astrocytes from white matter and the application of the siRNA technique for silencing the expression of their specific marker, S100A4
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DOI：
10.1016/j.brainresprot.2005.03.005
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发表时间：
2005-07-01
期刊：
影响因子：
--
通讯作者：
Takenaga, K
中科院分区：
文献类型：
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作者：
Kozlova, EN;Takenaga, K
白色物质星形胶质细胞具有与灰质中的星形胶质细胞不同的生理功能。白色物质在损伤后变得高度不允许神经突生长，但白色物质星形胶质细胞在这一过程中的作用还不完全清楚。目前的协议，使原代星形胶质细胞的文化是不够的，探索特定的特性，白色物质星形胶质细胞在体外。我们描述了一个程序，用于获得文化的白色物质星形胶质细胞从啮齿动物胼胝体。在这个过程中，我们利用我们以前的发现，白色，而不是灰质星形胶质细胞表达的钙结合蛋白S100 A4。S100 A4表达星形胶质细胞在胼胝体中是丰富的，我们表明，文化，高度富集S100 A4表达白色物质星形胶质细胞，可以再现地产生从这个地区。成功培养的关键因素是（i）仔细解剖4天大大鼠的胼胝体，以及（ii）Percoll密度梯度离心以纯化星形胶质细胞。作为探索S100 A4在白色星形胶质细胞中的可能作用的一种手段，我们描述了使用siRNA技术来消除S100 A4在我们的体外系统中的表达。(C)2005 Elsevier B.V保留所有权利。
White matter astrocytes have physiological functions which are distinct from those of astrocytes in gray matter. White matter becomes highly non-permissive to neurite growth after injury, but the role of white matter astrocytes in this process is incompletely understood. Current protocols for making primary astroglial cultures are inadequate for exploring the specific properties of white matter astrocytes in vitro. We describe a procedure for obtaining cultures of white matter astrocytes from the rodent corpus callosum. In this procedure, we take advantage of our previous finding that white, but not gray matter astrocytes express the calcium-binding protein S100A4. S100A4 expressing astrocytes are abundant in the corpus callosum, and we show that cultures, highly enriched in S100A4 expressing white matter astrocytes, can be reproducibly generated from this area. Key factors for successful cultures are (i) meticulous dissection of the corpus callosum from 4-day-old rats, and (ii) Percoll density gradient centrifugation to purify astrocytes. As a means of exploring the possible role of S100A4 in white matter astrocytes, we describe the use of the siRNA technique to eliminate the expression of S100A4 in our in vitro system. (C) 2005 Elsevier B.V All rights reserved.