Alexanian Arshak R, Crowe Maria J, Kurpad Shekar N
Neuroscience Research Labs, Department of Neurosurgery, Medical College of Wisconsin, VAMC, 5000 W. National Avenue 151, Milwaukee, WI 53295, USA.
J Neurosci Methods. 2006 Jan 15;150(1):41-6. doi: 10.1016/j.jneumeth.2005.05.024. Epub 2005 Aug 8.
Several recent studies have shown that highly undifferentiated neural stem cells (NSCs) grafted into the intact or injured adult spinal cord of animal either remain undifferentiated or show fate restriction to a astrocytic lineage. This indicates that functionally diverse roles expected of cellular replacement cannot be performed by the transplantation of highly immature precursors; rather, more differentiated or appropriate mixtures of more restricted neural precursors may be important in replacement strategies. In this study, we investigated the ability of lineage-restricted neural progenitors derived from adult mouse periventricular subependymal zone (SEZ) to integrate and differentiate into the chronically injured adult spinal cord. To this end, NSCs were grown as adherent cultures followed by expansion in non-adhesive dishes. This allowed us to grow NSCs as colonies of restricted neural precursors, illustrated by NCAM, nestin, Sox-2, A2B5, and GFAP immunostaining. The mixture of lineage-restricted precursors was directly implanted into the chronically injured spinal cord of immunosuppressed cats. The fate of the cells was traced with GFP fluorescence and immunocytochemistry for neural markers such as beta-III-tubulin, GFAP, and Ng2. After four weeks, transplanted cells survived, giving rise to neurons and in addition to cells with an astrocytic phenotype. We conclude that a mixture of more restricted neural precursors may be better suited than highly immature NSCs for neural replacement strategies after central nervous system (CNS) injuries.
最近的几项研究表明，移植到完整或受损成年动物脊髓中的高度未分化神经干细胞（NSCs）要么保持未分化状态，要么表现出向星形胶质细胞谱系的命运限制。这表明，高度不成熟的前体细胞移植无法实现细胞替代所期望的多种功能；相反，更分化或更具限制性的神经前体细胞的适当混合物在替代策略中可能很重要。在本研究中，我们研究了源自成年小鼠脑室周围室管膜下区（SEZ）的谱系限制性神经祖细胞整合并分化到慢性损伤的成年脊髓中的能力。为此，将神经干细胞培养成贴壁培养物，然后在非贴壁培养皿中扩增。这使我们能够将神经干细胞培养成受限制的神经前体细胞集落，通过NCAM、巢蛋白、Sox-2、A2B5和GFAP免疫染色得以证明。将谱系限制性前体细胞的混合物直接植入免疫抑制猫的慢性损伤脊髓中。用绿色荧光蛋白（GFP）荧光和针对神经标志物如β-III-微管蛋白、GFAP和Ng2的免疫细胞化学方法追踪细胞的命运。四周后，移植细胞存活下来，产生了神经元以及具有星形胶质细胞表型的细胞。我们得出结论，对于中枢神经系统（CNS）损伤后的神经替代策略，更具限制性的神经前体细胞混合物可能比高度不成熟的神经干细胞更适合。
