Kim Joo Yeon, Lee Ju-Hyun, Sun Woong
Department of Anatomy, Korea University College of Medicine.
Department of Anatomy, Korea University College of Medicine;
J Vis Exp. 2016 Dec 15(118):54929. doi: 10.3791/54929.
Adult neural stem cells (aNSCs) can be used for the regeneration of damaged brain tissue. NSCs have the potential for differentiation and proliferation into three types of cells: neurons, astrocytes, and oligodendrocytes. Identifying aNSC-derived regions and characterizing the aNSC properties are critical for the potential use of aNSCs and for the elucidation of their role in neural regeneration. The subcallosal zone (SCZ), located between white matter and the hippocampus, has recently been reported to contain aNSCs and continuously give rise to neuroblasts. A low percentage of aNSCs from the SCZ is differentiated into neurons; most cells are differentiated into glial cells, such as oligodendrocytes and astrocytes. These cells are suggested to have a therapeutic potential for traumatic cortical injury. This protocol describes in detail the process to generate SCZ-aNSCs from an adult mouse brain. A brain matrix with intervals of 1 mm is used to obtain the SCZ-containing coronal slices and to precisely dissect the SCZ from the whole brain. The SCZ sections are initially subjected to a neurosphere culture. A well-developed culture system allows for the verification of their characteristics and can increase research on NSCs. A neurosphere culture system provides a useful tool for determining proliferation and collecting the genuine NSCs. A monolayer culture is also an in vitro system to assay proliferation and differentiation. Significantly, this culture system provides a more homogenous environment for NSCs than the neurosphere culture system. Thus, using a discrete brain region, these culture systems will be helpful for expanding our knowledge about aNSCs and their applications for therapeutic uses.
成体神经干细胞（aNSCs）可用于受损脑组织的再生。神经干细胞具有分化和增殖为三种细胞类型的潜力：神经元、星形胶质细胞和少突胶质细胞。识别aNSC来源区域并表征aNSC特性对于aNSCs的潜在应用以及阐明它们在神经再生中的作用至关重要。胼胝体下区（SCZ）位于白质和海马体之间，最近有报道称其含有aNSCs并持续产生神经母细胞。来自SCZ的aNSCs中只有一小部分分化为神经元；大多数细胞分化为胶质细胞，如少突胶质细胞和星形胶质细胞。这些细胞被认为对创伤性皮质损伤具有治疗潜力。本方案详细描述了从成年小鼠大脑中生成SCZ-aNSCs的过程。使用间隔为1毫米的脑基质来获取包含SCZ的冠状切片，并从整个大脑中精确解剖出SCZ。SCZ切片最初进行神经球培养。一个完善的培养系统有助于验证它们的特性，并可增加对神经干细胞的研究。神经球培养系统为确定增殖和收集真正的神经干细胞提供了一个有用的工具。单层培养也是一种用于检测增殖和分化的体外系统。重要的是，与神经球培养系统相比，这种培养系统为神经干细胞提供了更均匀的环境。因此，使用离散的脑区，这些培养系统将有助于扩展我们对aNSCs及其治疗应用的认识。
