Singec Ilyas, Knoth Rolf, Meyer Ralf P, Maciaczyk Jaroslaw, Volk Benedikt, Nikkhah Guido, Frotscher Michael, Snyder Evan Y
Institute of Anatomy and Cell Biology, University of Freiburg, Albertstr. 17, D-79104 Freiburg, Germany.
Nat Methods. 2006 Oct;3(10):801-6. doi: 10.1038/nmeth926.
For more than a decade the 'neurosphere assay' has been used to define and measure neural stem cell (NSC) behavior, with similar assays now used in other organ systems and in cancer. We asked whether neurospheres are clonal structures whose diameter, number and composition accurately reflect the proliferation, self-renewal and multipotency of a single founding NSC. Using time-lapse video microscopy, coculture experiments with genetically labeled cells, and analysis of the volume of spheres, we observed that neurospheres are highly motile structures prone to fuse even under ostensibly 'clonal' culture conditions. Chimeric neurospheres were prevalent independent of ages, species and neural structures. Thus, the intrinsic dynamic of neurospheres, as conventionally assayed, introduces confounders. More accurate conditions (for example, plating a single cell per miniwell) will be crucial for assessing clonality, number and fate of stem cells. These cautions probably have implications for the use of 'cytospheres' as an assay in other organ systems and with other cell types, both normal and neoplastic.
十多年来，“神经球分析”一直被用于定义和测量神经干细胞（NSC）的行为，现在其他器官系统和癌症研究中也使用了类似的分析方法。我们研究了神经球是否为克隆结构，其直径、数量和组成是否能准确反映单个起始神经干细胞的增殖、自我更新和多能性。通过延时视频显微镜、与基因标记细胞的共培养实验以及对球体体积的分析，我们观察到神经球是高度动态的结构，即使在表面上“克隆”的培养条件下也容易融合。嵌合神经球普遍存在，与年龄、物种和神经结构无关。因此，按照传统方法检测，神经球的内在动态会引入混杂因素。更精确的条件（例如，每个微孔接种单个细胞）对于评估干细胞的克隆性、数量和命运至关重要。这些注意事项可能对在其他器官系统以及其他正常和肿瘤细胞类型中使用“细胞球”作为分析方法有影响。
