Moase C E, Trasler D G
Department of Biology, McGill University, Montreal, Quebec, Canada.
Teratology. 1990 Aug;42(2):171-82. doi: 10.1002/tera.1420420208.
Splotch (Sp) and splotch-delayed (Spd) are allelic mutations on chromosome 1 of the mouse. Embryos homozygous for either allele have neural tube defects (NTDs) and deficiencies in neural crest cell (NCC) derived structures. The fact that Spd mouse mutants sometimes have deficiencies in NCC derivatives in the absence of an NTD led to the hypothesis that neurulation and the release of NCCs may depend on a regulatory event that is common to both processes. Therefore, it may be possible to understand the cause of NTDs in these mutants by examining the basis of aberrant NCC derivatives. Caudal neural tubes were excised from day 9 Sp and Spd embryos and placed into gelatin-coated tissue culture dishes, or 3-dimensional basement membrane matrigel, and cultured for 72 hours. A cytogenetic marker was used to genotype the embryos. In planar cultures, no morphological differences were observed between NCCs from neural tube explants of Spd mutants compared to those from heterozygous or wild-type embryos. However, there appeared to be a delay in the release of NCCs from the neural tube in both Sp and Spd mutants, which was particularly evident in Sp. After 24 hours in culture, the extent of NCC outgrowth, as well as the number of NCCs emigrating from explanted neural tubes, was significantly lower in Sp and Spd mutant cultures than in controls. No differences were observed in the mitotic indices among cells which had emigrated. By 72 hours, mutant cultures and their non-mutant counterparts were similar in terms of outgrowth, cell number, and migratory capability. After 24 hours in 3-dimensional basement membrane matrigel, cell outgrowth from Sp explants was also significantly less than controls. The pattern of NCC outgrowth in both types of culture conditions indicates a 24 hour delay in mutant cultures compared to controls. This stems from a delay in the release of NCCs from the neural tube, suggesting that the defect lies within the neuroepithelium with respect to the release of NCCs.
斑点（Sp）和延迟斑点（Spd）是小鼠1号染色体上的等位基因突变。任一突变等位基因的纯合胚胎都有神经管缺陷（NTDs）以及神经嵴细胞（NCC）衍生结构的缺陷。Spd小鼠突变体有时在没有神经管缺陷的情况下存在NCC衍生物缺陷，这一事实导致了这样一种假说，即神经胚形成和NCC的释放可能依赖于这两个过程共有的一个调控事件。因此，通过研究异常NCC衍生物的基础，有可能了解这些突变体中神经管缺陷的原因。从第9天的Sp和Spd胚胎中切除尾侧神经管，放入涂有明胶的组织培养皿或三维基底膜基质胶中，培养72小时。使用细胞遗传学标记对胚胎进行基因分型。在平面培养中，与杂合子或野生型胚胎的神经管外植体相比，Spd突变体的神经管外植体的NCCs在形态上没有差异。然而，在Sp和Spd突变体中，NCCs从神经管的释放似乎都有延迟，这在Sp中尤为明显。培养24小时后，Sp和Spd突变体培养物中NCC的生长程度以及从外植神经管迁出的NCC数量均显著低于对照组。迁出细胞的有丝分裂指数没有观察到差异。到72小时时，突变体培养物及其非突变体对应物在生长、细胞数量和迁移能力方面相似。在三维基底膜基质胶中培养24小时后，Sp外植体的细胞生长也明显少于对照组。两种培养条件下NCC的生长模式表明，与对照组相比，突变体培养物有24小时的延迟。这是由于NCCs从神经管释放的延迟，表明在NCCs释放方面，缺陷存在于神经上皮内。
