Morriss-Kay G, Tuckett F
Department of Human Anatomy, Oxford, UK.
Development. 1989 Aug;106(4):787-98. doi: 10.1242/dev.106.4.787.
Studies on cell behaviour in vitro have indicated that the chondroitin sulphate proteoglycan (CSPG) family of molecules can participate in the control of cell proliferation, differentiation and adhesion, but its morphogenetic functions had not been investigated in intact embryos. Chondroitin/chondroitin sulphates have been identified in rat embryos at low levels at the start of neurulation (day 9) and at much higher levels on day 10. In this study we have sought evidence for the morphogenetic functions of CSPGs in rat embryos during the period of neurulation and neural crest cell migration by a combination of two approaches: immunocytochemical localization of CSPG by means of an antibody, CS-56, to the chondroitin sulphate component of CSPG, and exposure of embryos to the enzyme chondroitinase ABC. Staining of the CS-56 epitope was poor at the beginning of cranial neurulation; bright staining was at first confined to the primary mesenchyme under the convex neural folds late on day 9. In day 10 embryos, all mesenchyme cells were stained, but at different levels of intensity, so that primary mesenchyme, neural crest and sclerotomal cells could be distinguished from each other. Basement membranes were also stained, particularly bright staining being present where two epithelial were basally apposed, e.g., neural/surface ectoderms, dorsal aorta/neural tube, prior to migration of a population of cells between them. Staining within the neural epithelium was first confined to the dorsolateral edge region, and associated with the onset of neural crest cell emigration; after neural tube closure, neuroepithelial staining was more general. Neural crest cells were stained during migration, but the reaction was absent in areas associated with migration end-points (trigeminal ganglion anlagen, frontonasal mesenchyme). Embryos exposed to chondroitinase ABC in culture showed no abnormalities until early day 10, when cranial neural crest cell emigration from the neural epithelium was inhibited and neural tube closure was retarded. Sclerotomal cells failed to take their normal pathway between the dorsal aorta and neural tube. Correlation of the results of these two methods suggests: (1) that by decreasing adhesiveness within the neural epithelium at specific stages, CSPG facilitates the emigration of neural crest cells and the migratory movement of neuroblasts, and may also provide increased flexibility during the generation of epithelial curvatures; (2) that by decreasing the adhesiveness of fibronectin-containing extracellular matrices, CSPG facilitates the migration of neural crest and sclerotomal cells. This second function is particularly important when migrating cells take pathways between previously apposed tissues.
体外细胞行为研究表明，硫酸软骨素蛋白聚糖（CSPG）分子家族可参与细胞增殖、分化和黏附的调控，但其形态发生功能在完整胚胎中尚未得到研究。在大鼠胚胎中，神经胚形成开始时（第9天）硫酸软骨素/硫酸软骨素的含量较低，而在第10天含量则高得多。在本研究中，我们通过两种方法相结合，寻找CSPG在大鼠胚胎神经胚形成期和神经嵴细胞迁移过程中的形态发生功能的证据：一种方法是使用针对CSPG硫酸软骨素成分的抗体CS-56对CSPG进行免疫细胞化学定位；另一种方法是将胚胎暴露于软骨素酶ABC。在颅神经胚形成开始时，CS-56表位的染色较弱；明亮的染色最初仅限于第9天后期凸向的神经褶下方的初级间充质。在第10天的胚胎中，所有间充质细胞均被染色，但染色强度不同，因此可以区分初级间充质、神经嵴和硬骨细胞。基底膜也被染色，特别是在两个上皮细胞基底相对的部位，如神经/表面外胚层、背主动脉/神经管，在一群细胞在它们之间迁移之前染色特别明亮。神经上皮内的染色最初仅限于背外侧边缘区域，与神经嵴细胞迁出的开始相关；神经管闭合后，神经上皮染色更为普遍。神经嵴细胞在迁移过程中被染色，但在与迁移终点相关的区域（三叉神经节原基、额鼻间充质）没有反应。在培养中暴露于软骨素酶ABC的胚胎直到第10天早期都没有出现异常，此时颅神经嵴细胞从神经上皮的迁出受到抑制，神经管闭合延迟。硬骨细胞未能在背主动脉和神经管之间走正常路径。这两种方法结果的相关性表明：（1）通过在特定阶段降低神经上皮内的黏附性，CSPG促进神经嵴细胞的迁出和成神经细胞的迁移运动，并且在上皮弯曲形成过程中可能还提供了更大的灵活性；（2）通过降低含纤连蛋白的细胞外基质的黏附性，CSPG促进神经嵴和硬骨细胞的迁移。当迁移细胞在先前相邻组织之间通过时，这第二种功能尤为重要。
