Kubota Y, Ito K
Department of Biology, Graduate School of Science, Osaka University, Japan.
Dev Dyn. 2000 Feb;217(2):170-9. doi: 10.1002/(SICI)1097-0177(200002)217:2<170::AID-DVDY4>3.0.CO;2-9.
We examined the roles of fibroblast growth factor (FGF)-2 and FGF-8 in the migration of mesencephalic mouse neural crest cells. Our in vitro migration assay has shown that FGF-2 (basic FGF) and FGF-8 have chemotactic activity for these cells. Chemotaxis was inhibited by anti-FGF-2 and anti-FGF-8 neutralizing antibodies. In addition, anti-FGF-2 blocked neural crest cell migration in cranial organ cultures. This observation suggests that FGF-2 functions as a chemoattractant in migration of mesencephalic neural crest cells in vivo. In organ culture, the antagonist of FGF binding to a low-affinity fibroblast growth factor receptor (FGFR) heparan sulfate, inositolhexakisphosphate (InsP6), inhibited migration as well. Mesencephalic neural crest cells had high-affinity FGFRs, in particular FGFR-1 and FGFR-3. Thus, the chemotactic activities of FGF-2 can be mediated by the low-affinity FGFR alone or by a combination of low- and high-affinity FGFRs (FGFR-1, FGFR-3, or both). Moreover, differential localization of FGF-2 was found at the mesencephalic axial level of intact embryos during neural crest cell migration. FGF-2 protein expression was predominant in the target regions, in particular the mandibular mesenchyme, that are colonized by mesencephalic neural crest cells. This characteristic distribution supports the notion that FGF-2 acts as a chemoattractant in the mouse embryo that directs mesencephalic neural crest cell migration. Whereas FGF-8 showed chemotactic activity in vitro, neural crest cell dispersion was observed in explants that had been treated with anti-FGF-8 neutralizing antibodies. This result suggests that FGF-8 may not be a chemoattractant in vivo. However, the distribution of neural crest cells in explants treated with anti-FGF-8 differed from that in control explants or in intact embryos. Extreme FGF-2 distribution was observed in the mandibular arch and FGF-8 is expressed in the epithelium. FGF-8 may play a role in mesencephalic neural crest cell migration, and its role may be concerned with the differential localization of FGF-2. To establish this notion, we performed immunohistochemical examination of FGF-2 distribution in explants treated with FGF-8 and analysis of FGF-2 gene expression levels by reverse transcriptase-polymerase chain reaction by using RNA from explants. The data indicate that FGF-2 is distributed throughout the mesenchyme in FGF-8-treated explants and that expression of FGF-2 is promoted by FGF-8. Therefore, we conclude that the expression of FGF-8 in the mandibular arch epithelium is a prerequisite for the differential localization of FGF-2 and that the FGF-2 distribution pattern is essential for chemotaxis of mesencephalic neural crest cell migration.
我们研究了成纤维细胞生长因子（FGF）-2和FGF-8在小鼠中脑神经嵴细胞迁移中的作用。我们的体外迁移试验表明，FGF-2（碱性FGF）和FGF-8对这些细胞具有趋化活性。抗FGF-2和抗FGF-8中和抗体可抑制趋化作用。此外，抗FGF-2可阻断颅器官培养物中神经嵴细胞的迁移。这一观察结果表明，FGF-2在体内中脑神经嵴细胞迁移中作为一种化学引诱剂发挥作用。在器官培养中，FGF与低亲和力成纤维细胞生长因子受体（FGFR）硫酸乙酰肝素结合的拮抗剂肌醇六磷酸（InsP6）也可抑制迁移。中脑神经嵴细胞具有高亲和力FGFR，特别是FGFR-1和FGFR-3。因此，FGF-2的趋化活性可单独由低亲和力FGFR介导，或由低亲和力和高亲和力FGFR（FGFR-1、FGFR-3或两者）共同介导。此外，在神经嵴细胞迁移过程中，在完整胚胎的中脑轴水平发现了FGF-2的差异定位。FGF-2蛋白表达在中脑神经嵴细胞定植的靶区域，特别是下颌间充质中占主导地位。这种特征性分布支持了FGF-2在小鼠胚胎中作为化学引诱剂指导中脑神经嵴细胞迁移的观点。虽然FGF-8在体外显示出趋化活性，但在用抗FGF-8中和抗体处理的外植体中观察到神经嵴细胞分散。这一结果表明，FGF-8在体内可能不是化学引诱剂。然而，用抗FGF-8处理的外植体中神经嵴细胞的分布与对照外植体或完整胚胎中的不同。在下颌弓中观察到FGF-2的极端分布，而FGF-8在上皮中表达。FGF-8可能在中脑神经嵴细胞迁移中起作用，其作用可能与FGF-2的差异定位有关。为了证实这一观点，我们对用FGF-8处理的外植体中FGF-2的分布进行了免疫组织化学检查，并使用外植体的RNA通过逆转录-聚合酶链反应分析了FGF-2基因表达水平。数据表明，在FGF-8处理的外植体中，FGF-2分布于整个间充质，且FGF-8可促进FGF-2的表达。因此，我们得出结论，下颌弓上皮中FGF-8的表达是FGF-2差异定位的先决条件，且FGF-2的分布模式对中脑神经嵴细胞迁移的趋化作用至关重要。
