Analysis of teratogenic mechanisms of retinoic acid on mammalian craniofacial morphogenesis---as models for human congenital anomalies

视黄酸（RA）是维生素a的天然代谢物，对哺乳动物的正常发育至关重要。然而，胚胎也表现出异常，过量的维生素A和其他天然和合成的类维生素A，包括类风湿性关节炎。类维生素a在实验动物中实验性诱导的颅面畸形被认为是人类先天性异常的模型，如特克-柯林斯综合征和迪·乔治综合征。由于颅神经嵴细胞在头面发育中起着重要的作用，我们在体外建立了可重复诱导典型颅面异常表型的条件，并检查了嵴细胞的行为。将9.0天和9.5天的大鼠胚胎在体外用全反式ra处理6小时，并进一步培养到体内胚胎11.5天的阶段。前一种治疗方法导致中枢神经系统和咽弓的后融合，而后一种治疗方法导致第一和第二弓以及三叉神经和听面神经节的融合。有趣的是，DiI标记研究显示来自前脑和前后脑的嵴细胞在融合弓内不混合。它支持了嵴细胞群体的差异身份仍然保持的观点。相比之下，在第9天的培养大鼠胚胎中，来自后脑前部的嵴细胞异位迁移到第二弓，这表明RA不仅影响后脑神经外胚层的身份，而且影响来自后脑的嵴细胞的身份。因此，RA以分期依赖的方式影响嵴细胞的迁移和脑神经的特征。在分子水平上也观察到RA效应的这种阶段依赖性，因为CRABP-I蛋白的分布模式在晚期治疗中没有变化，但在早期治疗中发生了急剧变化。

Retinoic acid (RA), a natural metabolite of vitamin A,is essential for normal mammalian development. However, embryos exhibit anomalies also by excess vitamin A and other natural and synthesized retinoids including RA.Craniofacial malformations experimentally induced with retinoids in laboratory animals are regarded as models for human congenital anomalies such as Treacher-Collins and Di George syndromes. Since cranial neural crest cells play important roles in development of head and face, here we have established in vitro conditions which reproducibly induce typical phenotypes of craniofacial abnormalities and examined behavior of the crest cells.9.0 day and 9.5 day rat embryos were treated in vitro with all-trans-RA for 6 hours and further cultured up to the stage corresponding to 11.5 day in vivo embryos. The former treatment induced posteriorization in the CNS and pharyngeal arches, while the latter treatment resulted in fusion of the first and second arches as well as that of the trigeminal and acousticofacial ganglia. Interestingly, DiI labeling study revealed that crest cells derived from the anterior and preotic hindbrain do not mix within the fused arch. It supports the idea that differential identities of the crest cell populations are still maintained. In contrast, crest cells from the anterior hindbrain ectopically migrate into the second arch in cultured rat embryos treated at 9.0day, suggesting that RA affected not only the identity of the hindbrain neuroectoderm but also that of the crest cells derived from the hindbrain. Thus, RA affected migration of crest cells and features of cranial nerves in a stage-dependent manner. Such stage dependency in RA's effects was also observed at the molecular level since the distribution pattern of CRABP-I protein was unchanged in the late-stage treatment but drastically altered in the early-stage treatment.