Much information is currently available for molecules in early odontogenesis, but there is limited knowledge regarding terminal cytodifferentiation of ameloblasts and odontoblasts for the determination of normal crown morphology. The present differential display PCR (DD-PCR) revealed that insulin-like growth factor-binding protein 5 (IGFBP5) was differentially expressed in molar tooth germs between the cap (before crown mineralization) and root formation (after crown mineralization) stages. Real-time PCR confirmed that the expression levels of IGFBP1–4 were not significantly changed but those of IGFBP5–7 were upregulated in a time-dependent manner. Immunoreactivities for IGFBP5–7 were hardly seen in molar germs at the cap/early bell stage and protective-stage ameloblasts at the root formation stage. However, the reactivity was strong in odontoblasts and maturation-stage ameloblasts, which are morphologically and functionally characterized by wide intercellular space and active enamel matrix mineralization. The localization of each IGFBP was temporospatial. IGFBP5 was localized in the nuclei of fully differentiated odontoblasts and ameloblasts, while IGFBP6 was localized in the apical cytoplasm of ameloblasts and odontoblasts with dentinal tubules, and IGFBP7 was mainly found in the whole cytoplasm of odontoblasts and the intercellular space of ameloblasts. IGFBP silencing using specific siRNAs upregulated representative genes for dentinogenesis and amelogenesis, such as DMP1 and amelogenin, respectively, and augmented the differentiation media-induced mineralization, which was confirmed by alizarin red s and alkaline phosphatase staining. These results suggest that IGFBP5–7 may play independent and redundant regulatory roles in late-stage odontogenesis by modulating the functional differentiation of ameloblasts and odontoblasts.

目前，关于早期牙发生的分子有很多信息，但关于成釉细胞和成牙本质细胞的终末细胞分化以确定正常牙冠形态的知识有限。目前的差异显示PCR（DD-PCR）显示，胰岛素样生长因子结合蛋白5（IGFBP5）在牙冠（牙冠矿化之前）和牙根形成（牙冠矿化之后）阶段的磨牙牙胚中存在差异表达。实时PCR证实IGFBP1-4的表达水平没有显着变化，但IGFBP5-7的表达水平以时间依赖性方式上调。 IGFBP5-7 的免疫反应性在帽/早期钟形阶段的磨牙胚和根形成阶段的保护阶段成釉细胞中几乎没有观察到。然而，成牙本质细胞和成熟阶段成釉细胞的反应性很强，其形态和功能特征是宽的细胞间隙和活跃的牙釉质基质矿化。每个 IGFBP 的定位都是时空的。 IGFBP5定位于完全分化的成牙本质细胞和成釉细胞的细胞核，IGFBP6定位于成牙本质细胞和有牙本质小管的成牙本质细胞的顶端细胞质，IGFBP7主要存在于成牙本质细胞的整个细胞质和成釉细胞的细胞间隙中。使用特定 siRNA 沉默 IGFBP 上调了牙本质发生和牙釉质发生的代表性基因，例如 DMP1 和釉原蛋白，并增强了分化培养基诱导的矿化，这一点通过茜素红和碱性磷酸酶染色得到证实。 这些结果表明IGFBP5-7可能通过调节成釉细胞和成牙本质细胞的功能分化，在牙齿发生的晚期发挥独立和冗余的调节作用。