Abstract：
OBJECTIVE: This study aimed to reveal the effects of SET domain bifurcated 1 (SETDB1) on epithelial cells during tooth development.
METHODS: We generated conditional knockout mice (Setdb1fl/fl,Keratin14-Cre+ mice), in which Setdb1 was deleted only in epithelial cells. At embryonic day 14.5 (E14.5), immunofluorescence staining was performed to confirm the absence of SETDB1 within the epithelium of tooth embryos from Setdb1fl/fl,Keratin14-Cre+ mice. Mouse embryos were harvested after reaching embryonic day 13.5 (E13.5), and sections were prepared for histological analysis. To observe tooth morphology in detail, electron microscopy and micro-CT analysis were performed at postnatal months 1 (P1M) and 6 (P6M). Tooth embryos were harvested from postnatal day 7 (P7) mice, and the epithelial components of the tooth embryos were isolated and examined using quantitative RT-PCR for the expression of genes involved in tooth development.
RESULTS: Setdb1fl/fl,Keratin14-Cre+ mice exhibited enamel hypoplasia, brittle and fragile dentition, and significant abrasion. Coronal sections displayed abnormal ameloblast development, including immature polarization, and a thin enamel layer that detached from the dentinoenamel junction at P7. Electron microscopic analysis revealed characteristic findings such as an uneven surface and the absence of an enamel prism. The expression of Msx2, Amelogenin (Amelx), Ameloblastin (Ambn), and Enamelin (Enam) was significantly downregulated in the epithelial components of tooth germs in Setdb1fl/fl,Keratin14-Cre+ mice.
CONCLUSIONS: These results indicate that SETDB1 in epithelial cells is important for tooth development and clarify the relationship between the epigenetic regulation of SETDB1 and amelogenesis imperfecta for the first time.
METHODS: We generated conditional knockout mice (Setdb1fl/fl,Keratin14-Cre+ mice), in which Setdb1 was deleted only in epithelial cells. At embryonic day 14.5 (E14.5), immunofluorescence staining was performed to confirm the absence of SETDB1 within the epithelium of tooth embryos from Setdb1fl/fl,Keratin14-Cre+ mice. Mouse embryos were harvested after reaching embryonic day 13.5 (E13.5), and sections were prepared for histological analysis. To observe tooth morphology in detail, electron microscopy and micro-CT analysis were performed at postnatal months 1 (P1M) and 6 (P6M). Tooth embryos were harvested from postnatal day 7 (P7) mice, and the epithelial components of the tooth embryos were isolated and examined using quantitative RT-PCR for the expression of genes involved in tooth development.
RESULTS: Setdb1fl/fl,Keratin14-Cre+ mice exhibited enamel hypoplasia, brittle and fragile dentition, and significant abrasion. Coronal sections displayed abnormal ameloblast development, including immature polarization, and a thin enamel layer that detached from the dentinoenamel junction at P7. Electron microscopic analysis revealed characteristic findings such as an uneven surface and the absence of an enamel prism. The expression of Msx2, Amelogenin (Amelx), Ameloblastin (Ambn), and Enamelin (Enam) was significantly downregulated in the epithelial components of tooth germs in Setdb1fl/fl,Keratin14-Cre+ mice.
CONCLUSIONS: These results indicate that SETDB1 in epithelial cells is important for tooth development and clarify the relationship between the epigenetic regulation of SETDB1 and amelogenesis imperfecta for the first time.
摘要：
目的：本研究旨在揭示SET结构域分叉1（SETDB1）在牙齿发育过程中对上皮细胞的影响。
方法：我们产生了条件性敲除小鼠（Sedb1fl/fl，Keratin14-Cre+小鼠)，其中Setdb1仅在上皮细胞中删除。在胚胎第14.5天（E14.5），进行免疫荧光染色以确认来自Setdb1fl/fl的牙齿胚胎上皮内不存在SETDB1，Keratin14-Cre+小鼠。在达到胚胎第13.5天（E13.5）后收获小鼠胚胎，并准备切片进行组织学分析。为了详细观察牙齿形态，在出生后1个月（P1M）和6个月（P6M）进行电子显微镜和显微CT分析。从出生后第7天（P7）小鼠中收获牙齿胚胎，分离牙齿胚胎的上皮成分，并使用定量RT-PCR检测牙齿发育相关基因的表达。
结果：Setdb1fl/fl，Keratin14-Cre+小鼠表现出釉质发育不全，脆弱和脆弱的牙列，和显著的磨损。冠状切片显示出成釉细胞发育异常，包括不成熟的两极分化，以及在P7处从牙釉质交界处脱离的薄釉质层。电子显微镜分析显示了特征性的发现，例如不平坦的表面和没有搪瓷棱镜。表达Msx2,Amelogenin(Amelx),Ameloblastin（Ambn），在Setdb1fl/fl中，Enamelin（Enam）在牙胚的上皮成分中显著下调，Keratin14-Cre+小鼠。
结论：这些结果表明，上皮细胞中的SETDB1对牙齿发育很重要，并首次阐明了SETDB1的表观遗传调控与牙釉质发育不全之间的关系。
方法：我们产生了条件性敲除小鼠（Sedb1fl/fl，Keratin14-Cre+小鼠)，其中Setdb1仅在上皮细胞中删除。在胚胎第14.5天（E14.5），进行免疫荧光染色以确认来自Setdb1fl/fl的牙齿胚胎上皮内不存在SETDB1，Keratin14-Cre+小鼠。在达到胚胎第13.5天（E13.5）后收获小鼠胚胎，并准备切片进行组织学分析。为了详细观察牙齿形态，在出生后1个月（P1M）和6个月（P6M）进行电子显微镜和显微CT分析。从出生后第7天（P7）小鼠中收获牙齿胚胎，分离牙齿胚胎的上皮成分，并使用定量RT-PCR检测牙齿发育相关基因的表达。
结果：Setdb1fl/fl，Keratin14-Cre+小鼠表现出釉质发育不全，脆弱和脆弱的牙列，和显著的磨损。冠状切片显示出成釉细胞发育异常，包括不成熟的两极分化，以及在P7处从牙釉质交界处脱离的薄釉质层。电子显微镜分析显示了特征性的发现，例如不平坦的表面和没有搪瓷棱镜。表达Msx2,Amelogenin(Amelx),Ameloblastin（Ambn），在Setdb1fl/fl中，Enamelin（Enam）在牙胚的上皮成分中显著下调，Keratin14-Cre+小鼠。
结论：这些结果表明，上皮细胞中的SETDB1对牙齿发育很重要，并首次阐明了SETDB1的表观遗传调控与牙釉质发育不全之间的关系。
