School of Stomatology, Tianjin Medical University, Tianjin, 300014, China.
Immunology Department, Key Laboratory of Immune Microenvironment and Disease, Ministry of Education, Tianjin Medical University, Tianjin, 300014, China.
Biochem Biophys Res Commun. 2023 Jun 30;663:47-53. doi: 10.1016/j.bbrc.2023.04.053. Epub 2023 Apr 18.
Enamel hypoplasia is a tooth development defection due to the disruption of enamel matrix mineralization, manifesting as chalky white phenotype. Multiple genes may be involved in this tooth agenesis. It has been proved that ablation of coactivator Mediator1 (Med1) switches the cell fate of dental epithelia, resulting in abnormal tooth development via Notch1 signaling. Smad3 (-/-) mice displays the similar chalky white incisors. However, the expression of Smad3 in Med1 ablation mice and the impact of Med1 on functional integration between Smad3 and Notch1 remains unclear. Cre-loxP-based C57/BL6 mice with epithelial-specific Med1 knockout (Med1 KO) backgrounds were generated. Mandibles and dental epithelial stem cells (DE-SCs) from incisors cervical loop (CL) were isolated from wild-type (CON) mice and Med1 KO mice. Transcriptome sequencing was used to analyze the differences of CL tissue between KO and CON mice. The results revealed the enrichment of TGF-β signaling pathway. qRT-PCR and western blot were performed to show the gene and protein expression of Smad3, pSmad3, Notch1 and NICD, the key regulators of TGF-β and Notch1 signaling pathway. Expression of Notch1 and Smad3 was confirmed to be down-regulated in Med1 KO cells. Using activators of Smad3 and Notch1 on Med1 KO cells, both pSmad3 and NICD were rescued. Moreover, adding inhibitors and activators of Smad3 and Notch1 to cells of CON groups respectively, the protein expressions of Smad3, pSmad3, Notch1 and NICD were synergistically affected. In summary, Med1 participates in the functional integration of Smad3 and Notch1, thus promoting enamel mineralization.
釉质发育不全是一种由于釉质基质矿化中断导致的牙齿发育缺陷，表现为粉笔样白的表型。多个基因可能参与了这种牙齿缺失。已经证明，辅激活子 Mediator1 (Med1) 的消融会改变牙上皮细胞的命运，通过 Notch1 信号导致牙齿发育异常。Smad3 (-/-) 小鼠表现出类似的粉笔样白切牙。然而，Med1 在 Med1 消融小鼠中的表达以及 Med1 对 Smad3 和 Notch1 之间功能整合的影响尚不清楚。利用基于 Cre-loxP 的 C57/BL6 小鼠，构建了上皮细胞特异性 Med1 敲除 (Med1 KO) 背景的小鼠。从野生型 (CON) 小鼠和 Med1 KO 小鼠的切牙颈环 (CL) 中分离下颌骨和牙上皮干细胞 (DE-SCs)。利用转录组测序分析 KO 和 CON 小鼠 CL 组织之间的差异。结果显示 TGF-β 信号通路的富集。qRT-PCR 和 Western blot 用于显示 Smad3、pSmad3、Notch1 和 NICD 的基因和蛋白表达，它们是 TGF-β 和 Notch1 信号通路的关键调节因子。证实 Med1 KO 细胞中 Notch1 和 Smad3 的表达下调。在 Med1 KO 细胞上使用 Smad3 和 Notch1 的激活剂，pSmad3 和 NICD 均被挽救。此外，分别向 CON 组细胞添加 Smad3 和 Notch1 的抑制剂和激活剂，Smad3、pSmad3、Notch1 和 NICD 的蛋白表达受到协同影响。总之，Med1 参与了 Smad3 和 Notch1 的功能整合，从而促进了釉质矿化。
