牙齿缺失是口腔健康的重大问题，利用干细胞通过生物工程技术生成再生牙是重要的研究方向。临床可获取的成牙干细胞数量有限，需体外培养进行扩增。然而扩增细胞成牙潜能消失，这成为研究的最大瓶颈。我们之前的研究表明牙胚间充质的扩增细胞在与新鲜细胞混合产生的群效应下可恢复成牙潜能，其产生机制不明。预实验通过转录组测序分析发现经群效应恢复成牙潜能的扩增细胞的Dlk1表达上调；添加DLK1蛋白可使扩增细胞成牙相关基因Pax9及Msx1表达升高，Notch通路靶基因Hes1表达下降。结合文献报道我们提出假说：DLK1介导Notch通路在群效应恢复扩增细胞成牙潜能的过程中起到调控作用。本研究拟通过上皮-间充质重组成牙实验等方法验证DLK1处理及Notch通路活性调节对扩增细胞成牙潜能的影响，进一步揭示扩增细胞成牙潜能开关的分子调控机制，以期建立无需新鲜细胞参与的分子调控方法恢复扩增细胞成牙潜能，突破研究瓶颈。

Tooth loss is a major problem of oral health. Stem cell-based whole tooth bio-engineering is a promising alternative treatment to the traditional artificial teeth, where cell expansion is required due to the limited number of odontogenic cells that can be obtained in clinic. However, the odontogenic induction capacity of the cells is lost after in vitro culture, which becomes the greatest bottleneck of the research on tooth regeneration in a clinical scenario. We have demonstrated that cultured embryonic dental mesenchymal cells (eDMCs) can be rescued from failure of tooth induction by the community effect created through mixing with fresh eDMCs. However, the mechanism of this effect is still unclear. In pre-experiments, RNA-seq data analysis reveals that Dlk1 was upregulated in the cultured eDMCs with restored odontogenic inductivity; DLK1 treatment can upregulate odontogenic genes Pax9 and Msx1, downregulate Notch signaling pathway target gene Hes1 in cultured eDMCs. Hence, based on the preliminary data and relative studies reported in literature, we hypothesize that DLK1-mediated Notch signaling pathway plays an important role in the regulation of the recovery of odontogenic inductivity in cultured eDMCs through community effect. This study will utilize odontogenic epithelia-mesenchyme recombination experiment etc. to verify the effect of DLK1 treatment and Notch signaling pathway regulation on the odontogenic potential of cultured eDMCs. This study will further elaborate the molecular mechanism of the odontogenic potential regulation of proliferated eDMCs, provide basis to break through the bottleneck, which is to establish method to rescue the odontogenic potential of cultured eDMC through molecular manipulation without contribution from fresh eDMCs.