牙源间充质特异性表达Wnt10a在牙根发育中的作用及其调控机制

Root is an important part of a tooth, which maintain the stabilization and perform the physiological functions of tooth. The developmental defect of tooth root is a type of tooth agenesis, which lead to early loss of tooth, impair the masticatory function, craniofacial development and oral health. The process of tooth development, the critical molecules involved in tooth crown formation and its reciprocal induction events have been addressed by multiple studies in recent years. However, the molecular regulation of tooth root development is rare to study and remains unclear. Wnt10a is specifically expressed in the dental mesenchymal cells lining the inner dentin surface during tooth root development. In previous study, we developed a new mouse strain, Wnt10afl/fl, and generated EIIa-Cre;Wnt10afl/fl mice, which completely knock out Wnt10a in all kinds of cells. We found that the molar root showed severe developmental defect in EIIa-Cre;Wnt10afl/fl mice, suggesting that mesenchymal cell specific expression of Wnt10a plays crucial role during tooth root development. In this study, we plan to generate three different Wnt10a conditional knockout mice model, including Gli1-CreERT2;Wnt10afl/fl mice, which targeting the tooth root progenitor cells; Wnt1-Cre;Wnt10afl/fl mice, which targeting the dental mesenchymal cells, and K14-Cre;Wnt10afl/fl mice, which targeting the dental epithelial cells. We plan to perform a series of experiments, to illustrate the molecular regulatory mechanism of mesenchymal cell specific expressed Wnt10a during tooth root development from the transcriptome, methylation, and non-coding RNA level. The results of our study will provide useful information for therapeutic strategy for the congenital tooth root defect and tooth regeneration.

牙根是牙齿维持稳固和行使咀嚼功能的重要部分。牙根发育异常是牙齿发育异常的一种，常导致牙齿早期丧失，影响咀嚼功能和颌面部发育。与牙冠发育中的信号网络被广泛研究和详尽阐明不同，牙根发育的系统性研究匮乏，牙根发育的调控机制知之甚少。在牙根发育过程中，Wnt10a持续地特异性表达在紧邻新生牙本质的间充质细胞内。我们在前期研究中创新性地构建了全敲除Wnt10a基因的EIIa-Cre;Wnt10afl/fl，该小鼠的下颌磨牙牙根存在明显的发育缺陷，提示我们牙源间充质特异性表达Wnt10a在牙根发育中存在着重要的作用。为深入研究调控机制，本项目拟构建在牙根前体细胞、牙间充质细胞和牙上皮细胞特异性敲除Wnt10a基因的三种条件性基因敲除小鼠模型，从转录组、甲基化和非编码RNA水平上阐明并验证牙源间充质特异性表达Wnt10a在牙根发育中的作用和调控机制，寻找相关靶点并探讨其在牙齿再生治疗中的意义与可行性。

人类WNT10A (Wnt family member 10A) 双等位基因突变可导致牙根发育缺陷，即牛牙症。本研究通过构建三种Wnt10a基因条件性敲除小鼠模型，探讨Wnt10a在牙根发育中的作用及其调控机制。主要研究结果：1) 在牙根发育过程中，Wnt10a在上皮和间充质细胞中均有表达；2) EIIa-Cre;Wnt10afl/fl小鼠及K14-Cre;Wnt10afl/fl小鼠的牙根发育出现异常，表现为牛牙症。证实了上皮内的Wnt10a在牙根发育过程中起关键作用；3) 上皮内敲除Wnt10a后，邻近未来根分歧区域的Wnt4表达代偿性上升，导致该位置的间充质细胞增殖显著增加，可能阻碍了HERS在根分歧区域水平向生长和融合，从而导致牛牙症的发生。本研究证实了上皮内表达的Wnt10a在牙根形态发育过程中不可或缺的作用。Wnt10a和Wnt4信号调控等级的发现和功能的阐明，将极大地加深我们对牙齿正常发育和发育异常的分子和细胞机制的理解。本研究的结果为未来阻止和纠正牙根发育缺陷提供了治疗策略，同时对利用Wnt10a优化生物性牙根再生有新的启示。