低强度脉冲超声通过METTL3调控EphB4 m6A甲基化修饰促进正畸牙槽骨改建的机制研究

The insufficiency of orthodontic alveolar bone remodeling seriously affects the realization of orthodontic targets and the maintenance of the therapeutic outcome. How to promote bone formation in orthodontic alveolar bone remodeling is the focus of orthodontic clinical research. Based on the preliminary study of our project group, it is found that LIPUS can effectively promote the regeneration of periodontal tissue defects, the applicant propose that through utilizing the regulating effect of LIPUS on osteogenesis could increase bone formation in orthodontic alveolar bone remodeling, and so then further improve the efficiency and safety of orthodontic treatment. In this project, we first evaluate the effect of LIPUS on orthodontic alveolar bone remodeling through in vivo experiments to clarify its clinical application significance, subsequently the in vitro model will be used to study the effect of LIPUS on the proliferation and differentiation of BMSC and screen its effector molecules. To analyze the signal pathways and action targets of the LIPUS functional effects, our study combined with previous experimental results and then focus on METTL3-mediated m6A-demethylation. On this basis, the role of METTL3 in LIPUS regulation of osteogenic differentiation and the molecular mechanism of FOXO3-METTL3-EphB4 signaling axis will be validated in multi-angles. Results of the project will provide a new idea for broadening orthodontic adjuvant therapy and provide theoretical basis for the application of LIPUS in orthodontic clinic.

正畸牙槽骨改建不足严重影响着治疗目标的实现及效果的维持，已成为正畸治疗的生物学瓶颈。基于LIPUS在牙周组织再生中的有效应用，项目组前期实验发现LIPUS可促进正畸牙槽骨改建并加快正畸牙移动。为解析LIPUS发挥功能效应的信号途径及作用靶点，前期实验通过芯片数据挖掘聚焦于RNA甲基转移酶METTL3，并证实该分子在LIPUS处理后表达上调，且在LIPUS影响BMSC成骨分化中具有明确功能。进一步研究表明，LIPUS可能通过PI3K/Akt/FOXO3信号轴影响METTL3转录表达，进而通过其介导的EphB4 m6A甲基化修饰参与调控细胞成骨分化。本项目拟在此基础上结合体内、体外模型，多层次验证METTL3在LIPUS影响细胞成骨分化中的作用方式及其分子机制。项目结果将为我们拓宽正畸辅助治疗手段提供新的思路，为揭示LIPUS调控细胞成骨分化的深入机制实现突破。

LIPUS可以加速正畸牙移动并有利于正畸牙槽骨改建。成骨细胞的骨形成和破骨细胞的骨吸收之间的动态耦联平衡控制着骨改建，且其分化受到LIUPS声学机械效应的影响。然而，其响应LIPUS刺激的感受模式及其调节机制仍然未知。在这项研究中，我们通过大鼠正畸牙移动模型发现LIPUS诱导的正畸牙槽骨改建与成骨-破骨细胞间交互对话及EphrinB2/EphB4信号转导有关。体外实验发现LIPUS促进BMSCs成骨分化和EphB4表达，尤其是在与BMMs诱导的破骨细胞共培养条件下。我们证明了LIPUS加强了正畸力环境下牙槽骨组织中BMSCs与破骨细胞表面的EphrinB2/EphB4信号交互，上调BMSCs膜表面Ephb4受体并将 LIPUS 相关的机械信号转导到细胞内细胞骨架上，改变细胞骨架聚合并激活Hippo信号通路中的YAP入核，从而调节BMSCs细胞的增殖和成骨分化。