EndMT在正畸牙周改建中的作用及机制研究

The essence of orthodontic treatment is periodontal tissue remodeling under orthodontic force. The function of mesenchymal stem cells (MSCs) in periodontal remodeling has been confirmed. The pilot studies of our research group found that the number of multipotent stem-like cells in periodontal ligament increased significantly after orthodontic loading, which distributed mainly in perivascular area. However, the origin of these newly generated MSCs is not clear. In 2010, Nat Med reported vascular endothelial cells could convert into multipotent stem-like cells by the mechanism of endothelial-to-mesenchymal transition (EndMT); besides, this process was not just confined to the stages of embryonic development or pathological fibrosis, but also occurs under physiological states. In 2011, Cell Stem Cell suggested that the perivascular endothelial cells could be the in vivo counterparts of cultured MSCs. Based on these previous studies, we speculate that the newly emerged multipotent stem-like cells in periodontal ligament might have a substantial connection with vascular endothelial cells, and EndMT might be the mechanism that regulate the periodontal remodeling. Therefore, our project aims to observe the occurance of EndMT in periodontal remodeling by establishing animal models of orthodontic tooth movement. Moreover, we plan to establish the cultural system under stress stimuli, aiming to explore the effect of mechanical stimuli on EndMT and relevant pathway centered on ALK2. This project would not only lay solid foundation to optimize the theory of orthodontic periodontal remodeling, but also establish the approach based on EndMT to intervene the teeth movement in vivo.

口腔正畸治疗的实质是矫治力作用下的牙周组织改建。MSCs在牙周组织改建中的作用现已证实。课题组前期研究发现正畸牙移动牙周组织内MSCs数量明显上调且集中在血管周围。但新生MSCs来源目前尚不清楚。2010年Nat Med首次报道血管内皮细胞通过EndMT转化形成MSCs样细胞，并证实EndMT并非仅局限在胚胎发育阶段或组织纤维化的病理过程，生理状态下亦可发生。2011年Cell stem cell评述认为体内血管内皮细胞相当于MSCs的储存库。我们推测正畸牙周组织内MSCs新生与血管密切关系，EndMT是正畸牙周组织MSCs新生、牙周组织改建的重要机制。因此，本项目拟应用正畸牙移动模型，观察牙周组织改建中的EndMT发生；并通过应力培养体系，探讨应力刺激对EndMT的影响及以ALK2为核心的信号通路。该项目将为完善正畸牙周改建理论及构建基于EndMT机制的正畸牙移动临床干预路径提供依据。

口腔正畸治疗的实质是矫治力作用下的牙周组织改建。虽然MSCs在牙周组织改建中的作用现已被证实，但正畸牙移动中牙周组织新生MSCs的来源及部分功能目前尚不清楚。本项目建立了人正畸牙移动模型，证明正畸力学载荷力能刺激内皮细胞发生EndMT，正畸力刺激牙周膜内MSC数量增加，力学刺激下牙周膜内的MSCs被激活，这些干细胞可能来源于血管周围的微环境。建立血管内皮细胞（HUVECs）体外力学加载模型，研究显示压应力及张应力均可刺激EndMT的发生但作用有限，而低强度脉冲超声及BMP4的作用相对更加明显；建立过表达和缺失表达CHD7体系，通过siRNA介导的knockdown技术下调CHD7的表达后，MSC成骨能力下降，而CHD7的过表达可以增强MSC成骨能力，机制显示CHD7可与BMP信号通路的下游因子SMAD1相互作用，BMP2促进了CHD7与主要成骨转录因子SP7增强子位点的结合，体内实验也验证了CHD7下调后MSC成骨能力的下降，证实了CHD7在新生MSC成骨分化中的重要作用；研究发现正畸应力刺激可诱导牙髓发生无菌性炎症和牙髓微环境改变。通过提高组织内IL17A表达，引起实验大鼠牙髓内炎症反应，从而激活DPSCs，增强其自我复制及多向分化能力。；建立正畸力对大鼠牙移动模型，探究正畸力对大鼠牙移动模型DPSCs的影响及其影响机制。发现正畸应力刺激可诱导牙髓发生无菌性炎症和牙髓微环境改变。通过提高组织内IL17A表达，引起实验大鼠牙髓内炎症反应，从而激活DPSCs，增强其自我复制及多向分化能力。哌嗪类衍生物三甲氧苄嗪（TMZ）对EndMT的影响近年来已经被报道，本项目TMZ处理间充质干细胞，评估TMZ对维持新生间充质干细胞代谢及线粒体功能稳定性的作用，结果显示TMZ对氧化应激诱导的MSCs细胞凋亡具有保护作用，TMZ预处理后可一定程度上降低H2O2对线粒体的损伤作用，TMZ处理可降低细胞氧化应激水平，可能通过调节Caspase3、Bax、Bad 和Bcl2等凋亡相关关键蛋白发挥作用。该项目将为完善正畸牙周改建理论及理解正畸牙移动牙周间充质干细胞新生、分化、稳态维持提供实验依据。