S1P信号通路对牙骨质细胞应力转导的作用机制研究

Cementum covering the tooth root dentin is a barrier for the tooth to resist the mechanical strain, which is associated with orthodontic tooth movement and root resorption. Cementocytes, embedded in the cementum, may be involved in the regulation of mechanotransduction during orthodontic tooth movement. Much less is known about the biological property and the mechanism in mechanotransduction of cementocyte due to the difficulties in isolating cementocytes. We have isolated and established a novel murine immortalized cementocyte cell line and observed that these cells could response to flow fluid shear stress. Moreover, S1P signaling pathway was detected in the mineralization of the cell line in our recent study. S1P signaling pathway has been reported to modulate the process of osteocyte mechanotransduction, thus S1P signaling pathway may also play a key role in cementocyte. Therefore, we proposed that S1P signaling pathway may act as the upstream mediators, modulate the downstream gene expression, thus regulating cementocyte mechanotransduction. The experiment will be performed with our cell line under compressive loading in vitro and tooth intrusion in vivo to mimic the microenvironment in orthodontic tooth movement. The function and mechanism of S1P signaling pathway in cementocyte mechanotransduction will be investigated with μCT, scanning electron microscope, techniques of cryobiology and molecular biology. The agonist or antagonist will be administrated to up-regulate or down-regulate S1P signaling pathway. This, in turn, will elucidate the mechanism of S1P signaling pathway in cementocyte mechanotransduction. The other genes which are involved in cementocyte mechanotransduction will be investigated as well. The present study will contribute to proper tooth movement in clinic and provide insight into the mechanism of tooth resorption.

牙骨质是牙根抵抗应力的屏障，与牙移动及正畸力下的牙根吸收密切相关。牙骨质细胞可能是牙骨质感受应力并调控牙根吸收的关键细胞。然而由于其分离难度极大，国内外一直缺乏有效的研究模型对其应力感应机制进行深入研究。申请人前期建立新型永生化牙骨质细胞系，发现其能感受应力，且矿化过程中S1P信号通路基因表达活跃。S1P信号通路参与调控骨细胞应力转导，因此很可能也是维持牙骨质稳态的关键。由此我们提出：S1P信号通路可能作为上游关键因子调控下游的基因表达，介导了牙骨质的应力转导。本课题拟通过压应力下牙骨质细胞体外培养及牙压入移动的动物模型，模拟正畸临床中的应力环境，并通过上调或下调S1P信号通路，采用μCT、扫描电镜、细胞生物学及分子生物学等技术，研究压应力下S1P信号通路对牙骨质细胞应力转导的作用机制，并筛选介导应力转导的其他潜在关键基因。本课题的实施将为实现健康正畸牙移动及防治牙根吸收提供新的理论依据。

牙骨质是牙根抵抗应力的屏障，与牙移动及正畸力下的牙根吸收密切相关。牙骨质细胞可能是牙骨质感受应力并调控牙根吸收的关键细胞。然而由于其分离难度极大，国内外一直缺乏有效的研究模型对其应力感应机制进行深入研究。本项目利用申请人前期建立的新型永生化牙骨质细胞系进行体外压应力加载，阐明了牙骨质细胞应力转导过程中的关键因子及S1P信号通路在其中所起的重要调控作用；同时借助大鼠磨牙压低移动的体内实验模型，进一步验证了S1P信号通路对牙骨质细胞应力转导的作用，揭示了S1P信号通路对牙骨质稳态的调控机制。此外，我们还初步探索了S1P信号通路在牙骨质细胞受应力刺激后对破骨细胞的作用，为深入探讨牙根吸收的机制做了铺垫。研究结果提示：在压应力刺激下，牙骨质细胞中应力转导相关因子PGE2、β-catenin的表达水平升高，同时S1P浓度及S1PR1、S1PR2的表达量降低，说明S1P信号通路参与了牙骨质细胞的应力转导过程；通过激动剂及拮抗剂调节S1P信号通路后发现，ALP、Runx2、RANKL/OPG、SOST等牙骨质稳态相关因子表达出现变化，证明应力下S1P信号通路可能参与了应力刺激下牙骨质稳态的调控。牙根吸收的体内研究表明，S1P信号通路可能是通过其下游关键调控因子NFATc1来影响牙骨质稳态相关蛋白的表达水平。体内体外实验结果分析发现，不同力值的应力刺激可能对牙骨质稳态产生不同影响。因此，我们进行了初步探究，发现不同力值的应力刺激下牙骨质细胞能够对破骨前体细胞的分化、成熟产生不同影响。本项目完满完成了压应力下牙骨质细胞应力转导的作用机制探究，并确定了S1P信号通路在牙骨质细胞应力转导及维持牙骨质稳态中的重要作用，为后期深入探究牙根吸收的机制奠定了坚实的前期研究基础，也为精准预防正畸所致牙根吸收提供了可能的新药靶点，具有重要临床应用价值。