颅骨锁骨发育不全致病基因RUNX2通过调控miR-31介导的牙槽骨改建过程影响牙齿替换的机制研究

颅骨锁骨发育不全综合征患者出现牙齿替换障碍，严重影响患者生活质量，但其机制仍未阐明。我们在前期工作中发现，其致病基因RUNX2突变下调牙源性细胞的成骨向分化能力和诱导破骨细胞分化的能力，并影响miR-31表达水平，而miR-31可调控成骨细胞和破骨细胞分化。因此，我们推测RUNX2突变通过调控miR-31干扰成骨细胞和破骨细胞分化，导致牙槽骨改建异常。针对此假说，我们将深入研究RUNX2突变对牙囊细胞诱导成骨细胞和破骨细胞分化能力的影响。其次，通过荧光素酶报告基因实验、染色质免疫共沉淀、慢病毒转染、“拯救”实验、miR-31靶基因预测和验证等方法，逐级分析RUNX2突变通过调控miR-31影响牙槽骨改建的机制。最后，通过观察吸附有RUNX2突变体细胞的牙根片段在裸鼠体内的骨形成和骨吸收情况，体内验证miR-31在RUNX2介导的牙槽骨改建中的作用，以揭示该综合征患者牙齿替换障碍的发病机制。

Delayed or failed eruption of the permanent successors is a typical feature of cleidocranial dysplasia, which seriously affects the quality of the patients’ life. However, the underlying mechanism is still unclear. In previous study, we found that RUNX2 mutation, the pathogenic gene of cleidocranial dysplasia, down-regulated the osteogenic potential of odontogenic cells and its ability to support osteoclast differentiation and also the expression of miR-31, which regulates the differentiation of osteoblast and osteoclast. Therefore, the hypothesis is raised that RUNX2 mutations disturb the alveolar bone remodeling during tooth replacement by regulating miR-31. In order to test this hypothesis, the effect of RUNX2 mutation on the differentiation of osteoblasts and osteoclasts by dental follicle cells, pre-osteoblast and pre-osteoclast will be first investigated. Thereafter, the mechanism of RUNX2 regulating bone remodeling via miR-31 will be explored by luciferase reporter assay, ChIP assay, lentivirus transfection, Rescue assay, miR-31 target gene prediction and validation. Lastly, the bone formation and bone resorption of the tooth root slice combined with scaffold material and pre-osteoblasts or pre-osteoclasts transfected with RUNX2 mutant was explored in nude mice in order to verify the in vitro findings. This study explores the mechanism that underlies impaired tooth replacement in cleidocranial dysplasia.