p63基因调控非综合征型唇腭裂腭中嵴上皮细胞粘附功能的作用机制研究

Formation of the cleft palate (CP) is closely related to superficial morphology and reduction and/or loss of cell adhesion molecules of the medial edge epithelium (MEE). Our previous studies indicated that, in the MEE cells, TGF-β and dexamethasone (DEX) could affect formation of filopodia, and regulate cell polarity and cell adhesion. In addition, p63 gene, a key factor of regulating cell adhesion, played an important role in the embryonic development. However, there were not trials to take this as a breakthrough to explore the mechanism of CP. Based on these studies, the purposes of this project was using mouse model of CP from the cells, organs, maternal environment, animal experiments in vivo and in vitro in multi-level and multi-angle to explore changes of the superficial morphology and the mechanism of cell adhesion molecule expression in the MEE, to clarify the intrinsic regulation mechanisms of adhesion function of the MEE by investigating whether p63 could regulate the expression of chondroitin sulfate proteoglycan (CSPG) and E-cadherin through TGF-β signaling pathway. Simultaneously, we will attempt to reverse the changes of surface morphology and loss of adhesion molecule in the MEE which caused by DEX, to restore the adhesion of the MEE, to restart the palatal fusion, and to provide fresh ideas for the effective prevention and treatment of cleft palate.

腭中嵴上皮(medial edge epithelium, MEE)表面形态改变及粘附分子表达的减少和丧失与腭裂的形成具有密切关系，前期的研究表明TGF-β信号通路和地塞米松可以影响MEE细胞表面丝状伪足的形成，并参与调节MEE细胞的细胞极性和细胞粘附功能。p63基因在胚胎发育中也起着关键作用，是调节粘附分子表达的关键因子，但其内在的分子调节机制尚未有深入的探讨。基于此，本研究利用小鼠腭裂动物模型，从细胞、器官、母体内环境等体内体外多层次多角度来探讨p63基因如何通过TGF-β信号通路而影响MEE表面形态及粘附分子表达变化的内在调控机制，同时利用引入外源性p63基因尝试逆转地塞米松诱导的腭裂模型中MEE粘附分子丧失及表面形态改变的现象，恢复其粘附功能，重启腭融合过程，为腭裂的有效防治提供新的思路。

本项目以小鼠致畸腭裂模型的相关机制为背景，重点研究了氧化应激反应在TCDD致腭裂模型中的作用；TCDD对胎盘发育及母体免疫的影响；并正在研究microRNA在正常胚胎腭间充质细胞发育过程中的作用。 我们证实TCDD可在体内环境下影响tgf-β3的表达；体内实验证实谷胱甘肽稳态对于腭部的发育具有至关重要的作用：正常组E13.5腭胚突上皮表面细胞膨大，光滑，边界清晰，E14、E14.5腭胚突表面微丝，伪足增多；TCDD组E13.5腭胚突上皮表面细胞明显萎缩，边界欠清晰，E14、E14.5 腭胚突上皮表面变平滑，微丝，伪足数量减少；TCDD暴露可显著干扰小鼠胎盘的生长与发育，可干扰小鼠胎盘海绵滋养层细胞和迷路层血管的形成，引起小鼠外周免疫功能出现Th1极化，外周血IFH-γ和IL-4水平下降；体外培养腭胚突间充质细胞，转染mir-140-5p后，TGFβR1表达受抑制，TGFβ通路下游胶原蛋白亚单位col1a2表达受抑制。我们从不同角度研究了腭裂的病因机制，为了解揭示腭裂病因以及疾病的预防提供了科学依据。