炎性微环境下circBIRC6调控牙周膜干细胞成骨分化及牙周再生的机制研究

Periodontal ligament stem cells (PDLSCs) are important seed cells for periodontal tissue regeneration. However, the osteogenic differentiation and periodontal regeneration capacity of PDLSCs are significantly attenuated under the inflammatory microenvironment. Exploring the underlying molecular mechanisms is of great clinical significance for the repair and regeneration of periodontal tissues. Our preliminary studies compared the differentially expressed circRNAs between the PDLSCs from inflammatory environment and those from normal control, and found that the expression level of circBIRC6 was significantly increased following inflammatory stimulation. Downregulation of circBIRC6 promoted the osteogenic differentiation of PDLSCs in the inflammatory microenvironment. In addition, circBIRC6 might regulate prolyl hydroxylase domain-2 (PHD2) expression by absorbing miR-16-5p. Base on the above findings, this project will further elucidate the molecular mechanisms accounting for the role of circBIRC6 as the competitive endogenous RNA in regulating osteogenic differentiation and periodontal regeneration capacity of PDLSCs using in vitro inflammatory microenvironment model and rat model of periodontitis. Completion of our project will provide novel ideas for the experimental investigation and clinical translation of repairing periodontal defects.

牙周膜干细胞（PDLSCs）是实现牙周组织再生的重要种子细胞，但是其在炎症微环境下成骨分化及牙周再生能力明显下降，因此深入研究该现象的内在分子机制对于牙周组织修复再生有重要的临床意义。我们前期用高通量测序比较了炎症微环境与正常来源PDLSCs环状RNA的表达差异，发现炎症微环境下circBIRC6表达水平明显升高，抑制circBIRC6表达可促进体外炎症微环境下PDLSCs的成骨分化。初步证实了circBIRC6可能通过吸附miR-16-5p调控脯氨酸羟化酶-2（PHD2）的表达。本项目拟通过体外炎症微环境细胞模型和大鼠牙周炎骨缺损模型，进一步深入探讨circBIRC6作为竞争性内源RNA调控PDLSCs成骨分化及牙周再生的分子机制。本项目将为牙周组织缺损修复的实验研究及临床转化应用提供新思路。

项目背景：牙周韧带干细胞是牙周组织再生的理想种子细胞。众所周知，持续的炎症会显著损害牙周韧带干细胞的成骨分化能力。因此，在炎症微环境下保持牙周韧带干细胞的成骨潜能对于治疗牙周病骨损失很重要。我们的研究旨在探讨环状RNA BIRC6在调节炎症条件下牙周韧带干细胞成骨分化中的潜在作用。.研究内容：我们使用碱性磷酸酶染色、茜素红染色、定量实时聚合酶链反应、蛋白印迹和免疫荧光染色评估circBIRC6对牙周韧带干细胞成骨分化的影响。进行RNA pull-down和荧光素酶分析以探讨circBIRC6与miR-543之间的相互作用。然后进一步研究circBIRC6 / miR-543轴影响的下游信号通路。.重要结果及关键数据：我们发现在暴露于炎症刺激和牙周病组织的牙周韧带干细胞中，circBIRC6的表达水平比相应对照组高。降低circBIRC6增强了炎症条件下牙周韧带干细胞的成骨潜能，而升高circBIRC6则导致相反的结果。机理上，我们发现circBIRC6通过“吸附”miR-543来调节牙周韧带干细胞的成骨分化。更重要的是，我们证明circBIRC6 / miR-543轴通过PTEN / PI3K / AKT / mTOR信号通路调节牙周韧带干细胞的矿化能力在炎症微环境中。.科学意义：在炎症条件下，circBIRC6的下调通过调节miR-543/PTEN/PI3K/AKT/mTOR信号通路促进了牙周韧带干细胞的成骨分化。这些发现对于设计和实施针对circBIRC6的治疗策略以治疗牙周炎骨质丢失具有重要的启示。