LncRNA调控牙周病微环境来源PDLSCs对牵张力反应的机制研究

In the modern society, more and more periodontitis patients are seeking orthodontic treatment for a charming smile. However, the local inflammatory injury often results in the poor response of periodontal tissue to orthodontic force stimulation, and the underlying mechanism is still unknown until now. We have confirmed that the periodontitis microenvironment could damage the osteogenesis ability of periodontal ligament stem cells (PDLSCs), consequently leading to its more sensitivity and less tolerance to mechanical strain. The array analysis also showed the abnormal LncRNA expression, especially Lnc8604 and Lnc8781, in the PDLSCs from periodontitis tissue (PPDLSCs) when the cells received the mechanical strain. Additionally, the osteogenesis related MAPK signal pathway and Ca2+ signal pathway are inhibited. Therefore, the abnormal expression of Lnc8604 and Lnc8781 in PPDLSCs in the periodontitis microenvironment may disturb the MAPK signal pathway and Ca2+ signal pathway, thus affecting the periodontal tissue remodeling under the mechanical stimulus. For verifying the hypothesis, we will establish the mechanical strain loading system and periodontitis model. We will observe the influence of LncRNA8604 and LncRNA8781 expression to the osteogenesis ability of PPDLSCs under mechanical stimuli via cytology, molecular biology, bioinformatics and histomorphology. Moreover, with the aid of in vivo and in vitro experiments, we will explore the mechanism of MAPK and Ca2+ signaling pathways in mediating the process of LncRNA affecting PPDLSCs osteogenesis ability.The whole project is expected to provide new experimental basis for improving the regeneration ability of PDLSCs in periodontitis environment and the orthodontic effects of periodontitis patients in the future.

现代社会越来越多牙周炎患者寻求正畸治疗，但炎症损害常导致牙周组织对正畸力反应不佳，其调控机制尚不明确。我们前期发现，牙周炎微环境会破坏牙周膜干细胞（PDLSCs）成骨能力，并导致其对牵张力敏感性增强而耐受性降低；芯片分析发现，牵张力加载引发牙周炎来源PDLSCs（PPDLSCs）中LncRNA表达异常，尤以Lnc8604和Lnc8781较为明显，PPDLSCs中成骨相关MAPK及Ca2+信号通路也受到抑制，这可能是导致牙周病患者力学刺激下牙周组织改建较差的原因。本项目拟利用分子生物学、生物信息学和组织学等方法，观察Lnc8604和Lnc8781的表达对牵张力加载下PPDLSCs成骨能力的影响；并借助体内体外实验，深入探讨MAPK及Ca2+信号通路在介导两种LncRNA对PPDLSCs成骨能力影响中的调控机制，为提高牙周炎环境下PDLSCs的再生能力、改善牙周炎患者的正畸疗效提供策略和依据。

牙周病的炎症损害常导致牙周组织对正畸力反应不佳，我们前期发现，牙周炎微环境会破坏牙周膜干细胞（PDLSCs）成骨能力，并导致其对牵张力敏感性增强而耐受性降低，但具体机制仍不清晰。在本项目中，通过芯片分析我们发现，牵张力加载引发牙周炎来源PDLSCs（PPDLSCs）中大量LncRNA表达异常，KEGG生物学通路分析显示，在HPDLSCs中发生变化的mRNA多数与正常生物学过程相关，而在PPDLSCs中差异表达的mRNA则有很多涉及疾病相关通路，力学相关信号通路也被抑制，说明由于牙周病微环境的刺激导致PPDLSCs对静态牵张力的反应发生了病理性改变。为进一步探索其机制，我们对H-PDLSCs及P-PDLSCs进行成骨诱导筛选成骨相关lncRNA，结果显示lnc01135在H-PDLSCs及P-PDLSCs成骨分化后表达量明显增加，同时其表达量与Runx2的表达具有明显相关性。接下来我们通过慢病毒有效上调及下调lnc01135的表达，发现lnc01135对12%SMS作用下HPDLSCs和PPDLSCs的成骨分化能力均有积极的调控作用；对HPDLSCs促进程度优于PPDLSCs；而在未加力状态下，lnc01135在PPDLSCs中的表达量较HPDLSCs明显降低，其对12%SMS刺激的应答反应也明显减弱。CeRNA分析结果显示lnc01135 与miR-17-5p等5个成骨相关的microRNA存在结合位点，下一步我们将继续从lncRNA-microRNA相结合的角度试图深入分析lnc01135调控PDLSCs在牵张力作用下成骨分化的具体机制。本研究为改善牙周病患者的正畸治疗效果奠定了一定的理论基础。