Drp1介导的线粒体功能障碍及NLRP3炎症小体活化在牙周炎中的作用及调控机制研究

Periodontitis, a highly prevalent disease, results in massive costs to the patients as well as the whole society. Oxidative stress induced tissue injury and inflammatory response were recognized as a crucial pathogenesis in the development and progression of periodontitis. However, the underlying mechanisms are not well understood. Our previous studies revealed that mitochondrial dysfunction played a crucial role in the destruction of periodontal tissues. Mitochondrial dysfunction mediated by dynamin-related protein 1 (Drp1), a key mitochondrial fission protein, represented an important pathological factor of oxidative stress damage. Furthermore, Drp1 was proved to be closely associated with the activation of NLRP3 inflammasome, a master regulator of periodontal inflammation. However, the specific regulating effect of Drp1 on periodontal oxidative stress response and related mechanisms remain elusive. In this proposal, we will establish injury and inflammatory model of human periodontal ligament cells to unveil the role of Drp1 in periodontal tissue destruction and activation of NLRP3 inflammasome with chemical and genetic approaches. We will further identify the regulatory mechanism of Drp1 at protein and gene level using gene chip technology, pull-down assay as well as other molecular biological detection methods. Ultimately, we will verify the regulating effect of Drp1 and related mechanism by adopting periodontitis animal model with inhibitor used and gene knock-out mice. This comprehensive study will not only provide new insights into the pathological mechanism of periodontitis, but also make a solid foundation for the development of new therapeutic strategies and targets in the prevention or treatment of periodontitis.

牙周炎发病率高、危害大，而氧化应激介导的组织破坏及炎症反应是牙周炎发生发展的重要病理机制之一，但其确切机理未见阐明。课题组研究证实：线粒体功能障碍在牙周炎组织破坏中起重要作用，而线粒体分裂蛋白Drp1介导的线粒体功能障碍是诱发氧化应激破坏效应的关键因素；同时Drp1与牙周炎炎症反应关键调控因子（NLRP3炎症小体）的活化密切相关。然而，Drp1对牙周炎氧化应激反应的确切调节作用及Drp1的分子调控机制尚不清楚。本研究拟构建人牙周膜细胞损伤及炎症模型，采用化学及基因手段正反双重探究Drp1在细胞损伤及NLRP3炎症小体活化中的调控作用；利用基因芯片、Pull-down联合质谱分析等技术从基因及蛋白水平全面阐明Drp1的调控机制；联合牙周炎动物模型，应用抑制剂及基因敲除小鼠，最终验证Drp1的调控作用及分子机制。本研究不但为牙周炎病理发生提供新的分子机制，而且为其防治提供新策略与新靶点。

牙周炎发病率高、危害大，而氧化应激介导的组织破坏及炎症反应是牙周炎发生发展的重要病理机制之一，其确切机理有待阐明。课题组研究证实：线粒体功能障碍在牙周炎组织破坏中起重要作用，而线粒体分裂蛋白Drp1介导的线粒体功能障碍是诱发氧化应激破坏效应的关键因素；同时Drp1与牙周炎炎症反应关键调控因子（NLRP3炎症小体）的活化密切相关。然而，Drp1对牙周炎氧化应激反应的确切调节作用及Drp1的分子调控机制尚不清楚。本研究构建过氧化氢诱导人牙周膜细胞损伤及炎症模型，采用化学手段干预，通过对人牙周膜细胞凋亡水平、成骨分化能力、炎症因子水平及线粒体功能的检测，全面探究Drp1在细胞损伤及NLRP3炎症小体活化中的调控作用；联合丝线结扎构建牙周炎动物模型及抑制剂的应用，通过对牙槽骨吸收水平、氧化应激和炎症相关蛋白表达水平的检测，揭示Drp1-ROS（活性氧簇）串话在牙周组织损伤和炎症反应中的重要调控作用及分子机制；通过采用临床样本，针对细胞凋亡水平及氧化应激和炎症相关蛋白表达水平的检测，探究Drp1-ROS与牙周炎发生发展的临床相关性。研究结果证实，Drp1介导的线粒体功能障碍及NLRP3炎症小体活化在牙周膜细胞损伤及炎症反应中起关键作用；Drp1-ROS串话是介导牙周组织损伤及炎症反应的重要病理机制，与此同时，该串话与牙周炎的发生发展高度相关，可成为牙周炎防治新靶点。本研究结果进一步丰富了牙周炎发病机制的理论体系，也为建立新的防治策略提供线索和理论基础。