DOT1L介导肌肉细胞线粒体通透性在骨质疏松的分子机制研究

As China's population ages, the incidence of osteoporosis will rise rapidly, putting a heavy burden on the national economy and society. It is of great significance to study the pathogenesis of osteoporosis and find new strategies to prevent and treat osteoporosis. Muscle cells play an important role in the pathogenesis of osteoporosis. The applicant of this project found that the expression of histone H3K79 methyltransferase DOT1L was low in muscle cells, serum and serum derived exosome of patients with osteoporosis. DOT1L can affect the secretion of Irisin by muscle cells to regulate osteoblast differentiation, osteoclast differentiation and bone erosion through MPTP opening, but the specific molecular mechanism needs further clarification. Therefore, the project is planned to conduct the following studies: 1. To clarify the specific molecular mechanism by which DOT1L epigenetically regulates the transcription of CypD gene; 2. Does DOT1L-mediated opening of MPTP regulate FNDC5 expression and Irisin secretion through the ROS/Sirt1/PGC1-α pathway? 3. Explore whether there is a new target for the diagnosis and treatment of osteoporosis in the DOT1L-MPTP-Irisin pathway in muscle cells. Through the above research, it is hoped to provide new strategies and theoretical support for the diagnosis and prevention of osteoporosis.

随着我国人口的老龄化，骨质疏松的发病率将迅速上升，给国家经济和社会带来沉重的负担。深入研究骨质疏松发病机制，寻找防治骨质疏松的新策略，具有重要的意义。肌肉细胞在骨质疏松发病中具有重要作用。申请者前期发现，骨质疏松患者肌肉细胞、血清和血清外泌体中组蛋白H3K79甲基转移酶DOT1L低表达。DOT1L通过MPTP打开影响肌肉细胞分泌Irisin，调节成骨细胞分化、破骨细胞分化和蚀骨能力，但具体分子机制需要进一步阐明。因此，本项目将进行如下研究：1.明确DOT1L表观调控CypD基因转录的具体分子机制；2.DOT1L介导的MPTP的打开是否通过ROS/Sirt1/PGC1-α通路，对FNDC5表达和Irisin分泌进行调节；3.探索肌肉细胞DOT1L-MPTP-Irisin通路中骨质疏松诊治的新靶点。通过以上研究，希望为骨质疏松的诊断和防治提供新的策略和理论支持。

肌肉系统与骨骼系统在解剖学上紧密连接，并且存在分子水平的互相调控。临床上骨质疏松患者往往伴随肌肉萎缩。本项目前期发现DOT1L通过控制线粒体通透性转换孔（Mitochondrial permeability transition pore, MPTP）的开放，调节肌肉细胞功能和肌细胞因子分泌，进而影响破骨细胞分化，为探索骨质疏松的发病机制提供了新的研究思路。利用生物信息学、分子生物学、细胞生物学手段，以及动物实验，本项目继续探讨其中的具体机制。结果显示，DOT1L结合STAT1，STAT1以非直接转录调节的方式调控CypD基因的表达，从而调控MPTP的开放。MPTP的开放进一步影响了ROS/Sirt1/PCG1-α通路激活。其中PCG1-α结合FNDC5基因转录区，直接调控FNDC5的表达以及Irisin的分泌。在卵巢摘除骨质疏松模型鼠中，我们发现Irisin能够显著减缓股骨和胫骨的骨量减少速度，缓解骨质疏松。这表明DOT1L调控的MPTP开放介导的ROS/Sirt1/PCG1-α通路可以作为治疗骨质疏松的新靶点。在实施本项目的过程中，利用收集的血清，我们发现血清细胞外囊泡（Extracellular vesicles, EVs）调节骨代谢平衡，其机制可能是通过细胞核蛋白Nucleolin介导的线粒体稳态进行调控。此外，利用蛋白质组学、免疫组织化学和体外细胞实验，本课题还探索骨代谢和阿尔兹海默病（Alzheimer's disease, AD）共有发病机制，发现DOT1L可能作为关键纽带，联系骨质疏松和AD。