miR-106b-5p启动子甲基化状态介导MAPK/ERK信号通路调控腰椎间盘退变机制的研究

Lumbar disc degeneration (LDD) is one of the most common spinal disorders. LDD poses considerable financial burden to many households and society. Several studies showed that microRNAs play a key role in development of LDD as post-transcriptional regulators of target genes. In prior study, we found that miR-106b-5p was significant up-regulated in human degenerated nucleus pulposus. Bioinformatics analysis and validation study confirmed that MAPK1 is a potential target gene of miR-106b-5p, and miR-106b-5p could regulate the proliferation and apoptosis of nucleus pulposus cell via MAPK/ERK signal pathway. As two CpG islands in miR-106b-5p promoter region, we propose a hypothesis that miR-106b-5p may be involved in the development of LDD due to promoter region methylation modification. In this study, we will measure the methylation status of miR-106b-5p promoter region using MeDIP-Seq technique in human nucleus pulposus, and explore the association between methylation modification levels and miR-106b-5p expression, and evaluate the regulation of miR-106b-5p on proliferation and apoptosis of nucleus pulposus cell via MAPK/ERK signal pathway. Our results may contribute to developing our understanding of pathogenesis of LDD and provide an important reference for early prevention and treatment of LDD.

腰椎间盘退变（lumbar disc degeneration，LDD）是最常见的脊柱退行性疾病之一，给家庭及社会造成巨大的经济负担。课题组前期研究发现miR-106b-5p在人LDD髓核组织中表达显著升高，生物信息学分析及靶标验证证实MAPK1是miR-106b-5p的靶基因，高表达的miR-106b-5p可能通过MAPK/ERK信号通路调控髓核细胞的凋亡和增殖。由于miR-106b-5p启动子区域存在CpG岛，我们推测miR-106b-5p可能存在异常甲基化状态而呈高表达，进而影响LDD的发生与发展。本研究在前期研究基础上，通过MeDIP-Seq技术检测人LDD髓核组织中miR-106b-5p启动子甲基化水平；探讨miR-106b-5p甲基化水平与表达量之间的关系，以及靶向调控MAPK/ERK信号通路影响髓核细胞的凋亡和增殖，为LDD防治提供新思路。

MicroRNA在腰椎间盘退变（lumbar disc degeneration，LDD）发病机制中的作用日益受到关注。本项目发现miR-106b-5p在LDD患者的髓核组织和细胞中表达升高；过表达或沉默miR-106b-5p显著影响髓核细胞的增殖、凋亡及细胞外基质的合成；体内沉默miR-106b-5p可抑制椎间盘的退变进程；重要的是，miR-106b-5p靶向SIRT2调控MAPK/ERK信号通路参与LDD的病理过程。体内治疗实验显示miR-106b-5p的表达与其启动子区域中基化状态存在直接的相关性，且过表达miR-106b-5p可导致髓核细胞的凋亡增高、细胞外基质降解增多，促进LDD的发生发展，而沉默miR-106b-5p可逆转上述作用。综上所述，研究结果表明miR-106b-5p同时在体外与体内发挥重要的调控作用，可作为潜在治疗LDD的靶点。