颞下颌骨关节炎中外泌体miRNA介导的滑膜对软骨功能调控的机制研究

Pathological changes occur in synovium and cartilage, when temporomandibular osteoarthritis occurs. The interaction and regulation between different tissues are still unclear in osteoarthritis. The content of exosome mainly are miRNA . The existence of exosomes and overexpression of proinflammatory molecule miRNA21 was conformed in our former research. This project based on previous results is designed to explore whether fibroblast secreted exosomal miRNA21 regulates the function of chondrocyte. Firstly miRNA sequencing, synovial fluid collection and animal model are used to verify the inflammatory induction of synovial cells on chondrocytes . Secondly, siRNA transfection, double fluorescence test and chondrocyte sequencing are used to prove exosome miRNA21 secreted by fibroblast regulates function of chondrocyte by repressing expression of TIMP-3 and Sox-5. The result could expound pathologic mechanism of TMJ osteoarthritis and provide theoretical basis for clinical treatment of osteoarthritis.

颞下颌骨关节炎发生时，关节腔内滑膜和软骨均会出现明显的炎性改变。滑膜和软骨之间是否通过信息交流来促进炎症的进展目前尚不清楚。外泌体是细胞之间信息交流的重要载体，主要成分包括miRNA。项目组前期证实了滑液中存在外泌体及高含量的促炎分子miRNA21，现将进一步探讨滑膜细胞释放的外泌体miRNA21是否对软骨细胞进行炎性调控。本项目通过患者滑液收集、体外细胞试验、miRNA基因测序和动物模型的构建来验证滑膜外泌体对软骨细胞的炎性诱导机制；再通过siRNA转染、双荧光试验和软骨细胞基因测序，验证骨关节炎中滑膜细胞来源的外泌体miRNA21通过抑制软骨细胞内TIMP-3和Sox-5基因表达介导软骨内的炎性发生。该课题旨在阐述滑膜细胞在受到炎性刺激后释放外泌体对软骨细胞进行炎性调控，从而促进骨关节炎进展的病理机制，为今后临床治疗提供前期理论基础。

颞下颌关节炎是一种以炎症及软骨退化为特征的常见病，而促炎性M1型巨噬细胞在颞下颌关节（TMJ）炎症中起了重要作用。本实验旨在研究源自M1型巨噬细胞的细胞外囊泡（M1-EVs）对颞下颌关节是否具有促炎作用以及相关的机制。本研究在体外将人的单核细胞系（THP-1）来源的M1-EVs应用于人的髁突软骨细胞；将源自大鼠骨髓的巨噬细胞（BMDM）的M1-EVs注射到大鼠颞下颌关节用于体外实验；体内体外实验均发现IL-6、IL-8、IL-β和基质金属蛋白酶的水平上调。其次还采用了miRNA测序确定了和M0-EVs 相比M1-EVs中表达差异最大的miRNA为miR-1246;并且RT-PCR结果也显示了miR-1246在TMJ骨关节炎和滑膜炎患者滑液中的高表达。此外再通过TargetScan，miRTarBase和miRDB筛选了其靶基因，即GSK-3β和Axin2；研究的结果显示，miR-1246抑制了GSK-3β和Axin2的表达，并激活了 Wnt /β-catenin通路从而引起了髁突软骨细胞的炎症。总之，本研究表明，含有miR-1246 的M1-EVs可以促进髁突软骨细胞的炎症，并为颞下颌关节炎症的发病机理提供了新的见解