长链非编码RNA NR_122076对RA成纤维样滑膜细胞迁移和侵袭的调控及其机制研究

Abnormal migration and invasion of fibroblast like synoviocytes (FLS) play a key role in controlling the progress of synovial inflammation and joint destruction of rheumatoid arthritis (RA). In recent years, it has been shown that long noncoding RNA (lncRNA) is involved in the modulation of autoimmune response and growth and invasion in several kinds of cancer. We found that expression of lncRNA NR_122076 is significantly decreased in FLS from RA patients compared with OA patients and healthy controls. lncRNA NR_122076 is mainly located in cytoplasm. Overexpression of NR_122076 by lentivirus infection suppressed migration and invasion of RA FLS. Therefore, based on our previous studies, this proposal will be aimed at identifying the role of NR_122076 in regulating migration and invasion of RA FLS. We further search for the "molecular target" that interacts with NR_122076 and regulates FLS function by using bioinformatics analysis, RNA pulldown and RNA immunoprecipitation (IP). We propose to clarify the mechanisms underlying synovial inflammation and joint destruction of RA in the term of epigenetics, providing the scientific basis for establishment of a novel therapeutic target for RA.

成纤维样滑膜细胞（FLS）异常迁移和侵袭是类风湿关节炎（RA）滑膜病变和关节破坏持续进展的关键因素。长链非编码RNA（lncRNA）可能在自身免疫免疫反应以及肿瘤生长和侵袭中发挥重要作用。我们前期通过基因芯片筛选以及Q-PCR验证，发现lncRNA NR_122076 在RA患者FLS中表达显著低于正常人和骨关节炎患者，并表达于细胞质中；过表达lncRNA NR_122076 明显抑制FLS体外迁移和侵袭。本课题拟在现有工作基础上，进一步明确 NR_122076对RA FLS迁移、侵袭等功能的调控作用，并利用生物信息学、RNA pulldown、RNA免疫沉淀等方法“捕获”能与NR_122076相互结合并能调节FLS功能的“目标分子”，以阐明NR_122076调控RA FLS功能的分子机制。本项目将进一步揭示RA滑膜病变和关节破坏的发生机制，并为RA治疗提供新思路。

滑膜组织的增殖和侵袭作用是类风湿关节炎（RA）发生关节破坏的关键因素，成纤维样滑膜细胞（fibroblast like synoviocytes, FLS）在RA滑膜组织发挥侵袭作用的过程中扮演关键角色。长链非编码RNA（long noncoding RNＡ，lncRNA）是一类长度大于200 nt的非编码RNA，其在RA等自身免疫疾病的作用远未得到阐明。本课题运用分子生物学和现代免疫学技术研究发现，lncRNA NR_122076 （或AK309896）在RA FLS和滑膜组织中表达下调，我们将其命名为LERFS（Lowly Expressed in Rheumatoid Fibroblast-like Synoviocyte）。过表达LERFS抑制RA FLS的体外迁移、侵袭和增殖，使细胞周期阻滞于G2/M期，并抑制RA FLS的体内迁移、侵袭。进一步利用RNA pulldown和质谱分析等发现，在正常人的 FLS中，LERFS与hnRNP Q蛋白结合形成复合体，进而促进hnRNP Q与RhoA、Rac1以及CDC42 mRNA之间的结合，降低上述mRNA的稳定性或翻译过程，从而使相应蛋白质保持在较低水平；而在RA FLS中LERFS表达水平下调导致LERFS-hnRNP Q复合体形成减少，对RhoA、Rac1和CDC42的mRNA抑制作用减弱，使上述Rho家族蛋白表达增高，活性增强，从而导致RA FLS增殖和侵袭能力增强。上述研究结果提示滑膜中LERFS表达下调可能是RA发生关节破坏的重要因素，揭示了FLS参与RA发病的新机制，为RA关节破坏的治疗提供新的靶点。