膜型/可溶型CD305分子调控FLS参与类风湿性关节炎骨损伤的机制研究

Rheumatoid arthritis (RA) is a chronic, systemic inflammatory disorder that may affect many tissues and organs, but principally attacks synovial joints. Fibroblast-like synoviocytes (FLS) are among the principal effector cells in the pathogenesis of RA. CD305，also named Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1), is a form of immunoreceptor tyrosine-based inhibition motifs (ITIMs)-harboring receptors primarily expressed in immune cells. In the previous studies, we found that CD305 moleclue highly expressed on the FLS of RA patients. Interestingly, when we focused on the expression of CD305 on the FLS, we found that the membrane CD305 actually was a low level expression in the RA patients. At same time, ELISA analysis observed that the soluble CD305 was significantly higher in the synovial fluids of RA patients compared with the osteoarthritis(OA) patients, and endocytosis was also observed by immunocytochemistry. All of these data suggested that CD305 moleclue may participate in the pathological process of RA by switch the membrane/soluble form. Based on these observations, it is worth to investigate the relationship between the level of membrane/soluble CD305 and the RA disease activity; the effect of membrane CD305 on the proliferation, erosiveness and inflammatory cytokines production of FLS from RA patients; the mechanisms of CD305 molecule been sheded or endocytosed. Furthermore, by using the CD305 transgenic mice we previously constructed, we intend to verify the role of this molecule in the FLS-induced cartilage injury in an in vivo mouse model. In particular, CD305 could be used as a novel therapeutic target for inflammatory arthritis diseases.

类风湿关节炎（RA）是一种以滑膜炎症为病理中心的自身免疫性疾病，关节的软骨侵蚀是RA的主要特点，成纤维样滑膜细胞（FLS）在其中扮演关键角色。CD305分子是一种在多种免疫细胞上起重要作用的抑制性受体，通过前期的研究我们发现CD305高表达于RA患者滑膜FLS中。值得注意的是，膜型CD305在RA FLS表面水平并不高，而RA患者关节液中可溶型CD305水平显著升高，同时伴CD305内吞现象，提示CD305分子通过调节膜型/可溶型水平参与RA病理过程。因此，本课题旨在研究膜型/可溶型CD305水平与RA疾病活动度之间的联系、探讨膜型CD305与FLS增殖、侵蚀、炎症因子分泌等关系、以及RA状态下CD305被剪切或内吞的调节机制，并利用前期已构建的CD305转基因小鼠进一步验证CD305分子与RA FLS过度增殖活化以及软骨损伤的关系，为RA的治疗提供新的理论依据。

本研究旨在探讨免疫抑制性分子，白细胞相关免疫球蛋白样受体LAIR-1/CD305在类风湿关节炎（RA）患者滑膜成纤维样细胞（FLS）中的表达。研究该分子对炎症因子、基质金属蛋白酶（MMPs）和FLS侵袭性的作用。结果发现，LAIR-1/CD305高表达于类风湿关节炎患者滑膜组织中，免疫荧光染色发现波形蛋白/ CD305蛋白具有共定位的格局。采用流式细胞仪检测，证实在原代分离的RA FLS中，LAIR-1/CD305膜型表达水平低，而在细胞内有表达。结合我们以往的结果，即RA患者关节滑膜液中可溶型LAIR-1/CD305显著升高，提示在RA炎症微环境的刺激下，膜型LAIR-1/CD305可被削切，导致FLS LAIR-1/CD305水平降低，影响正常的功能。本研究发现，肿瘤坏死因子（TNF）α是导致RA FLS膜型LAIR-1/CD305被削切的主要原因，而此过程与丝氨酸蛋白酶的活性直接相关。进一步的实验证实，过表达CD305/LAIR-1降低FLS的侵袭，抑制TNF-α诱导的MMP-13，炎症因子IL-6和IL-8的表达。这些研究结果提示LAIR-1/CD305是RA疾病过程中的一个重要的调控因子，参与RA疾病的发生发展过程。