Lee Angela, Qiao Yu, Grigoriev Galina, Chen Janice, Park-Min Kyung-Hyun, Park Sung Ho, Ivashkiv Lionel B, Kalliolias George D
Hospital for Special Surgery and Weill Cornell Graduate School of Medical Sciences, New York, New York 10021, USA.
Arthritis Rheum. 2013 Apr;65(4):928-38. doi: 10.1002/art.37853.
The nonresolving character of synovial inflammation in rheumatoid arthritis (RA) is a conundrum. To identify the contribution of fibroblast-like synoviocytes (FLS) to the perpetuation of synovitis, we investigated the molecular mechanisms that govern the tumor necrosis factor α (TNFα)-driven inflammatory program in human FLS.
FLS obtained from the synovial tissues of patients with RA or osteoarthritis were stimulated with TNFα and assayed for gene expression and cytokine production by real-time quantitative reverse transcription-polymerase chain reaction analysis and enzyme-linked immunosorbent assay. NF-κB signaling was evaluated by Western blotting. Histone acetylation, chromatin accessibility, and NF-κB p65 and RNA polymerase II (Pol II) occupancy at the interleukin-6 (IL-6) promoter were measured by chromatin immunoprecipitation and restriction enzyme accessibility assays.
In FLS, TNFα induced prolonged transcription of messenger RNA (mRNA) for IL-6 and progressive accumulation of IL-6 protein over 4 days. Similarly, induction of mRNA for CXCL8/IL-8, CCL5/RANTES, matrix metalloproteinase 1 (MMP-1), and MMP-3 after TNFα stimulation was sustained for several days. This contrasted with the macrophage response to TNFα, which characteristically involved a transient increase in the expression of proinflammatory genes. In FLS, TNFα induced prolonged activation of NF-κB signaling and sustained transcriptional activity, as indicated by increased histone acetylation, chromatin accessibility, and p65 and Pol II occupancy at the IL-6 promoter. Furthermore, FLS expressed low levels of the feedback inhibitors A20-binding inhibitor of NF-κB activation 3 (ABIN-3), IL-1 receptor-associated kinase M (IRAK-M), suppressor of cytokine signaling 3 (SOCS-3), and activating transcription factor 3 (ATF-3), which terminate inflammatory responses in macrophages.
TNFα signaling is not effectively terminated in FLS, which leads to an uncontrolled inflammatory response. The results suggest that prolonged and sustained inflammatory responses by FLS in response to synovial TNFα contribute to the persistence of synovial inflammation in RA.
类风湿关节炎（RA）中滑膜炎症的不消退特性是一个难题。为了确定成纤维样滑膜细胞（FLS）对滑膜炎持续存在的作用，我们研究了调控人FLS中肿瘤坏死因子α（TNFα）驱动的炎症程序的分子机制。
用TNFα刺激从RA患者或骨关节炎患者滑膜组织中获取的FLS，并通过实时定量逆转录-聚合酶链反应分析和酶联免疫吸附测定法检测基因表达和细胞因子产生情况。通过蛋白质免疫印迹法评估核因子κB（NF-κB）信号传导。通过染色质免疫沉淀和限制性酶切可及性测定法测量组蛋白乙酰化、染色质可及性以及NF-κB p65和RNA聚合酶II（Pol II）在白细胞介素-6（IL-6）启动子上的占据情况。
在FLS中，TNFα诱导IL-6信使核糖核酸（mRNA）的转录延长，并在4天内导致IL-6蛋白逐渐积累。同样，TNFα刺激后CXCL8/IL-8、CCL5/趋化因子调节激活正常T细胞表达和分泌因子（RANTES）、基质金属蛋白酶1（MMP-1）和MMP-3的mRNA诱导持续数天。这与巨噬细胞对TNFα的反应形成对比，巨噬细胞对TNFα的反应典型地表现为促炎基因表达的短暂增加。在FLS中，TNFα诱导NF-κB信号传导的长期激活和持续的转录活性，这表现为组蛋白乙酰化增加、染色质可及性增加以及p65和Pol II在IL-6启动子上的占据增加。此外，FLS表达低水平的反馈抑制剂NF-κB激活3的A20结合抑制剂（ABIN-3）、白细胞介素-1受体相关激酶M（IRAK-M）、细胞因子信号传导抑制因子3（SOCS-3）和激活转录因子3（ATF-3），这些因子可终止巨噬细胞中的炎症反应。
TNFα信号在FLS中不能有效终止，这导致炎症反应不受控制。结果表明，FLS对滑膜TNFα的延长和持续的炎症反应促成了RA中滑膜炎症的持续存在。
