Rheumatoid arthritis (RA) is a chronic inflammatory disease, featured by erosive arthritis, which will eventually lead to deprivation normal functions of the joint and joint malformations. Continued illness also results in more serious complications, such as cardiovascular diseases and disability. Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) function in various conditions, including RA. LncRNA NEAT1 was reported to promote migration and invasion in RA-FLSs, functioning as a promising diagnostic and therapeutic indicator in RA. The present work focused on the role of lncRNA NEAT1 in RA and the related mechanism. We collected the synovial tissue samples of 30 RA patients and 20 healthy controls. Moreover, RA fibroblast-like synoviocytes (RA-FLSs) cell line was bought and treated with tumor necrosis factor-α (TNF-α) to establish in vitro model of RA. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the expression of NEAT1 in synovial tissue and RA-FLSs. NEAT1 silencing plasmid were synthesized and co-trasnfected with miR-204-5p inhibitor into RA-FLSs. MTT and 5-Ethynyl-2′-deoxyuridine staining were used to assess cell proliferation. Flow cytometry and TUNEL assay were used to determine the cell apoptosis. miR-204-5p has been predicted as a target miRNA of NEAT1, and the interaction between NEAT1 and miR-204-5p was verified by dual-luciferase assay and RNA pull-down assay. qRT-PCR and enzyme-linked immunosorbent assay were used to determine the mRNA and protein concentration of interleukin‐1β and interleukin‐6. Finally, western blot assay was applied to measure the effect of NEAT1 and on p53, NF-κB, and p-NF-κB expressions. We found that NEAT1 was up-regulated, and miR-204-5p was down-regulated in the RA patients’ synovial tissue and TNF-α treated RA-FLSs. TNF-α increased NEAT1 level and decreased miR-204-5p level in RA-FLSs. There was no significant variance of p53 after transfected with NEAT1 in RA-FLSs. Meanwhile, Knockdown of NEAT1 attenuated TNF-α-induced RA-FLSs cell proliferation and inflammatory cytokine production while promoted cell apoptosis by targeting miR-204-5p through NF-κB pathway. These findings indicated that NEAT1 may be developed as a potential target for patients with RA.

类风湿性关节炎（RA）是一种慢性炎症性疾病，以侵蚀性关节炎为特征，最终会导致关节的正常功能丧失和关节畸形。持续患病还会导致更严重的并发症，例如心血管疾病和残疾。长链非编码 RNA (lncRNA) 和微 RNA (miRNA) 在各种条件下发挥作用，包括 RA。据报道，LncRNA NEAT1 可促进 RA-FLS 的迁移和侵袭，在 RA 中作为一种有前途的诊断和治疗指标。目前的工作重点是 lncRNA NEAT1 在 RA 中的作用及其相关机制。我们收集了 30 名 RA 患者和 20 名健康对照的滑膜组织样本。而且，购买RA成纤维样滑膜细胞（RA-FLSs）细胞系，用肿瘤坏死因子-α（TNF-α）处理，建立RA体外模型。定量实时聚合酶链反应 (qRT-PCR) 用于确定 NEAT1 在滑膜组织和 RA-FLS 中的表达。合成 NEAT1 沉默质粒并与 miR-204-5p 抑制剂共转染到 RA-FLS 中。MTT 和 5-Ethynyl-2'-deoxyuridine 染色用于评估细胞增殖。流式细胞术和TUNEL法用于测定细胞凋亡。miR-204-5p 已被预测为 NEAT1 的目标 miRNA，NEAT1 和 miR-204-5p 之间的相互作用通过双荧光素酶测定和 RNA 下拉测定验证。qRT-PCR 和酶联免疫吸附试验用于测定白细胞介素 1β 和白细胞介素 6 的 mRNA 和蛋白质浓度。最后，采用蛋白质印迹法检测 NEAT1 对 p53、NF-κB 和 p-NF-κB 表达的影响。我们发现在 RA 患者的滑膜组织和 TNF-α 治疗的 RA-FLS 中，NEAT1 上调，miR-204-5p 下调。TNF-α 增加了 NEAT1 水平并降低了 RA-FLS 中的 miR-204-5p 水平。在RA-FLSs中转染NEAT1后p53没有显着变化。同时，敲除 NEAT1 减弱了 TNF-α 诱导的 RA-FLSs 细胞增殖和炎性细胞因子的产生，同时通过 NF-κB 通路靶向 miR-204-5p 促进细胞凋亡。这些发现表明 NEAT1 可能被开发为 RA 患者的潜在靶点。和 miR-204-5p 在 RA 患者的滑膜组织和 TNF-α 治疗的 RA-FLS 中下调。TNF-α 增加了 NEAT1 水平并降低了 RA-FLS 中的 miR-204-5p 水平。在RA-FLSs中转染NEAT1后p53没有显着变化。同时，敲除 NEAT1 减弱了 TNF-α 诱导的 RA-FLSs 细胞增殖和炎性细胞因子的产生，同时通过 NF-κB 通路靶向 miR-204-5p 促进细胞凋亡。这些发现表明 NEAT1 可能被开发为 RA 患者的潜在靶点。和 miR-204-5p 在 RA 患者的滑膜组织和 TNF-α 治疗的 RA-FLS 中下调。TNF-α 增加了 NEAT1 水平并降低了 RA-FLS 中的 miR-204-5p 水平。在RA-FLSs中转染NEAT1后p53没有显着变化。同时，敲除 NEAT1 减弱了 TNF-α 诱导的 RA-FLSs 细胞增殖和炎性细胞因子的产生，同时通过 NF-κB 通路靶向 miR-204-5p 促进细胞凋亡。这些发现表明 NEAT1 可能被开发为 RA 患者的潜在靶点。NEAT1 的敲低减弱了 TNF-α 诱导的 RA-FLSs 细胞增殖和炎性细胞因子的产生，同时通过 NF-κB 通路靶向 miR-204-5p 促进细胞凋亡。这些发现表明 NEAT1 可能被开发为 RA 患者的潜在靶点。NEAT1 的敲低减弱了 TNF-α 诱导的 RA-FLSs 细胞增殖和炎性细胞因子的产生，同时通过 NF-κB 通路靶向 miR-204-5p 促进细胞凋亡。这些发现表明 NEAT1 可能被开发为 RA 患者的潜在靶点。