Objective: To discuss how IRF9 affects the fibroblast-like synoviocytes (FLS) in TNF-induced rheumatoid arthritis (RA) via the SIRT-1/NF-κB signaling pathway. Methods: RA-FLS were isolated and divided into control, sh-IRF9, TNF, TNF + sh-Ctrl, TNF + sh-IRF9, TNF + sh-SIRT1, and TNF + sh-IRF9 + sh-SIRT1 groups. Biological features of FLS were evaluated by MTT, wound healing, and Transwell assays, respectively. Cell apoptosis and cycle were assessed flow cytometrically. Inflammatory cytokines were determined through enzyme-linked immunosorbent assay (ELISA), while IRF9 expression and SIRT1/NF-κB signaling pathway activity were measured by Western blotting. Results: TNF increased IRF9 expression as well as NF-κB signaling activity and down-regulated SIRT1 of RA-FLS. Silencing IRF9 resulted in up-regulation of SIRT1 and blocked NF-κB signaling, with significant decreases in TNF-induced cell viability, migration, and invasion, prominent enhancement in apoptosis and the proportion of cells in G0/G1 phase, but a decrease in the proportion of cells in S and G2/M phases, and reduced levels of inflammatory cytokines. However, these changes were totally abolished after silencing SIRT1, i.e., the IRF9 shRNA-induced inhibitory effect on the growth of RA-FLS was reversed. Conclusion: Silencing IRF9 curbs the activity of the NF-κB signaling pathway via up-regulating SIRT-1, to further suppress TNF-induced changes in the malignant features of RA-FLS, and the secretion of inflammatory cytokines, with the promoted apoptosis.

中文翻译：

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IRF9 通过调节 SIRT-1/NF-κB 信号通路影响类风湿性关节炎成纤维细胞样滑膜细胞的 TNF 诱导表型

目的：探讨 IRF9 如何通过 SIRT-1/NF-κB 信号通路影响 TNF 诱导的类风湿性关节炎（RA）中成纤维细胞样滑膜细胞（FLS）。方法：分离RA-FLS，分为对照、sh-IRF9、TNF、TNF+sh-Ctrl、TNF+sh-IRF9、TNF+sh-SIRT1、TNF+sh-IRF9+sh-SIRT1组。FLS 的生物学特征分别通过 MTT、伤口愈合和 Transwell 测定进行评估。通过流式细胞术评估细胞凋亡和周期。通过酶联免疫吸附试验 (ELISA) 测定炎症细胞因子，而通过蛋白质印迹法测量 IRF9 表达和 SIRT1/NF-κB 信号通路活性。结果：TNF 增加 IRF9 表达以及 NF-κB 信号活性并下调 RA-FLS 的 SIRT1。沉默 IRF9 导致 SIRT1 上调并阻断 NF-κB 信号，TNF 诱导的细胞活力、迁移和侵袭能力显着降低，细胞凋亡和 G0/G1 期细胞比例显着增强，但 S 期和 G2/M 期细胞比例下降，炎性细胞因子。然而，这些变化在沉默 SIRT1 后完全消失，即 IRF9 shRNA 诱导的对 RA-FLS 生长的抑制作用被逆转。结论：沉默IRF9通过上调SIRT-1抑制NF-κB信号通路的活性，进一步抑制TNF诱导的RA-FLS恶性特征的变化，抑制炎症细胞因子的分泌，促进细胞凋亡。但 S 期和 G2/M 期细胞比例减少，炎性细胞因子水平降低。然而，这些变化在沉默 SIRT1 后完全消失，即 IRF9 shRNA 诱导的对 RA-FLS 生长的抑制作用被逆转。结论：沉默IRF9通过上调SIRT-1抑制NF-κB信号通路的活性，进一步抑制TNF诱导的RA-FLS恶性特征的变化，抑制炎症细胞因子的分泌，促进细胞凋亡。但 S 期和 G2/M 期细胞比例减少，炎性细胞因子水平降低。然而，这些变化在沉默 SIRT1 后完全消失，即 IRF9 shRNA 诱导的对 RA-FLS 生长的抑制作用被逆转。结论：沉默IRF9通过上调SIRT-1抑制NF-κB信号通路的活性，进一步抑制TNF诱导的RA-FLS恶性特征的变化，抑制炎症细胞因子的分泌，促进细胞凋亡。