Department of Immunology, School of Medicine, Kermanshah University of Medical Sciences, Kermanshah, Iran.
Rheumatology Research Center, Tehran University of Medical Sciences, Tehran, Iran.
Inflammation. 2022 Apr;45(2):665-676. doi: 10.1007/s10753-021-01575-z. Epub 2022 Feb 3.
The inflammatory interleukin (IL)-23/IL-17 axis plays an important role in the pathogenesis of ankylosing spondylitis (AS), but with an unknown regulatory mechanism. This study aimed to investigate the role of endoplasmic reticulum (ER) stress and autophagy pathway in the expression of IL-23 in peripheral blood-derived macrophages in AS patients. Peripheral blood samples were obtained from 15 AS and 15 healthy control subjects. MACS was used to isolate monocytes from PBMCs. Then, M-CSF was used to differentiate monocytes to M2 macrophages. IFN-γ and/or LPS were used to activate macrophages and M2 polarization towards M1 macrophages. Thapsigargin was used to induce ER stress and 3-MA to inhibit autophagy. The purity of extracted monocytes and macrophage markers was evaluated by flow cytometry. mRNA expression of HLA-B and-B27, ER stress-related genes, autophagy-related genes, and IL-23p19 was performed using RT-qPCR. Soluble levels of IL-23p19 were measured using ELISA. Significant increase in mRNA expression of HLA-B, HLA-B27, BiP, XBP1, CHOP, and PERK mRNAs was observed in macrophages of AS patients before and after stimulation with IFN-γ and LPS. No significant change in autophagy gene expression was detected. mRNA and soluble levels of IL-23p19 demonstrated a significant increase in macrophages of AS patients compared to healthy subjects. ER stress induction led to a significant increase in IL-23p19 in macrophages. Inhibition of autophagy did not affect IL-23 expression. ER stress, unlike autophagy, is associated with increased IL-23 levels in macrophages of AS patients.Key Messages ER stress in macrophages from AS patients plays a role in the increased production of IL-23. The autophagy pathway is not involved in the modulation of IL-23 production by AS macrophages.
炎症性白细胞介素 (IL)-23/IL-17 轴在强直性脊柱炎 (AS) 的发病机制中起重要作用，但调控机制尚不清楚。本研究旨在探讨内质网 (ER) 应激和自噬通路在 AS 患者外周血源性巨噬细胞中 IL-23 表达中的作用。从 15 例 AS 患者和 15 名健康对照者中获得外周血样本。使用 MACS 从 PBMC 中分离单核细胞。然后，使用 M-CSF 将单核细胞分化为 M2 巨噬细胞。使用 IFN-γ 和/或 LPS 激活巨噬细胞并使其向 M1 巨噬细胞极化。使用 Thapsigargin 诱导 ER 应激，使用 3-MA 抑制自噬。通过流式细胞术评估提取单核细胞和巨噬细胞标志物的纯度。使用 RT-qPCR 检测 HLA-B 和-B27、ER 应激相关基因、自噬相关基因和 IL-23p19 的 mRNA 表达。使用 ELISA 测量 IL-23p19 的可溶性水平。在 IFN-γ 和 LPS 刺激前后，AS 患者的巨噬细胞中观察到 HLA-B、HLA-B27、BiP、XBP1、CHOP 和 PERK mRNA 的 mRNA 表达显著增加。未检测到自噬基因表达的显著变化。与健康受试者相比，AS 患者的巨噬细胞中 IL-23p19 的 mRNA 和可溶性水平均显著增加。ER 应激诱导导致巨噬细胞中 IL-23p19 显著增加。自噬抑制不影响 IL-23 的表达。与自噬不同，ER 应激与 AS 患者巨噬细胞中 IL-23 水平的升高有关。
关键信息 AS 患者的巨噬细胞中的 ER 应激在增加 IL-23 的产生中起作用。自噬途径不参与调节 AS 巨噬细胞中 IL-23 的产生。
