炎症微环境中的细胞外基质（ECM）代谢紊乱在椎间盘退变（IDD）的发病机制中发挥着关键作用。据报道，白介素 32 (IL-32) 参与多种炎症性疾病的进展；然而，其是否参与髓核（NP）细胞的基质代谢尚不清楚。因此，本研究旨在探讨IL-32调节炎症微环境中ECM代谢的机制。 RNA-seq 用于识别炎症微环境中 NP 细胞中异常表达的基因。采用蛋白质印迹法、实时定量PCR、免疫组织化学和免疫荧光分析来测量经或未经肿瘤坏死因子-α（TNF-α）处理的人NP组织或NP细胞中IL-32和代谢标志物的表达。在体内，将过表达 IL-32 的腺相关病毒注射到大鼠尾椎间盘中，以评估其对 IDD 的影响。通过免疫沉淀和质谱法鉴定与 IL-32 相互作用的蛋白质。采用过表达IL-32或敲低脂肪非典型钙粘蛋白4（FAT4）的慢病毒、YAP相关蛋白（YAP）抑制剂维替泊芬（VP）处理人NP细胞，探讨IL-32的发病机制。 Hippo/YAP 信号传导活性已在人类 NP 组织中得到验证。如临床样本所示，退行性 NP 组织中 IL-32 的表达显着上调。此外，IL-32 在 TNF-α 诱导的退行性 NP 细胞中显着过度表达。 IL-32 过表达诱导大鼠模型中 IDD 进展。机制上，炎症微环境中IL-32的升高增强了其与FAT4和哺乳动物不育20样激酶1/2（MST1/2）蛋白的相互作用，促使MST1/2磷酸化，并激活Hippo/YAP信号通路，引起基质NP细胞代谢紊乱。我们的研究结果表明，IL-32通过FAT4/MST/YAP轴介导炎症微环境中NP细胞的基质代谢紊乱，为IDD的精准治疗提供理论基础。版权所有©2024。Elsevier B.V.出版。

Extracellular matrix (ECM) metabolism disorders in the inflammatory microenvironment play a key role in the pathogenesis of intervertebral disc degeneration (IDD). Interleukin-32 (IL-32) has been reported to be involved in the progression of various inflammatory diseases; however, it remains unclear whether it participates in the matrix metabolism of nucleus pulposus (NP) cells. Therefore, this study aimed to investigate the mechanism of IL-32 on regulating the ECM metabolism in the inflammatory microenvironment. RNA-seq was used to identify aberrantly expressed genes in NP cells in the inflammatory microenvironment. Western blotting, real-time quantitative PCR, immunohistochemistry and immunofluorescence analysis were performed to measure the expression of IL-32 and metabolic markers in human NP tissues or NP cells treated with or without tumor necrosis factor-α (TNF-α). In vivo, an adeno-associated virus overexpressing IL-32 was injected into the caudal intervertebral discs of rats to assess its effect on IDD. Proteins interacting with IL-32 were identified via immunoprecipitation and mass spectrometry. Lentivirus overexpressing IL-32 or knocking down Fat atypical cadherin 4 (FAT4), yes-associated protein (YAP) inhibitor-Verteporfin (VP) were used to treat human NP cells, to explore the pathogenesis of IL-32. Hippo/YAP signaling activity was verified in human NP tissues. IL-32 expression was significantly upregulated in degenerative NP tissues, as indicated in the clinical samples. Furthermore, IL-32 was remarkably overexpressed in TNF-α-induced degenerative NP cells. IL-32 overexpression induced IDD progression in the rat model. Mechanistically, the elevation of IL-32 in the inflammatory microenvironment enhanced its interactions with FAT4 and mammalian sterile 20-like kinase1/2 (MST1/2) proteins, prompting MST1/2 phosphorylation, and activating the Hippo/YAP signaling pathway, causing matrix metabolism disorder in NP cells. Our results suggest that IL-32 mediates matrix metabolism disorders in NP cells in the inflammatory micro-environment via the FAT4/MST/YAP axis, providing a theoretical basis for the precise treatment of IDD.Copyright © 2024. Published by Elsevier B.V.