Department of Orthopedic Surgery, Sun Yat-sen Memorial Hospital of Sun Yat-sen University, Guangzhou, Guangdong, China; Department of Orthopedic Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China.
Department of Orthopedic Surgery, Sun Yat-sen Memorial Hospital of Sun Yat-sen University, Guangzhou, Guangdong, China; Department of Orthopedic Surgery, The Affiliated Qingyuan Hospital (Qingyuan People's Hospital), Guangzhou Medical Universit, Qingyuan, Guangdong, China.
Int Immunopharmacol. 2024 Nov 15;141:112966. doi: 10.1016/j.intimp.2024.112966. Epub 2024 Aug 22.
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.
细胞外基质（ECM）代谢紊乱在炎症微环境中在椎间盘退行性变（IDD）的发病机制中起关键作用。白细胞介素 32（IL-32）已被报道参与各种炎症性疾病的进展；然而，它是否参与核髓核（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。使用 RNA-seq 鉴定炎症微环境中 NP 细胞中异常表达的基因。通过 Western blot、实时定量 PCR、免疫组织化学和免疫荧光分析测量人 NP 组织或经肿瘤坏死因子-α（TNF-α）处理的 NP 细胞中 IL-32 和代谢标志物的表达。在体内，将过表达 IL-32 的腺相关病毒注入大鼠尾椎间盘中，以评估其对 IDD 的影响。通过免疫沉淀和质谱鉴定与 IL-32 相互作用的蛋白质。使用过表达 IL-32 的慢病毒或敲低 Fat atypical cadherin 4 (FAT4)、yes-associated protein (YAP)抑制剂-Verteporfin (VP)处理人 NP 细胞，以探讨 IL-32 的发病机制。验证人 NP 组织中的 Hippo/YAP 信号活性。IL-32 在退行性 NP 组织中的表达明显上调，在临床样本中得到证实。此外，TNF-α诱导的退行性 NP 细胞中 IL-32 表达显著上调。IL-32 过表达在大鼠模型中诱导 IDD 进展。在机制上，炎症微环境中 IL-32 的升高增强了它与 FAT4 和哺乳动物无菌 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，导致 NP 细胞的基质代谢紊乱。我们的结果表明，IL-32 通过 FAT4/MST/YAP 轴介导 NP 细胞在炎症微环境中的基质代谢紊乱，为 IDD 的精确治疗提供了理论依据。
