Department of Bioscience and Biotechnology, Konkuk University, Seoul, Republic of Korea.
College of Pharmacy & Medical Research Center, Chungbuk National University, Cheongju, Republic of Korea.
Front Immunol. 2023 May 9;14:1160301. doi: 10.3389/fimmu.2023.1160301. eCollection 2023.
Interleukin-32 (IL-32), first reported in 2005, and its isoforms have been the subject of numerous studies investigating their functions in virus infection, cancer, and inflammation. IL-32θ, one of the IL-32 isoforms, has been shown to modulate cancer development and inflammatory responses. A recent study identified an IL-32θ mutant with a cytosine to thymine replacement at position 281 in breast cancer tissues. It means that alanine was also replaced to valine at position 94 in amino acid sequence (A94V). In this study, we investigated the cell surface receptors of IL-32θA94V and evaluated their effect on human umbilical vein endothelial cells (HUVECs). Recombinant human IL-32θA94V was expressed, isolated, and purified using Ni-NTA and IL-32 mAb (KU32-52)-coupled agarose columns. We observed that IL-32θA94V could bind to the integrins αVβ3 and αVβ6, suggesting that integrins act as cell surface receptors for IL-32θA94V. IL-32θA94V significantly attenuated monocyte-endothelial adhesion by inhibiting the expression of Intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in tumor necrosis factor (TNF)-α-stimulated HUVECs. IL-32θA94V also reduced the TNF-α-induced phosphorylation of protein kinase B (AKT) and c-jun N-terminal kinases (JNK) by inhibiting phosphorylation of focal adhesion kinase (FAK). Additionally, IL-32θA94V regulated the nuclear translocation of nuclear factor kappa B (NF-κB) and activator protein 1 (AP-1), which are involved in ICAM-1 and VCAM-1 expression. Monocyte-endothelial adhesion mediated by ICAM-1 and VCAM-1 is an important early step in atherosclerosis, which is a major cause of cardiovascular disease. Our findings suggest that IL-32θA94V binds to the cell surface receptors, integrins αVβ3 and αVβ6, and attenuates monocyte-endothelial adhesion by suppressing the expression of ICAM-1 and VCAM-1 in TNF-α-stimulated HUVECs. These results demonstrate that IL-32θA94V can act as an anti-inflammatory cytokine in a chronic inflammatory disease such as atherosclerosis.
白细胞介素-32（IL-32）于 2005 年首次报道，其同种型一直是研究其在病毒感染、癌症和炎症中功能的众多研究的主题。IL-32θ 是 IL-32 的同种型之一，已被证明可调节癌症的发展和炎症反应。最近的一项研究在乳腺癌组织中发现了一个 IL-32θ 突变体，该突变体在第 281 位的胞嘧啶被胸腺嘧啶取代。这意味着在氨基酸序列中第 94 位的丙氨酸也被缬氨酸取代（A94V）。在这项研究中，我们研究了 IL-32θA94V 的细胞表面受体，并评估了它们对人脐静脉内皮细胞（HUVECs）的影响。使用 Ni-NTA 和 IL-32 mAb（KU32-52）-偶联琼脂糖柱表达、分离和纯化重组人 IL-32θA94V。我们观察到 IL-32θA94V 可以与整合素 αVβ3 和 αVβ6 结合，这表明整合素是 IL-32θA94V 的细胞表面受体。IL-32θA94V 通过抑制肿瘤坏死因子（TNF）-α 刺激的 HUVECs 中细胞间黏附分子-1（ICAM-1）和血管细胞黏附分子-1（VCAM-1）的表达，显著减弱单核细胞-内皮细胞的黏附。IL-32θA94V 还通过抑制粘着斑激酶（FAK）的磷酸化来减少 TNF-α 诱导的蛋白激酶 B（AKT）和 c-jun N 末端激酶（JNK）的磷酸化。此外，IL-32θA94V 通过调节核因子 kappa B（NF-κB）和激活蛋白 1（AP-1）的核转位来调节 ICAM-1 和 VCAM-1 的表达，这些因子参与 ICAM-1 和 VCAM-1 的表达。ICAM-1 和 VCAM-1 介导的单核细胞-内皮细胞黏附是动脉粥样硬化的一个重要早期步骤，动脉粥样硬化是心血管疾病的主要原因。我们的研究结果表明，IL-32θA94V 与细胞表面受体整合素 αVβ3 和 αVβ6 结合，并通过抑制 TNF-α 刺激的 HUVECs 中 ICAM-1 和 VCAM-1 的表达来减弱单核细胞-内皮细胞的黏附。这些结果表明，IL-32θA94V 可以作为一种抗炎细胞因子在动脉粥样硬化等慢性炎症性疾病中发挥作用。
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