Lu Shan, Duan Mengyuan, Guo Zehao, Zhou Yongting, Wu Danwei, Zhang Xiaojuan, Wang Yicheng, Ye Caiying, Ju Rui, Li Juan, Zhang Dechang, Zhu Lei
Department of Pharmacology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, Beijing 100005, P.R. China.
Department of Pharmacy, Beijing Jishuitan Hospital, Beijing 100035, P.R. China.
Exp Ther Med. 2020 Aug;20(2):1455-1466. doi: 10.3892/etm.2020.8889. Epub 2020 Jun 12.
Carboxyamidotriazole (CAI), originally developed as a non-cytotoxic anti-cancer drug, was shown to have anti-inflammatory activity according to recent studies in a number of animal models of inflammation. However, its mechanism of action has not been characterized. Therefore, the present study was performed to identify the anti-inflammatory action of CAI in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages and to identify the signal transduction pathways involved. The results revealed that CAI had no direct effect on the activity of cyclooxygenase (COX), suggesting a different anti-inflammatory mechanism compared with that of COX-inhibiting non-steroidal anti-inflammatory drugs. Further investigation in RAW264.7 macrophages revealed that CAI decreased the production of nitric oxide via decreasing the LPS-stimulated expression of inducible nitric oxide synthase, and downregulated both mRNA and protein expression levels of the cytokines tumor necrosis factor-α, interleukin (IL)-1β, and IL-6. CAI also significantly reduced the increased DNA-binding activity of nuclear factor (NF)-κB induced by LPS stimulation. With respect to the mechanisms involved on NF-κB activity, CAI exhibited suppression of the phosphorylation and degradation of the inhibitor of nuclear factor-κBα (IκB), and decreased the phosphorylation levels of the p65 subunit and its subsequent nuclear translocation. In addition, CAI significantly decreased the phosphorylated forms of p38, JNK and ERK, which were increased following LPS stimulation, while the total expression levels of p38, JNK and ERK remained unaltered. The results in the present study indicate that CAI alleviates the inflammatory responses of RAW 264.7 macrophages in response to LPS stimulation via attenuating the activation of NF-κB and MAPK signaling pathways and decreasing the levels of pro-inflammatory mediators. This offers a novel perspective for understanding the anti-inflammatory mechanism of CAI and suggests its potential use as a therapeutic treatment in inflammatory diseases with excessive macrophage activation.
羧酰胺三唑（CAI）最初被开发为一种非细胞毒性抗癌药物，近期在多种炎症动物模型中的研究表明它具有抗炎活性。然而，其作用机制尚未明确。因此，本研究旨在确定CAI在脂多糖（LPS）诱导的RAW 264.7巨噬细胞中的抗炎作用，并确定相关的信号转导途径。结果显示，CAI对环氧化酶（COX）的活性没有直接影响，这表明其抗炎机制与抑制COX的非甾体抗炎药不同。对RAW264.7巨噬细胞的进一步研究表明，CAI通过降低LPS刺激诱导型一氧化氮合酶的表达来减少一氧化氮的产生，并下调细胞因子肿瘤坏死因子-α、白细胞介素（IL）-1β和IL-6的mRNA和蛋白质表达水平。CAI还显著降低了LPS刺激诱导的核因子（NF）-κB的DNA结合活性增加。关于NF-κB活性的相关机制，CAI抑制了核因子-κBα（IκB）抑制剂的磷酸化和降解，并降低了p65亚基的磷酸化水平及其随后的核转位。此外，CAI显著降低了LPS刺激后增加的p38、JNK和ERK的磷酸化形式，而p38、JNK和ERK的总表达水平保持不变。本研究结果表明，CAI通过减弱NF-κB和MAPK信号通路的激活以及降低促炎介质水平，减轻RAW 264.7巨噬细胞对LPS刺激的炎症反应。这为理解CAI的抗炎机制提供了新的视角，并表明其在巨噬细胞过度激活的炎症性疾病中具有潜在的治疗用途。
