Metabolic Disorders and Inflammatory Pathologies Laboratory (MDIPL), School of Life Sciences, Central University of Gujarat, Sector 30, Gandhinagar, Gujarat, 382030, India.
Department of Biotechnology, Atmiya University, Rajkot, Gujarat, 360005, India.
Chem Biol Interact. 2022 Jan 5;351:109771. doi: 10.1016/j.cbi.2021.109771. Epub 2021 Dec 2.
Cigarette smoke exposure leads to upregulation of cyclooxygenase-2 (COX-2), an inducible enzyme that synthesizes prostaglandin E2 (PGE2) and promotes airway inflammation. COX-2 overexpression is frequently implicated in inflammation, invasion, metastasis, and epithelial-mesenchymal transition (EMT). However, its detailed molecular mechanism in cigarette smoke induced EMT is not clear. Further, fisetin, a bioflavonoid, exhibits antioxidant and anti-inflammatory properties, but its effect in modulating COX-2-mediated inflammation and downstream sequelae remains unexplored. Therefore, we have investigated the mechanism of cigarette smoke-induced COX-2-mediated EMT in airway epithelial cells and examined the role of fisetin in controlling this aberration. MTT, trypan blue staining, gelatin zymography, Western blotting, invasion, wound healing, and tumor sphere formation assays in cigarette smoke extract (CSE) and/or fisetin treated airway epithelial cells, and in-silico molecular docking studies were performed. Results revealed that CSE exposure increased the expression and activity of COX-2, MMP-2/9, and β-catenin and also enhanced expression of EMT markers leading to higher migration and invasion potential of airway epithelial cells. A specific COX-2 inhibitor NS-398 as well as fisetin treatment reversed the expression of EMT biomarkers, reduced the activity of MMP-2/9, and blocked the migration and invasion potential induced by CSE. Further, PGE2 also increased MMPs activity, invasion, and migration potential similar to CSE, which were significantly reversed by fisetin. In-silico studies showed a high binding affinity of fisetin to key EMT associated proteins, validating its anti-EMT potential. Thus, our study firstly unearths the mechanism of CSE-induced EMT in airway epithelial cells via COX-2/MMP/β-catenin pathway, and secondly, it reveals that fisetin could significantly reverse CSE-induced EMT by inhibiting COX-2, indicating that fisetin could be an effective drug candidate for cigarette smoke-induced lung dysfunction.
香烟烟雾暴露导致环氧化酶-2（COX-2）的上调，COX-2 是一种诱导酶，可合成前列腺素 E2（PGE2）并促进气道炎症。COX-2 的过表达常与炎症、侵袭、转移和上皮-间充质转化（EMT）有关。然而，其在香烟烟雾诱导的 EMT 中的详细分子机制尚不清楚。此外，漆黄素是一种生物类黄酮，具有抗氧化和抗炎特性，但它在调节 COX-2 介导的炎症及其下游后果方面的作用仍未得到探索。因此，我们研究了香烟烟雾诱导的 COX-2 介导的 EMT 在气道上皮细胞中的机制，并研究了漆黄素在控制这种异常中的作用。在香烟烟雾提取物（CSE）和/或漆黄素处理的气道上皮细胞中进行了 MTT、台盼蓝染色、明胶酶谱、Western 印迹、侵袭、划痕愈合和肿瘤球体形成测定，以及计算机分子对接研究。结果表明，CSE 暴露增加了 COX-2、MMP-2/9 和 β-连环蛋白的表达和活性，并增强了 EMT 标志物的表达，导致气道上皮细胞更高的迁移和侵袭能力。特异性 COX-2 抑制剂 NS-398 以及漆黄素治疗逆转了 EMT 生物标志物的表达，降低了 MMP-2/9 的活性，并阻断了 CSE 诱导的迁移和侵袭能力。此外，PGE2 也增加了 MMPs 的活性、侵袭和迁移能力，与 CSE 相似，这与漆黄素的显著逆转有关。计算机模拟研究表明，漆黄素与 EMT 相关蛋白具有很高的结合亲和力，验证了其抗 EMT 的潜力。因此，我们的研究首次揭示了 CSE 通过 COX-2/MMP/β-连环蛋白途径诱导气道上皮细胞 EMT 的机制，其次表明漆黄素通过抑制 COX-2 可显著逆转 CSE 诱导的 EMT，表明漆黄素可能是一种有效的药物候选物，用于治疗香烟烟雾引起的肺功能障碍。
