Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Seoul National University Hospital, Seoul, 03080, Korea.
Department of Internal Medicine, Seoul National University College of Medicine, Seoul, 03080, Korea.
Exp Mol Med. 2018 Jul 6;50(7):1-9. doi: 10.1038/s12276-018-0114-1.
Although inflammation, oxidative stress, and protease-antiprotease imbalance have been referred to as a pathogenic triad in chronic obstructive pulmonary disease (COPD), little is known about how they interact. The objectives of this study were to elucidate the effect of cigarette smoke extract (CSE) on the neutrophil elastase (NE)-induced inflammatory response and its molecular mechanism in bronchial epithelial cells. We observed that NE activated extracellular signal-regulated kinase (ERK) and induced IL-8 production. Blocking ERK activation using a MEK inhibitor (U0126) suppressed NE-induced IL-8 secretion and knockdown of proteinase-activated receptor 2 (PAR2) using siRNAs inhibited both NE-induced ERK activation and subsequent IL-8 release, suggesting that NE-induced IL-8 production is dependent on PAR2-mediated ERK activation. Interestingly, pre-exposure to CSE markedly enhanced NE-induced IL-8 production. As PAR2 acts as a receptor for NE, we next investigated the effect of CSE on PAR2 expression as a molecular mechanism for the increased IL-8 production induced by NE in CSE exposed cells. CSE, but not NE, increased the expression of PAR2 mRNA and surface membrane protein. Inhibition of p38 MAPK reduced PAR2 expression induced by CSE while inhibition of the ERK and Akt pathway had no effect. Consequently, p38 inhibition significantly abrogated CSE-induced enhancement of IL-8 production in NE-treated cells. Of note, we observed increased PAR2 levels in lung homogenates and lung epithelial cells from CSE-treated mice and from both smokers and patients with COPD. Taken together, these results suggest that CSE upregulates PAR2 in normal human bronchial epithelial cells, thereby enhancing the inflammatory response to NE.
虽然炎症、氧化应激和蛋白酶-抗蛋白酶失衡被认为是慢性阻塞性肺疾病（COPD）的致病三联征，但它们之间的相互作用知之甚少。本研究旨在阐明香烟烟雾提取物（CSE）对中性粒细胞弹性蛋白酶（NE）诱导的支气管上皮细胞炎症反应的影响及其分子机制。我们观察到 NE 激活细胞外信号调节激酶（ERK）并诱导 IL-8 产生。使用 MEK 抑制剂（U0126）阻断 ERK 激活抑制了 NE 诱导的 IL-8 分泌，并且使用 siRNA 敲低蛋白酶激活受体 2（PAR2）抑制了 NE 诱导的 ERK 激活和随后的 IL-8 释放，表明 NE 诱导的 IL-8 产生依赖于 PAR2 介导的 ERK 激活。有趣的是，预先暴露于 CSE 可显著增强 NE 诱导的 IL-8 产生。由于 PAR2 作为 NE 的受体起作用，因此我们接下来研究了 CSE 对 PAR2 表达的影响，这是 CSE 暴露细胞中 NE 诱导的 IL-8 产生增加的分子机制。CSE 而非 NE 增加了 PAR2 mRNA 和表面膜蛋白的表达。抑制 p38 MAPK 减少了 CSE 诱导的 PAR2 表达，而抑制 ERK 和 Akt 途径则没有影响。因此，p38 抑制可显著消除 CSE 诱导的 NE 处理细胞中 IL-8 产生的增强。值得注意的是，我们观察到 CSE 处理的小鼠以及吸烟者和 COPD 患者的肺匀浆和肺上皮细胞中 PAR2 水平升高。总之，这些结果表明 CSE 在正常的人支气管上皮细胞中上调 PAR2，从而增强对 NE 的炎症反应。
