尼达尼布通过调节TGF-β/Smad信号通路抑制A549肺泡上皮细胞的上皮-间质转化。

Nintedanib inhibits epithelial-mesenchymal transition in A549 alveolar epithelial cells through regulation of the TGF-β/Smad pathway.

作者信息

Ihara Hiroaki, Mitsuishi Yoichiro, Kato Motoyasu, Takahashi Fumiyuki, Tajima Ken, Hayashi Takuo, Hidayat Moulid, Winardi Wira, Wirawan Aditya, Hayakawa Daisuke, Kanamori Koichiro, Matsumoto Naohisa, Yae Toshifumi, Sato Tadashi, Sasaki Shinichi, Takamochi Kazuya, Suehara Yoshiyuki, Ogura Dai, Niwa Shin-Ichiro, Suzuki Kenji, Takahashi Kazuhisa

机构信息

Department of Respiratory Medicine, Juntendo University, Graduate School of Medicine, Tokyo, Japan; Research Institute for Diseases of Old Ages, Juntendo University, Graduate School of Medicine, Tokyo, Japan.

出版信息

Respir Investig. 2020 Jul;58(4):275-284. doi: 10.1016/j.resinv.2020.01.003. Epub 2020 Apr 29.

DOI:10.1016/j.resinv.2020.01.003

PMID:32359980

Abstract

BACKGROUND

Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic lung disorder. Recent studies have suggested that epithelial-mesenchymal transition (EMT) of alveolar epithelial cells influences development of pulmonary fibrosis, which is mediated by transforming growth factor β (TGF-β). Tumor necrosis factor α (TNF-α), an important proinflammatory cytokine in IPF, has been shown to enhance TGF-β-induced EMT. Nintedanib, a multiple tyrosine kinase inhibitor that is currently used to treat IPF, has been shown to suppress EMT in various cancer cell lines. However, the mechanism of EMT inhibition by nintedanib and its effect on TGF-β and TNF-α signaling pathways in alveolar epithelial cells have not been fully elucidated.

METHODS

A549 alveolar epithelial cells were stimulated with TGF-β2 and TNF-α, and the effects of nintedanib on global gene expression were evaluated using microarray analysis. Furthermore, Smad2/3 phosphorylation was assessed using western blotting.

RESULTS

We found that in A549 cells, TGF-β2 and TNF-α treatment induces EMT, which was inhibited by nintedanib. Gene ontology analysis showed that nintedanib significantly attenuates the gene expression of EMT-related cellular pathways and the TGF-β signaling pathway, but not in the TNF-α-mediated signaling pathway. Furthermore, hierarchical cluster analysis revealed that EMT-related genes were attenuated in nintedanib-treated cells. Additionally, nintedanib was found to markedly suppress phosphorylation of Smad2/3.

CONCLUSION

Nintedanib inhibits EMT by mediating EMT-related gene expression and the TGF-β/Smad pathway in A549 alveolar epithelial cells.

摘要

背景

特发性肺纤维化（IPF）是一种进行性纤维化肺部疾病。最近的研究表明，肺泡上皮细胞的上皮-间质转化（EMT）影响肺纤维化的发展，这一过程由转化生长因子β（TGF-β）介导。肿瘤坏死因子α（TNF-α）是IPF中一种重要的促炎细胞因子，已被证明可增强TGF-β诱导的EMT。尼达尼布是一种目前用于治疗IPF的多酪氨酸激酶抑制剂，已被证明可抑制多种癌细胞系中的EMT。然而，尼达尼布抑制EMT的机制及其对肺泡上皮细胞中TGF-β和TNF-α信号通路的影响尚未完全阐明。

方法

用TGF-β2和TNF-α刺激A549肺泡上皮细胞，并用微阵列分析评估尼达尼布对整体基因表达的影响。此外，用蛋白质印迹法评估Smad2/3磷酸化。

结果

我们发现，在A549细胞中，TGF-β2和TNF-α处理诱导EMT，而尼达尼布可抑制EMT。基因本体分析表明，尼达尼布显著减弱EMT相关细胞途径和TGF-β信号通路的基因表达，但对TNF-α介导的信号通路无影响。此外，层次聚类分析显示，在尼达尼布处理的细胞中，EMT相关基因减弱。另外，发现尼达尼布可显著抑制Smad2/3的磷酸化。