MAPK Pathway Mediates Epithelial-Mesenchymal Transition Induced by Paraquat in Alveolar Epithelial Cells
MAPK Pathway Mediates Epithelial-Mesenchymal Transition Induced by Paraquat in Alveolar Epithelial Cells
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MAPK 通路介导百草枯诱导的肺泡上皮细胞上皮间质转化
DOI：
10.1002/tox.22146
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发表时间：
2016-11-01
影响因子：
4.5
通讯作者：
Yang, Hui-Fang
中科院分区：
文献类型：
--
作者：
Huang, Min;Wang, Ya-Peng;Yang, Hui-Fang
上皮-间充质转化(EMT)被认为参与了百草枯(PQ)诱导的肺纤维化过程，但其分子机制尚未明确。本研究调查了PQ中毒后EMT的潜在参与。免疫印迹法检测不同浓度PQ作用后多个时间点EMT标志物E-钙粘附素和α-SMA的表达。PQ处理后，在显微镜下观察EMT的诱导。采用逆转录聚合酶链式反应(RT-PCR)检测肝纤维化相关基因，包括基质金属蛋白酶2(MMP2)、基质金属蛋白酶9(MMP9)、I型胶原(Col I)和III型胶原(Col III)的表达水平。使用MAPK的选择性药物抑制剂分析信号通路。用划痕和Transwell法检测细胞的迁移能力。结果显示，PQ可诱导RLE-6NT细胞形成间充质细胞特征，表现为上皮细胞标志物E-钙粘附素显著降低，细胞外基质(ECM)标志物α-平滑肌肌动蛋白显著增加，且呈剂量和时间依赖性。此外，PQ处理的RLE-6NT细胞具有EMT样表型，表现为MMP2、MMP9、Col I和Col III表达增加，迁移能力增强。信号通路分析表明，PQ诱导的EMT通过激活MAPK通路导致ERK-1和Smad2的磷酸化。目前的研究结果表明，PQ诱导的肺纤维化是通过EMT发生的，而EMT是由MAPK通路介导的。这表明MAPK通路在肺泡上皮细胞中是一个有希望的治疗靶点。(C)2015年威利期刊公司。
Epithelial-mesenchymal transition (EMT) is believed to be involved in lung fibrosis process induced by paraquat (PQ); however, the molecular mechanism of this process has not been clearly established. The present study investigated the potential involvement of EMT after PQ poisoning. The expressions of EMT markers, such as E-cadherin and alpha-smooth muscle actin (alpha-SMA), at multiple time points after exposure to different concentrations of PQ were evaluated by western blot analysis. Following PQ treatment, EMT induction was observed under microscopy. Related fibrosis genes, including Matrix metalloproteinase 2 (MMP-2), Matrix metalloproteinase 9 (MMP-9), collagens type I (COL I), and type III (COL III), were also evaluated by measuring their mRNA levels using RT-PCR analysis. Signaling pathways were analyzed using selective pharmacological inhibitors for MAPK. Cell migration ability was evaluated by scratch wound and Transwell assays. The data showed that PQ-induced epithelial RLE-6NT cells to develop mesenchymal cell characteristics, as indicated by a significant decrease in the epithelial marker E-cadherin and a significant increase in the extracellular matrix (ECM) marker alpha-smooth muscle actin in a dose and time-dependent manner. Moreover, PQ-treated RLE-6NTcells had an EMT-like phenotype with elevated expression of MMP-2, MMP-9, and COL I and COL III and enhanced migration ability. Signal pathway analysis revealed that PQ-induced EMT led to ERK-1 and Smad2 phosphorylation through activation of the MAPK pathway. The results of the current study indicate that PQ-induced pulmonary fibrosis occurs via EMT, which is mediated by the MAPK pathway. This implies that the MAPK pathway is a promising therapeutic target in alveolar epithelial cells. (C) 2015 Wiley Periodicals, Inc.