Methotrexate-Induced Epithelial–Mesenchymal Transition in the Alveolar Epithelial Cell Line A549

Kawami, Masashi; Harabayashi, Rika; Miyamoto, Mioka; Harada, Risako; Yumoto, Ryoko; Takano, Mikihisa

doi:10.1007/s00408-016-9935-7

Cited by 35 publications

(27 citation statements)

References 34 publications

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“…Reverse transcription of the total RNA into cDNA was performed using ReverTra Ace (TOYOBO, Osaka, Japan). Quantitative evaluation for indicated mRNA expression was performed by real-time PCR analysis using the CFX Connect™ Real-Time PCR Detection System (Bio-Rad Laboratories, Inc., Hercules, CA, USA) and the KAPA SYBR ® FAST qPCR Kit (NIPPON Genetics, Tokyo, Japan), as previously described (16). The primer sequences were as follows:…”

Section: Real-time Pcr Analysismentioning

confidence: 99%

Transport of AOPP-Albumin into Human Alveolar Epithelial A549 Cell

et al. 2018

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Section: Real-time Pcr Analysismentioning

confidence: 99%

Transport of AOPP-Albumin into Human Alveolar Epithelial A549 Cell

et al. 2018

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“…Several clinical reports suggested that MTX induces serious pulmonary side effects, such as pulmonary fibrosis . Recently, various efforts have been made to clarify the mechanism underlying MTX‐induced lung injury, showing the association with several factors including cytokine release, reactive oxygen species and epithelial‐mesenchymal transition . Nevertheless, the detailed mechanisms of MTX toxicity in the lungs are poorly understood.…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, mRNA expression of some type II alveolar epithelial cell markers such as peptide transporter 2 (PEPT2) in H441 cells was higher than that in A549 cells, indicating that H441 would be a better alveolar type II epithelial cell model than A549. On the other hand, some reports suggested that MTX‐induced lung injury may be associated with epithelial‐mesenchymal transition (EMT) . In addition, EMT would occur in type II cells, but not in type I cells, in the lungs of patients with idiopathic pulmonary fibrosis and in human primary cultured alveolar type II epithelial cells .…”

Section: Introductionmentioning

confidence: 99%

“…[6,7] Recently, various efforts have been made to clarify the mechanism underlying MTX-induced lung injury, showing the association with several factors including cytokine release, reactive oxygen species and epithelial-mesenchymal transition. [8][9][10][11] Nevertheless, the detailed mechanisms of MTX toxicity in the lungs are poorly understood. In addition, the investigation on MTX transport in alveolar epithelium is limited, although MTX-induced pulmonary toxicity is likely associated with the distribution of MTX into the lungs.…”

Section: Introductionmentioning

confidence: 99%

“…On the other hand, some reports suggested that MTX-induced lung injury may be associated with epithelial-mesenchymal transition (EMT). [10,21] In addition, EMT would occur in type II cells, but not in type I cells, in the lungs of patients with idiopathic pulmonary fibrosis and in human primary cultured alveolar type II epithelial cells. [22,23] Therefore, we focused on MTX transport in alveolar type II epithelial cells using H441 as a model cell line.…”

Section: Introductionmentioning

confidence: 99%

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Reduced folate carrier-mediated methotrexate transport in human distal lung epithelial NCl-H441 cells

Kawami

Honda

Miyamoto

et al. 2019

Journal of Pharmacy and Pharmacology

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Objectives We had previously found that reduced folate carrier (RFC; SLC19A1) is mainly involved in an influx of transport of methotrexate (MTX), a folate analogue, using alveolar epithelial A549 cells. Therefore, we examined MTX uptake in NCl‐H441 (H441) cells, another in vitro alveolar epithelial model, focusing on the localization of RFC in the present study. Methods Transport function of RFC in H441 cells was studied using [3H]MTX. Key findings The uptake of MTX was increased remarkably after pretreatment of the cell monolayer with ethylenediaminetetraacetic acid (EDTA) in H441 cells but not in A549 cells, indicating the contribution of the basolaterally located transporter. In addition, folic acid and thiamine monophosphate, RFC inhibitors, inhibited the uptake of MTX from the basolateral side of the H441 cells. In order to compare the function of RFC on the apical and basolateral sides of the cells, the uptake of MTX from each side was examined using a Transwell chamber. Intracellular MTX amounts from the basolateral side were found to be significantly higher than those from the apical side. Conclusions These findings suggest that the distribution of MTX in the lung alveolar epithelial cells may be mediated by basolaterally located RFC in alveolar epithelial cells.

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Effect of transforming growth factor-β1 on functional expression of monocarboxylate transporter 1 in alveolar epithelial A549 cells

Uddin

Kawami

Yumoto

et al. 2020

Naunyn-Schmiedeberg's Arch Pharmacol

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Epithelial-mesenchymal transition (EMT) contributes to the development of severe lung diseases, such as pulmonary fibrosis. Recently, it has been reported that EMT involves complex metabolic reprogramming triggered by several factors including transforming growth factor (TGF-β1) and that monocarboxylate transporter (MCT1) plays an essential role in these metabolic changes. The aim of the present study was to clarify the functional expression of MCT1 during TGF-β1-induced EMT in alveolar epithelial A549 cells. The transport function of MCT1 in A549 cells was examined using [ 3 H]γ-hydroxybutyrate (GHB) and [ 3 H]lactic acid (LA) as substrates and α-cyano-4-hydroxycinnamate (CHC), lactic acid, phloretin, and AR-C155858 (AR) as inhibitors of MCT1. EMT was induced by treating the cells with TGF-β1. mRNA and protein expression levels were analyzed using real-time PCR and western blotting, respectively. Time-, temperature-, and pH-dependent GHB and LA uptake were observed in A549 cells. CHC, lactic acid, phloretin, and AR significantly inhibited the uptake of GHB in a concentration-dependent manner, suggesting that MCT1 is primarily responsible for transport of monocarboxylates such as GHB and LA in A549 cells. TGF-β1 treatment significantly enhanced GHB and LA uptake as well as the mRNA and protein expression levels of MCT1 in A549 cells. These changes were neutralized by co-treatment with SB431542, an inhibitor for the TGF-β1 signaling pathway.CHC and AR had no effect on TGF-β1-induced EMT-related gene expression changes.Here, we have clearly characterized functional expression of MCT1 in A549 cells and have shown that MCT1 may be upregulated via the TGF-β1 signaling pathway.

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Methotrexate-Induced Epithelial–Mesenchymal Transition in the Alveolar Epithelial Cell Line A549

Abstract: We have demonstrated that phenotypic changes of A549 cells induced by MTX are partly mediated by a TGF-β1-related intracellular signaling pathway, although TGF-β1 itself is not directly involved in this process.

Cited by 35 publications

References 34 publications

Transport of AOPP-Albumin into Human Alveolar Epithelial A549 Cell

Transport of AOPP-Albumin into Human Alveolar Epithelial A549 Cell

Reduced folate carrier-mediated methotrexate transport in human distal lung epithelial NCl-H441 cells

Effect of transforming growth factor-β1 on functional expression of monocarboxylate transporter 1 in alveolar epithelial A549 cells

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