Podoplanin-mediated TGF-β-induced epithelial-mesenchymal transition and its correlation with bHLH transcription factor DEC in TE-11 cells

Wu,Yunyan; Liu,Qiang; Yan,Xu; Kato,Yukio; Tanaka,Makiko; Inokuchi,Sadaki; Yoshizawa,Tadashi; Morohashi,Satoko; Kijima,Hiroshi

doi:10.3892/ijo.2016.3445

Podoplanin-mediated TGF-β-induced epithelial-mesenchymal transition and its correlation with bHLH transcription factor DEC in TE-11 cells

Authors:
- Yunyan Wu
- Qiang Liu
- Xu Yan
- Yukio Kato
- Makiko Tanaka
- Sadaki Inokuchi
- Tadashi Yoshizawa
- Satoko Morohashi
- Hiroshi Kijima
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Affiliations: Department of Pathology and Bioscience, Hirosaki University Graduate School of Medicine, Hirosaki, Aomori 036-8562, Japan, Department of Dental and Medical Biochemistry, Hiroshima University Graduate School of Biomedical Science, Hiroshima 734-8553, Japan, Department of Critical Care and Emergency Medicine, Tokai University School of Medicine, Isehara, Kanagawa 259-1193, Japan
Published online on: March 18, 2016 https://doi.org/10.3892/ijo.2016.3445
Pages: 2310-2320
Copyright: © Wu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Podoplanin is reported involved in the collective cell invasion, another tumor invasion style which is distinct from the single cell invasion, so-called epithelial-mesenchymal transition (EMT). In this study, we investigated the correlation between podoplanin and EMT-related markers in esophageal squamous cell carcinoma (ESCC), and evaluated its linkage with the basic helix-loop-helix (bHLH) transcription factor differentiated embryonic chondrocyte (DEC) 1 and DEC2. Three ESCC cell lines and human squamous cell carcinoma A431 cells were subjected to western blot analyses for podoplanin and EMT markers, as well as the expression of DEC1 and DEC2. By RT-qPCR and western blotting, we found that TGF-β increased the expression of podoplanin and mensenchymal markers (e.g., N-cadherin and vimentin), while decreased the expression of epithelial markers (e.g., Claudin-4 and E-cadherin), accompanied by Smad2 phosphorylation and slug activation. Moreover, TGF-β has different effects on the expression of DEC1 and DEC2, that is, it upregulates DEC1, but downregulates DEC2. Capability of cell proliferation, invasion and migration were further analyzed using CCK-8 assay, Matrigel-invasion assay, and the wound-healing assay, respectively. The proliferation, invasion and migration ability were significantly lost in podoplanin-knockdown cells when compared with the scrambled siRNA group. In addition to these changes, the expression of Claudin-4, but not that of Claudin-1 or E-cadherin, was induced by the siRNA against podoplanin. On the contrary, overexpression of DEC1 and DEC2 exhibits opposite effects on podoplanin, but only slight effect on Claudin-4 was detected. These data indicated that podoplanin is significantly associated with EMT of TE-11 cells, and may be directly or indirectly regulated by bHLH transcription factors DEC1 and DEC2.

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