Basic Study
Copyright ©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. May 21, 2020; 26(19): 2349-2373
Published online May 21, 2020. doi: 10.3748/wjg.v26.i19.2349
MiR-301a transcriptionally activated by HIF-2α promotes hypoxia-induced epithelial-mesenchymal transition by targeting TP63 in pancreatic cancer
Kun-Dong Zhang, Bin Hu, Gang Cen, Yu-Han Yang, Wei-Wei Chen, Zeng-Ya Guo, Xiao-Feng Wang, Qian Zhao, Zheng-Jun Qiu
Kun-Dong Zhang, Bin Hu, Gang Cen, Yu-Han Yang, Wei-Wei Chen, Zeng-Ya Guo, Xiao-Feng Wang, Zheng-Jun Qiu, Department of General Surgery, Shanghai Key Laboratory of Pancreatic Disease, Shanghai General Hospital, Shanghai Jiaotong University, Shanghai 200080, China
Qian Zhao, Key Laboratory of Cell Differentiation and Apoptosis of National Ministry of Education, Department of Pathophysiology and Rui-Jin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China
Author contributions: Zhang KD prepared the manuscript, which was critically reviewed by all co-authors; Zhao Q and Qiu ZJ designed the study; Zhang KD, Hu B, Cen G, and Yang YH conducted the study; Chen WW, Guo ZY, and Wang XF performed the statistical analyses; Zhang KD, Hu B, and Cen G contributed equally to this work; Zhao Q is an equal corresponding author; all authors approved the final version of the manuscript.
Supported by National Natural Science Foundation of China, No. 81372640.
Institutional review board statement: This study protocol was reviewed and approved by the Medical Ethics Committee of Shanghai General Hospital.
Institutional animal care and use committee statement: This study protocol was reviewed and approved by the Animal Experimental Ethical Committee of Shanghai General Hospital.
Conflict-of-interest statement: The authors declare that they have no competing interests.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Corresponding author: Zheng-Jun Qiu, MD, PhD, Chief Doctor, Professor, Surgeon, Department of General Surgery, Shanghai Key Laboratory of Pancreatic Disease, Shanghai General Hospital, Shanghai Jiaotong University, No. 100, Haining Road, Shanghai 200080, China. qiudoctor@sina.com
Received: November 18, 2019
Peer-review started: November 18, 2019
First decision: January 19, 2020
Revised: February 20, 2010
Accepted: April 18, 2020
Article in press: April 18, 2020
Published online: May 21, 2020
Processing time: 174 Days and 21.7 Hours
Abstract
BACKGROUND

Pancreatic cancer (PC) is one of the deadliest cancers worldwide. PC metastasis involves a complex set of events, including epithelial-mesenchymal transition (EMT), that increase tumor cell invasiveness. Recent evidence has shown that hypoxia is a major EMT regulator in pancreatic cancer cells and facilitates metastasis; however, the mechanisms remain elusive.

AIM

To investigate the role of miR-301a in hypoxia-induced EMT in PC cells.

METHODS

Real-time PCR and Western blot analysis were used to detect the expression of miR-301a and EMT markers in PDAC cells cultured in hypoxic and normoxic conditions. Western blot analysis was used to detect the expression of EMT markers in PDAC cells with miR-301a overexpression. Wound healing assay and Transwell assay were used to detect the migration capabilities of PDAC cells with miR-301a overexpression and knockout. Luciferase assay was used to detect the miR-301a promoter and the 3’ untranslated region activity of TP63. Orthotopic PC mouse models were used to study the role of miR-301a in metastasis of PDAC cells in vivo. In situ hybridization assay was used to detect the expression of miR-301a in PDAC patient samples (adjacent paratumor and paired tumor tissues).

 

RESULTS

Hypoxic environment could directly promote the EMT of PC cells. The expression level of miR-301a was increased in a HIF2α dependent manner in hypoxia-cultured CFPAC-1 and BxPC-3 cells. Overexpression of miR-301a enhanced the hypoxia-induced EMT of PC cells, while knocking out miR-301a result in the suppression of hypoxia-induced EMT. TP63 was a direct target of miR-301a and involved in the metastatic process of PC cells. Furthermore, miR-301a upregulation facilitated PDAC distant metastasis and lymph node metastasis in vivo. Additionally, miR-301a overexpression was indicative of aggressive clinicopathological behaviors and poor prognosis.

CONCLUSION

The newly identified HIF-2α-miR301a-TP63 signaling pathway may play a crucial role in hypoxia-induced EMT in PDAC cells.

Keywords: MiR-301a; Epithelial-mesenchymal transition; Pancreatic cancer; Hypoxia; HIF-2α; TP63

Core tip: In this study, we found that miR-301a expression was increased during the process of hypoxia-induced epithelial-mesenchymal transition (EMT) in pancreatic cancer (PC) cells. miR-301a overexpression facilitated hypoxia-induced EMT, while miR-301a knockout inhibited hypoxia-induced EMT in PC cells. The increased expression of miR-301a was transcriptionally regulated by HIF-2α. In addition, we identified a new target gene of miR-301a, namely, TP63, and confirmed that TP63 was involved in EMT and metastasis of PC cells. Collectively, our data suggest that the newly identified HIF-2α-miR301a-TP63 signaling pathway plays a crucial role in hypoxia-induced EMT in pancreatic ductal adenocarcinoma and that miR-301a may serve as a new prognostic biomarker and candidate miRNA for tumor diagnosis and treatment.