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Ultrasound-Targeted Microbubble Destruction-Mediated Downregulation of EZH2 Inhibits Stemness and Epithelial-Mesenchymal Transition of Liver Cancer Stem Cells

Authors Wu J, Sun L, Liu T, Dong G

Received 26 June 2020

Accepted for publication 18 November 2020

Published 11 January 2021 Volume 2021:14 Pages 221—237

DOI https://doi.org/10.2147/OTT.S269589

Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Professor Gaetano Romano


Jie Wu, Lulu Sun, Tingting Liu, Gang Dong

Department of Ultrasound Intervention, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan Province, People’s Republic of China

Correspondence: Gang Dong
Department of Ultrasound Intervention, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan Province 450052, People’s Republic of China
Tel +86-371-66913289
Email dongg_dongang@163.com

Background: Cancer cells could show the characteristics of cancer stem cells (CSCs) through epithelial-mesenchymal transition (EMT). EZH2 was associated with EMT. Ultrasound-targeted microbubble destruction (UTMD) could enhance gene transfection efficiency. Here, we explored the effect of UTMD-mediated shEZH2 on liver CSCs.
Methods: EZH2 expression in liver cancer and the overall survival of liver cancer patients were analyzed by bioinformatics. Liver CSCs (CD133+HuH7) were sorted by flow cytometry. After transfection of shEZH2 through UTMD (UTMD-shEZH2) or liposome (LIP-shEZH2), the viability, proliferation, sphere formation, migration, and invasion of CD133+HuH7 cells were detected by MTT, colony formation, tumor-sphere formation, wound healing, and transwell assays, respectively. A mice subcutaneous-xenotransplant tumor model was established by injecting CD133+HuH7 or CD133HuH7 cells into the limbs of mice. Tumor weight and volume were documented. The expressions of EZH2, EMT-related factors, and STAT3/PI3K/AKT pathway-related factors in CD133+HuH7 cells or tumor tissues were detected by RT-qPCR, Western blot, or immunohistochemical.
Results: EZH2 was high-expressed in liver cancer, and the patients with high expression of EZH2 had a poor survival. CD133+ HuH7 cells had higher EZH2 expression, higher viability, and stronger sphere-forming and tumor-forming abilities than CD133 HuH7 cells. ShEZH2 inhibited the viability, proliferation, sphere formation, migration, and invasion of CD133+ HuH7 cells, decreased the weight and volume of the xenotransplant tumor, inhibited the expressions of EZH2, Vimentin, N-Cadherin, Twist-1, p-STAT3, p-PI3K, and p-AKT, and increased E-Cadherin expression. UTMD-shEZH2 caused a stronger effect on CD133+ HuH7 cells than LIP-shEZH2.
Conclusion: UTMD-mediated shEZH2 inhibited the stemness and EMT of liver CSCs in vitro and in vivo through regulating the STAT3/PI3K/AKT pathway.

Keywords: liver cancer, cancer stem cells, ultrasound-targeted microbubble destruction, epithelial-mesenchymal transition, xenotransplant

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