Basic Study
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World J Gastroenterol. Dec 28, 2021; 27(48): 8302-8322
Published online Dec 28, 2021. doi: 10.3748/wjg.v27.i48.8302
Hepatitis B core antigen modulates exosomal miR-135a to target vesicle-associated membrane protein 2 promoting chemoresistance in hepatocellular carcinoma
Xiao-Cui Wei, Ya-Ru Xia, Ping Zhou, Xing Xue, Shuang Ding, Li-Juan Liu, Fan Zhu
Xiao-Cui Wei, Ya-Ru Xia, Ping Zhou, Xing Xue, Shuang Ding, Li-Juan Liu, Fan Zhu, State Key Laboratory of Virology, Hubei Province Key Laboratory of Allergy and Immunology, Department of Medical Microbiology, School of Medicine, Wuhan University, Wuhan 430071, Hubei Province, China
Author contributions: Zhu F conceived the study, was in charge of overall direction and planning, and revised the manuscript; Wei XC designed and performed the experiments, and wrote the paper; Wei XC and Liu LJ revised the manuscript; Xia YR and Zhou P participated in data collection; Xue X was responsible for bioinformatics data; Ding S, and Liu LJ analyzed data; all authors read and approved the final manuscript and agreed to be accountable for all aspects of the report.
Supported by National Natural Science Foundation of China, No. 81971943 and 81772196; and the Medical Science Advancement Program (Basic Medical Sciences) of Wuhan University, No. TFJC 2018002.
Institutional review board statement: The study was reviewed and approved by the Institutional Review Board of Wuhan University, School of Basic Medical Sciences, and the study was carried out following The Code of Ethics of the World Medical Association (Declaration of Helsinki) for experiments involving humans. Informed consent was obtained for experimentation with human subjects, and their privacy rights were consistently observed.
Conflict-of-interest statement: The authors hereby declare that no conflict of interest exists.
Data sharing statement: Datasets available from the corresponding author upon request.
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: Fan Zhu, PhD, Professor, State Key Laboratory of Virology, Hubei Province Key Laboratory of Allergy and Immunology, Department of Medical Microbiology, School of Medicine, Wuhan University, No. 185 Donghu Road, Wuhan 430071, Hubei Province, China. fanzhu@whu.edu.cn
Received: April 27, 2021
Peer-review started: April 27, 2021
First decision: June 13, 2021
Revised: June 22, 2021
Accepted: November 11, 2021
Article in press: December 11, 2021
Published online: December 28, 2021
Abstract
BACKGROUND

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors. The association of hepatitis B virus (HBV) infection with HCC is hitherto documented. Exosomal miRNAs contribute to cancer progression and chemoresistance. HBV X protein has been known to modulate miRNAs that facilitate cell proliferation and the process of hepatocarcinogenesis. However, there has been no report on hepatitis B core antigen (HBc) regulating exosomal miRNAs to induce drug resistance of HCC cells.

AIM

To elucidate the mechanism by which HBc promotes Doxorubicin hydrochloride (Dox) resistance in HCC.

METHODS

Exosomes were isolated by ultracentrifugation. The morphology and size of exosomes were evaluated by Dynamic Light Scattering (DLS) and transmission electron microscopy (TEM). The miRNAs differentially expressed in HCC were identified using The Cancer Genome Atlas (TCGA) database. The level of miR-135a-5p in patient tissue samples was detected by quantitative polymerase chain reaction. TargetScan and luciferase assay were used to predict and prove the target gene of miR-135a-5p. Finally, we identified the effects of miR-135a-5p on anti-apoptosis and the proliferation of HCC in the presence or absence of Dox using flow cytometry, Cell counting kit 8 (CCK-8) assay and western blot.

RESULTS

We found that HBc increased the expression of exosomal miR-135a-5p. Integrated analysis of bioinformatics and patient samples found that miR-135a-5p was increased in HCC tissues in comparison with paracancerous tissues. Bioinformatic analysis and in vitro validation identified vesicle-associated membrane protein 2 (VAMP2) as a novel target gene of miR-135a-5p. Functional assays showed that exosomal miR-135a-5p induced apoptosis protection, cell proliferation, and chemotherapy resistance in HCC. In addition, the rescue experiment demonstrated that VAMP2 reversed apoptosis protection, cell growth, and drug resistance by miR-135a-5p. Finally, HBc promoted HCC anti-apoptosis, proliferation, and drug resistance and prevented Dox-induced apoptosis via the miR-135a-5p/VAMP2 axis.

CONCLUSION

These data suggested that HBc upregulated the expression of exosomal miR-135a-5p and promoted anti-apoptosis, cell proliferation, and chemical resistance through miR-135a-5p/VAMP2. Thus, our work indicated an essential role of the miR-135a-5p/VAMP2 regulatory axis in chemotherapy resistance of HCC and a potential molecular therapeutic target for HCC.

Keywords: Hepatocellular carcinoma, Exosomes, miR-135a-5p, Anti-apoptosis, Proliferation, Chemoresistance

Core Tip: Hepatitis B virus infection is the most common cause of hepatocellular carcinoma (HCC). Drug resistance is the primary reason for the high mortality of HCC patients. We demonstrated that hepatitis B core antigen (HBc) increased exosomal miR-135a-5p. Tissue samples showed that the level of miR-135a-5p was significantly elevated in HCC tissues. Vesicle-associated membrane protein 2 (VAMP2) was demonstrated to be a target gene of miR-135a-5p. Further investigation recommended that HBc enhanced the anti-apoptosis, cell proliferation, and chemotherapy resistance of HCC cells through exosomal miR-135a-5p by targeting VAMP2. Our findings reveal that HBc can cause anti-cancer drug resistance in HCC and provide us with a novel mechanism underlying drug resistance in cancer chemotherapy.