Basic Study
Copyright ©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Dec 21, 2020; 26(47): 7513-7527
Published online Dec 21, 2020. doi: 10.3748/wjg.v26.i47.7513
Extracellular histones stimulate collagen expression in vitro and promote liver fibrogenesis in a mouse model via the TLR4-MyD88 signaling pathway
Zhi Wang, Zhen-Xing Cheng, Simon T Abrams, Zi-Qi Lin, Edwin Yates, Qian Yu, Wei-Ping Yu, Ping-Sheng Chen, Cheng-Hock Toh, Guo-Zheng Wang
Zhi Wang, Zhen-Xing Cheng, Wei-Ping Yu, Ping-Sheng Chen, Department of Pathology and Pathophysiology, Medical School, Southeast University, Nanjing 210009, Jiangsu Province, China
Zhi Wang, Qian Yu, Department of Gastroenterology, Zhongda Hospital, Nanjing 210009, Jiangsu Province, China
Zhen-Xing Cheng, Simon T Abrams, Cheng-Hock Toh, Guo-Zheng Wang, Department of Clinical Infection, Microbiology and Immunology, University of Liverpool, Liverpool L69 7BE, United Kingdom
Zhen-Xing Cheng, Department of Gastroenterology, The First Affiliated Hospital, Anhui Medical University, Hefei 230032, Anhui Province, China
Zi-Qi Lin, Department of Integrated Traditional Chinese and Western Medicine, Sichuan Provincial Pancreatitis Centre and West China-Liverpool Biomedical Research Centre, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
Edwin Yates, Department of Integrative Biology, University of Liverpool, Liverpool L69 7ZB, United Kingdom
Cheng-Hock Toh, Roald Dahl Haemostasis & Thrombosis Ctr, Royal Liverpool University Hospital, Liverpool L69 7BE, United Kingdom
Author contributions: Wang Z conceived the study; Cheng ZX assisted animal experiments and hydroxyproline measurements; Lin ZQ and Abrams ST assisted in performing in vitro experiments; Yates E synthesized and characterised non-anticoagulant heparin; Abrams ST helped edit figures; Yu Q, Yu WP, Chen PS, Toh CH and Wang GZ supervised the work and were involved in data analysis and manuscript writing; and all authors have read and agreed to the published version of the manuscript.
Supported by Key R & D Program of Jiangsu Province, No. BE2019712; British Heart Foundation, No. PG/14/19/30751 and No. PG/16/65/32313.
Institutional review board statement: The study was reviewed and approved by Medical School of Southeast University.
Institutional animal care and use committee statement: All procedures were performed according to State laws and monitored by local inspectors, and approved by the Animal Research Ethics Committee at the Medical School of the Southeast University.
Conflict-of-interest statement: No conflict interested has been claimed by any author.
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:
Corresponding author: Ping-Sheng Chen, MBChB, MD, PhD, Professor, Research Dean, Department of Pathology and Pathophysiology, Medical School, Southeast University, No. 87 Dingjiaqiao, Nanjing 210009, Jiangsu Province, China.
Received: August 27, 2020
Peer-review started: August 27, 2020
First decision: October 17, 2020
Revised: November 8, 2020
Accepted: December 6, 2020
Article in press: December 6, 2020
Published online: December 21, 2020
Research background

Currently, the molecular mechanisms of liver fibrosis are not fully understood. Recurrent liver injury or inflammation initiates wound healing along with fibrogenesis. However, what initiates this process is not clear. When cells die, damage-associated molecular patterns (DAMPs) are released. Histones are the most abundant DAMPs and are also ligands for TLR4, which in turn has been demonstrated to be involved in bile-duct-ligation-induced liver fibrosis. Lipopolysaccharide (LPS) is proposed to be a ligand for TLR4. Since recurrent liver injury does not naturally produce LPS but abundant extracellular histones, this study sought to investigate the potential roles of extracellular histones as TLR4 ligands in liver fibrosis.

Research motivation

Since our laboratory has been involved in studying the roles of DAMPs in critical illnesses, extracellular histones in liver fibrosis are of interest in terms of biological and clinical significance.

Research objectives

Our study aimed to clarify the roles of extracellular histones in fibrogenesis in vitro and in vivo.

Research methods

In our study, a hepatic stellate (HSC) cell line and animal models of liver fibrosis were used. Intervention studies with non-anticoagulant heparin (NAHP) to detoxify histones and TLR4-blocking antibodies to inhibit TLR4 were performed. In addition, TLR4 and MyD88 knockout mice were used to support the theory that the TLR4-MyD88 signaling pathway is involved in liver fibrosis in the CCl4 mouse model.

Research results

High levels of circulating histones were present when fibrosis was induced by CCl4 in the mouse model. Extracellular histones stimulated HSC cells in vitro to increase production of collagen I and alpha-smooth muscle actin. NAHP inhibited histone-enhanced collagen production in vitro, and reduce liver injury and fibrosis in vivo. TLR4 was involved in histone-enhanced collagen I production by HSC cells. In vivo, the TLR4-MyD88 signaling pathway mediated liver fibrosis, but whether circulating histones were the major activators of the pathway was not clear.

Research conclusions

Recurrent liver injury releases extracellular histones that potentially activate TLR4-MyD88 signaling to promote liver fibrosis. The ability of NAHP to detoxify circulating histones has the potential for treatment of liver injury and prevention of liver fibrosis.

Research perspectives

Future studies demonstrating the contribution of circulating histones to activation of the TLR4-MyD88 signaling and downstream pathways will validate their role in liver fibrosis. Development of effective anti-histone therapies to reduce liver injury and prevent liver fibrosis have potential in the management of diseases with recurrent liver injury.