Basic Study
Copyright ©The Author(s) 2021. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Nov 21, 2021; 27(43): 7509-7529
Published online Nov 21, 2021. doi: 10.3748/wjg.v27.i43.7509
Liver injury changes the biological characters of serum small extracellular vesicles and reprograms hepatic macrophages in mice
Xiu-Fang Lv, An-Qi Zhang, Wei-Qi Liu, Min Zhao, Jing Li, Li He, Li Cheng, Yu-Feng Sun, Gang Qin, Peng Lu, Yu-Hua Ji, Ju-Ling Ji
Xiu-Fang Lv, An-Qi Zhang, Wei-Qi Liu, Min Zhao, Jing Li, Li He, Li Cheng, Yu-Feng Sun, Peng Lu, Ju-Ling Ji, Department of Pathology, Medical School of Nantong University, Nantong 226001, Jiangsu Province, China
Xiu-Fang Lv, An-Qi Zhang, Wei-Qi Liu, Min Zhao, Yu-Feng Sun, Peng Lu, Ju-Ling Ji, Key Laboratory of Microenvironment and Translational Cancer Research, Science and Technology Bureau of Nantong City, Nantong 226001, Jiangsu Province, China
Jing Li, Department of Pathology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
Li He, Department of Pathology, Nanjing Jinling Hospital, Nanjing University School of Medicine, Nanjing 210093, Jiangsu Province, China
Li Cheng, Department of Pathology, Affiliated Jiangyin Hospital of Southeast University Medical College, Jiangyin 214400, Jiangsu Province, China
Gang Qin, Nantong Institute of Liver Diseases, Nantong Third People’s Hospital, Nantong University, Nantong 226006, Jiangsu Province, China
Yu-Hua Ji, Key Laboratory of Neuroregeneration of Jiangsu and Ministry of Education, Nantong University, Nantong 226001, Jiangsu Province, China
Yu-Hua Ji, Institute of Immunology, College of Life Science and Technology, Jinan University, Guangzhou 510632, Guangdong Province, China
Author contributions: Ji JL and Ji YH conceived the study and interpreted the data; Lv XF performed a systematic review and statistical analysis; Zhang AQ performed the Western blot analysis; Liu WQ performed qRT-PCR; Zhao M isolated the serum sEVs and performed the characterization; Li J, He L and Lu P performed the mouse model experiments; Cheng L performed the mouse model and the primary mouse hepatic macrophage-related experiments; Sun YF performed the histopathologic examination and the sEV characterization; Qin G performed the liver function test; Ji JL and Lv XF prepared the figures and drafted the manuscript; All authors read and approved the final version of the manuscript.
Supported by National Natural Science Foundation of China, No. 81761128018, No. 81572871 and No. 81272027; Natural Science Foundation of Jiangsu Province, No. BK20151277; and Science and Technology Project of Nantong City, No. MS12020010.
Institutional review board statement: This study was reviewed and approved by the Nantong University Medical School Ethics Committee (License No. NTUMEC2017-20).
Institutional animal care and use committee statement: All experimental protocols were approved by the Animal Ethics Committee of Nantong University (License No. S20170102-011). The animal care and experiments were performed in accordance with the relevant guidelines and regulations.
Conflict-of-interest statement: The authors declare no competing interests.
Data sharing statement: All data used for the present study are presented in the main manuscript and its supplementary information files.
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: Ju-Ling Ji, MD, PhD, Director, Professor, Department of Pathology, Medical School of Nantong University, No. 19 Qixiu Road, Nantong 226001, Jiangsu Province, China.
Received: April 21, 2021
Peer-review started: April 21, 2021
First decision: July 14, 2021
Revised: July 21, 2021
Accepted: September 14, 2021
Article in press: September 14, 2021
Published online: November 21, 2021

Serum small extracellular vesicles (sEVs) and their small RNA (sRNA) cargoes could be promising biomarkers for the diagnosis of liver injury. However, the dynamic changes in serum sEVs and their sRNA components during liver injury have not been well characterized. Given that hepatic macrophages can quickly clear intravenously injected sEVs, the effect of liver injury-related serum sEVs on hepatic macrophages deserves to be explored.


To identify the characteristics of serum sEVs and the sRNAs during liver injury and explore their effects on hepatic macrophages.


To identify serum sEV biomarkers for liver injury, we established a CCL4-induced mouse liver injury model in C57BL/6 mice to simulate acute liver injury (ALI), chronic liver injury (CLI) and recovery. Serum sEVs were obtained and characterized by transmission electron microscopy and nanoparticle tracking analysis. Serum sEV sRNAs were profiled by sRNA sequencing. Differentially expressed microRNAs (miRNAs) were compared to mouse liver-enriched miRNAs and previously reported circulating miRNAs related to human liver diseases. The biological significance was evaluated by Ingenuity Pathway Analysis of altered sEV miRNAs and conditioned cultures of ALI serum sEVs with primary hepatic macrophages.


We found that both ALI and CLI changed the concentration and morphology of serum sEVs. The proportion of serum sEV miRNAs increased upon liver injury, with the liver as the primary contributor. The altered serum sEV miRNAs based on mouse studies were consistent with human liver disease-related circulating miRNAs. We established serum sEV miRNA signatures for ALI and CLI and a panel of miRNAs (miR-122-5p, miR-192-5p, and miR-22-3p) as a common marker for liver injury. The differential serum sEV miRNAs in ALI contributed mainly to liver steatosis and inflammation, while those in CLI contributed primarily to hepatocellular carcinoma and hyperplasia. ALI serum sEVs decreased both CD86 and CD206 expression in monocyte-derived macrophages but increased CD206 expression in resident macrophages in vitro.


Serum sEVs acquired different concentrations, sizes, morphologies and sRNA contents upon liver injury and could change the phenotype of liver macrophages. Serum sEVs therefore have good diagnostic and therapeutic potential for liver injury.

Keywords: MicroRNA, Small RNA sequencing, Biomarker, Monocyte-derived macrophage, Resident macrophage

Core Tip: The liver injury changed the concentration, morphology and small RNA contents of serum small extracellular vesicles (sEVs). Altered serum sEV microRNAs (miRNAs) based on mouse studies were highly consistent with the circulating miRNAs reported in human liver diseases. Serum sEV miRNA signatures for acute liver injury and chronic liver injury and a panel of miRNAs that can be used as a common marker for liver injury were established. Acute liver injury serum sEVs depolarized monocyte-derived macrophages and educated resident liver macrophages to transform into M2-like cells. Serum sEVs have good diagnostic and therapeutic potential for liver injury.