Basic Study
Copyright ©The Author(s) 2025. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jul 21, 2025; 31(27): 106166
Published online Jul 21, 2025. doi: 10.3748/wjg.v31.i27.106166
Myeloperoxidase, extracellular DNA and neutrophil extracellular trap formation in the animal models of metabolic dysfunction-associated steatotic liver disease
Andrej Feješ, Paulína Belvončíková, Emil Bečka, Tomáš Strečanský, Michal Pastorek, Jakub Janko, Barbora Filová, Pavel Babál, Katarína Šebeková, Veronika Borbélyová, Roman Gardlík
Andrej Feješ, Paulína Belvončíková, Emil Bečka, Tomáš Strečanský, Michal Pastorek, Jakub Janko, Katarína Šebeková, Veronika Borbélyová, Roman Gardlík, Institute of Molecular Biomedicine, Comenius University Faculty of Medicine, Bratislava 81108, Slovakia
Barbora Filová, Institute of Histology and Embryology, Comenius University Faculty of Medicine, Bratislava 81108, Slovakia
Pavel Babál, Institute of Pathological Anatomy, Comenius University Faculty of Medicine, Bratislava 81108, Slovakia
Author contributions: Belvončíková P, Feješ A and Gardlík R conceived and designed research; Belvončíková P, Feješ A, Bečka E, Strečanský T, Janko J, Filová B and Pastorek M performed experiments; Bečka E, Strečanský T and Šebeková K analyzed data; Šebeková K, Babál P, Pastorek M and Gardlík R interpreted results of experiments; Feješ A prepared figures; Feješ A and Belvončíková P drafted manuscript; Babál P, Borbélyová V and Gardlík R edited and revised manuscript; Gardlík R approved final version of manuscript; All authors have read and approved the final manuscript.
Supported by Slovak Research and Development Agency, No. APVV-21-0370; and Ministry of Education, Science, Research and Sport of the Slovak Republic, No. VEGA 1/0706/25 and No. VEGA 1/0341/23.
Institutional review board statement: The study was reviewed and approved by the Institute of Pathophysiology, Faculty of Medicine, Comenius University Institutional Review Board (SK U 06021 Approval No. 07/2023).
Institutional animal care and use committee statement: All procedures involving animals were reviewed and approved by the Institutional Animal Care and Use Committee of the State Veterinary and Food Administration of the Slovak Republic (IACUC protocol number: 4503-3/2024-220).
Conflict-of-interest statement: The authors declare no conflicts of interest.
ARRIVE guidelines statement: The authors have read the ARRIVE Guidelines, and the manuscript was prepared and revised according to the ARRIVE Guidelines.
Data sharing statement: Technical appendix, statistical code, and dataset available from the corresponding author at roman.gardlik@fmed.uniba.sk. No additional data are available.
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: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Roman Gardlík, MD, PhD, Associate Professor, Head, Institute of Molecular Biomedicine, Comenius University Faculty of Medicine, Sasinkova 4, Bratislava 81108, Slovakia. roman.gardlik@fmed.uniba.sk
Received: February 25, 2025
Revised: June 12, 2025
Accepted: June 16, 2025
Published online: July 21, 2025
Processing time: 146 Days and 21 Hours
Abstract
BACKGROUND

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a prevalent chronic liver disorder driven by obesity and metabolic dysfunction. MASLD progresses to metabolic dysfunction-associated steatohepatitis, which is characterized by inflammation, hepatocyte injury, and fibrosis, increasing the risk of cirrhosis and liver failure. Recent studies suggest that neutrophil extracellular traps (NETs) and extracellular DNA (ecDNA) contribute to liver inflammation and fibrogenesis. However, their role in MASLD pathogenesis remains incompletely understood.

AIM

To investigate the dynamics of circulating NETs and ecDNA as potential biomarkers of liver injury in MASLD.

METHODS

Using three complementary mouse models, thioacetamide (TAA)-induced fibrosis, choline-deficient L-amino acid-defined (CDAA) diet-induced metabolic dysfunction-associated steatohepatitis, and cafeteria (CAF) diet-induced MASLD, we assessed the association between NET-related markers and liver damage. Blood samples were collected biweekly to analyze ecDNA and NET markers, including myeloperoxidase (MPO) and MPO-DNA complexes, using ELISA and real-time PCR. Liver histopathology was assessed for inflammation, fibrosis, and neutrophil infiltration.

RESULTS

The TAA and CDAA models exhibited significant liver injury, characterized by increased plasma alanine aminotransferase and aspartate aminotransferase levels, hepatocellular damage, and fibrosis. Elevated circulating NET markers (MPO and ecDNA) were observed in these models, with a strong correlation between NET formation and liver pathology. The CAF diet model induced steatosis but failed to elicit significant liver fibrosis or an increase in NET markers, suggesting that NETosis is associated with more severe liver damage. Notably, ecDNA and MPO levels correlated with neutrophil infiltration and fibrosis scores, indicating their potential as biomarkers of MASLD progression.

CONCLUSION

NETosis and ecDNA levels reflect liver injury severity in MASLD. NET markers and liver fibrosis were strongly associated in TAA and CDAA models, whereas CAF model showed minimal NET involvement.

Keywords: Cell-free DNA; Mouse models; Metabolic dysfunction-associated steatotic liver disease; Neutrophil extracellular traps; Steatohepatitis; Myeloperoxidase; Neutrophil elastase

Core Tip: Metabolic dysfunction-associated steatotic liver disease (MASLD) is a growing global health concern with limited therapeutic options. This study demonstrated that extracellular DNA (ecDNA) and neutrophil extracellular traps (NETs) serve as potential biomarkers of liver injury severity in MASLD. Using three distinct animal models, we showed that NET markers, including myeloperoxidase and ecDNA, correlated with liver fibrosis and inflammation. These findings highlighted NETs and ecDNA as promising targets for early diagnosis and intervention in MASLD, offering new insights into disease progression and potential therapeutic strategies.