Introduction
Neutrophil extracellular traps (NETs) have emerged as important mediators in liver fibrosis, promoting inflammatory responses, activating hepatic stellate cells, and accelerating extracellular matrix accumulation. Meanwhile, anticoagulants have demonstrated potential antifibrotic effects in preclinical models, though the underlying mechanisms remain to be fully elucidated. As coagulation factors are known to influence neutrophil activation and NETs release, it is hypothesized that anticoagulation may indirectly suppress NETs formation and thereby attenuate fibrosis.
Aims & Methods
This study aimed to investigate whether anticoagulant therapy mitigates liver fibrosis through modulation of NETs and to explore the feasibility of combination therapy. Advanced liver fibrosis was induced in mice via intraperitoneal injection of thioacetamide (TAA, 150 mg/kg), three times per week for 12 weeks. Mice were then randomly divided into a treatment group (the factor X antagonist Edoxaban, Edo) and a control group (Ctr) (each n=6). Edo was given as a suspension by gavage (30 mg/kg in 0.2 ml) daily for 3 weeks, while controls received the same volume of saline, from week 10-12 of TAA administration. Liver fibrosis was evaluated by collagen morphometry, RT-qPCR and WB. NETs formation was assessed via RNA sequencing, WB and IHC.
Results
Masson's trichrome and Sirius Red morphometry demonstrated significantly decreased hepatic collagen deposition in the Edo group. RT-qPCR and WB also showed significant downregulation of fibrosis-related genes including Col1a1, Fn and Acta2 in the Edo group. Except for downregulation of fibrosis and inflammation related genes, differential gene expression analysis revealed enrichment in the NETs formation pathway. HE staining of the Edo group showed a significantly reduced neutrophil infiltration and IHC showed lower expression of NETs markers including Myeloperoxidase (MPO) and Neutrophil elastase (NE), and WB showed decreased levels of citrullinated histone H3 (CitH3), another marker of NETs formation, in the Edo vs the control group.
Conclusion
Anticoagulation may alleviate liver fibrosis by targeting neutrophil extracellular traps, highlighting a novel therapeutic target for treating liver fibrosis.
References
1. Guerrero A, Campo LD, Piscaglia F, Scheiner B, Han G, Violi F, et al. Anticoagulation improves survival in patients with cirrhosis and portal vein thrombosis: The IMPORTAL competing-risk meta-analysis. J Hepatol 2023; 79(1):69-78.
2. Babuta M, Morel C, de Carvalho Ribeiro M, Calenda C, Ortega-Ribera M, Thevkar Nagesh P, et al. Neutrophil extracellular traps activate hepatic stellate cells and monocytes via NLRP3 sensing in alcohol-induced acceleration of MASH fibrosis. Gut 2024; 73(11):1854-1869.
3. Xia Y, Wang Y, Xiong Q, He J, Wang H, Islam M, et al. Neutrophil extracellular traps promote MASH fibrosis by metabolic reprogramming of HSC. Hepatology 2025; 81(3):947-961.
4. Ballestri S, Capitelli M, Fontana MC, Arioli D, Romagnoli E, Graziosi C, et al. Direct Oral Anticoagulants in Patients with Liver Disease in the Era of Non-Alcoholic Fatty Liver Disease Global Epidemic: A Narrative Review. Adv Ther 2020; 37(5):1910-1932.
5. Mahmoud NI, Messiha BAS, Abo-Saif AA, Abdel-Bakky MS. Inhibition of activated factor X; a new pathway in ameliorating carbon tetrachloride-induced liver fibrosis in rats. J Biochem Mol Toxicol 2019; 33(5):e22287.