Interleukin 8-CXCR2-mediated neutrophil extracellular trap formation in biliary atresia associated with neutrophil extracellular trap-induced stellate cell activation

Yuhuan Luo¹, Lisa Fraser², Julia Jezykowski², Nitika A Gupta³, Alexander G Miethke⁴, Sarah A Taylor¹, Estella M Alonso⁵, Simon Horslen⁶, Rohit Kohli⁷, Jean P Molleston⁸, Binita M Kamath⁹, Stephen L Guthery¹⁰, Kathleen M Loomes¹¹, John C Magee¹², Phillip Rosenthal¹³, Pamela Valentino¹⁴, Ronald J Sokol¹, Cara L Mack^{1

2}; Childhood Liver Disease Research Network

Affiliations

¹ Department of Pediatrics, University of Colorado Anschutz, Aurora, Colorado, USA.
² Department of Pediatrics, Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
³ Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, USA.
⁴ Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA.
⁵ Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA.
⁶ Department of Pediatrics, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
⁷ Department of Pediatrics, University of Southern California, Los Angeles, California, USA.
⁸ Department of Pediatrics, James Whitcomb Riley Hospital for Children, Indianapolis, Indiana, USA.
⁹ Department of Pediatrics, University of Toronto, The Hospital for Sick Children, Toronto, Ontario, Canada.
¹⁰ Department of Pediatrics, University of Utah College of Medicine, Salt Lake City, Utah, USA.
¹¹ Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA.
¹² Department of Surgery, University of Michigan, Ann Arbor, Michigan, USA.
¹³ Department of Pediatrics, University of California, San Francisco, California, USA.
¹⁴ Department of Pediatrics, University of Washington School of Medicine, Seattle, Washington, USA.

PMID: 39693274
PMCID: PMC12174471 (available on 2025-12-18)
DOI: 10.1097/HEP.0000000000001195

Interleukin 8-CXCR2-mediated neutrophil extracellular trap formation in biliary atresia associated with neutrophil extracellular trap-induced stellate cell activation

Yuhuan Luo et al. Hepatology. 2025.

. 2025 Sep 1;82(3):552-565.

doi: 10.1097/HEP.0000000000001195. Epub 2024 Dec 18.

Authors

Affiliations

¹ Department of Pediatrics, University of Colorado Anschutz, Aurora, Colorado, USA.
² Department of Pediatrics, Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
³ Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, USA.
⁴ Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA.
⁵ Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA.
⁶ Department of Pediatrics, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
⁷ Department of Pediatrics, University of Southern California, Los Angeles, California, USA.
⁸ Department of Pediatrics, James Whitcomb Riley Hospital for Children, Indianapolis, Indiana, USA.
⁹ Department of Pediatrics, University of Toronto, The Hospital for Sick Children, Toronto, Ontario, Canada.
¹⁰ Department of Pediatrics, University of Utah College of Medicine, Salt Lake City, Utah, USA.
¹¹ Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA.
¹² Department of Surgery, University of Michigan, Ann Arbor, Michigan, USA.
¹³ Department of Pediatrics, University of California, San Francisco, California, USA.
¹⁴ Department of Pediatrics, University of Washington School of Medicine, Seattle, Washington, USA.

PMID: 39693274
PMCID: PMC12174471 (available on 2025-12-18)
DOI: 10.1097/HEP.0000000000001195

Abstract

Background and aims: Biliary atresia (BA) entails an inflammatory sclerosing lesion of the biliary tree, with prominent fibrosis in infancy. Previous studies revealed that neutrophil-activating IL-8 and neutrophil extracellular traps (NETs) positively correlated with bilirubin and the risk of liver transplant. The aims of this study were to determine the mechanism of NET formation (NETosis) in BA and whether NETs induce stellate cell activation.

Approach and results: BA and other liver disease control plasma and tissue were obtained at diagnosis and transplant. Elastase, NETs, and IL-8 were quantified by ELISA for plasma and by immunohistochemistry for liver tissue. FACS analysis of neutrophils co-cultured with BA or control plasma measured BA-specific NETosis. Stellate cell activation from co-culture studies of stellate cells with NETs was measured by real-time quantitative PCR, ELISA, and FACS. Liver neutrophils and NETs, and plasma elastase, NETs, and IL-8, were significantly increased in BA at diagnosis and transplant. Normal neutrophils co-cultured with BA plasma had increased NETosis and activation of CXCR2, an IL-8 receptor; CXCR2 inhibition decreased NET production. Immunohistochemistry identified increased NET expression of profibrogenic tissue factor and IL-17. NETs co-cultured with stellate cells resulted in stellate cell activation based on increased ACTA2 and COL1A1 mRNA, collagen protein, and cell surface expression of actin, collagen1A, and platelet-derived growth factor receptor-beta.

Conclusions: Patients with BA have persistent IL-8-CXCR2-mediated NETosis that correlates with biomarkers of injury and fibrosis, and NETs induce stellate cell activation, suggesting a role for NETs in the immunopathogenesis of disease. Future investigations should focus on therapeutic agents that inhibit NETs in BA.

Keywords: cholangiopathy; fibrogenesis; innate immunity.

PubMed Disclaimer

Conflict of interest statement

Conflicts of Interest: Nitika A. Gupta advises Mallinkrodt, Albireo, Chronic Liver Disease Foundation, and Alexion. Alexander G. Miethke consults for and received grants from Mirum. Sarah A. Taylor consults for Ipsen and Mirum. Simon Horslen advises Albireo. He received grants from Mirum. Rohit Kohli consults for and advises Mirum and Ipsen. He advises Sanofi. He received grants from Epigen. Jean P. Molleston received grants from CF Foundation, Mirum, Albireo, AbbVie, and Gilead. Binita M. Kamath consults for and received grants from Ipsen.

Kathleen M. Loomes consults for and received grants from Albireo and Mirum. Philip Rosenthal consults for Mirum, Takeda, and Taysha. He received grants from Gilead. Pamela Valentino advises and is on the speakers’ bureau for Mirum. She advises Grifols. The remaining authors have no conflicts to report.

References

1. Sundaram SS, Mack CL, Feldman AG, Sokol RJ. Biliary atresia: Indications and timing of liver transplantation and optimization of pretransplant care. Liver Transpl. 2017;23(1):96–109. - PMC - PubMed
1. Bezerra JA, Wells RG, Mack CL, et al. BILIARY ATRESIA: Clinical and Research Challenges for the 21(st) Century. Hepatology. 2018;68(3):1163–1173. - PMC - PubMed
1. Mack CL, Tucker RM, Lu BR, et al. Cellular and humoral autoimmunity directed at bile duct epithelia in murine biliary atresia. Hepatology. 2006;44(5):1231–1239. - PMC - PubMed
1. Mack CL, Falta MT, Sullivan AK, et al. Oligoclonal expansions of CD4+ and CD8+ T-cells in the target organ of patients with biliary atresia. Gastroenterology. 2007;133(1):278–287. - PMC - PubMed
1. Lages CS, Simmons J, Maddox A, et al. The dendritic cell-T helper 17-macrophage axis controls cholangiocyte injury and disease progression in murine and human biliary atresia. Hepatology. 2017;65(1):174–188. - PMC - PubMed

MeSH terms

Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions

Save citation to file

Email citation

Add to Collections

Add to My Bibliography

Your saved search

Create a file for external citation management software

Your RSS Feed

Interleukin 8-CXCR2-mediated neutrophil extracellular trap formation in biliary atresia associated with neutrophil extracellular trap-induced stellate cell activation

Affiliations

Interleukin 8-CXCR2-mediated neutrophil extracellular trap formation in biliary atresia associated with neutrophil extracellular trap-induced stellate cell activation

Authors

Affiliations

Abstract

Conflict of interest statement

References

MeSH terms

Substances

Grants and funding

LinkOut - more resources

Full Text Sources

Miscellaneous