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Urolithin A attenuates inflammation and enhances barrier integrity in an experimental NEC-in-a-Dish model

Pediatric Research (2026) Cite this article

Abstract

Background and hypothesis

Necrotizing enterocolitis (NEC) is a devastating gastrointestinal disease in preterm infants, characterized by exaggerated inflammation and poorly understood pathogenesis. Aryl hydrocarbon receptor (AhR) signaling has been shown to reduce intestinal inflammation in colitis models. In these models, a microbial-derived AhR ligand, Urolithin A (UroA), has demonstrated anti-inflammatory properties and improved gut barrier function. We hypothesize that UroA attenuates inflammation and enhances epithelial barrier integrity in an in vitro NEC-in-a-Dish model.

Methods

Enteroids derived from human neonatal intestinal tissue were pretreated with 100 µM UroA or 0.1% DMSO (vehicle) before a 6-h incubation with dysbiotic enteric bacteria cultured from an infant with NEC totalis (NEC-in-a-Dish). RNA sequencing was performed and analyzed for pro-inflammatory cytokine expression and pathway enrichment. Epithelial barrier integrity was evaluated using a fluorescein-isothiocyanate-dextran permeability assay on two-dimensional monolayers.

Results

UroA pretreatment decreased pro-inflammatory cytokine expression and enhanced barrier integrity. Transcriptomic analysis showed downregulation of NOD-like receptor, IL-17/Th17, NF-κB, and TLR signaling, and upregulation of AhR-dependent genes involved in antioxidant responses.

Conclusion

UroA reduces inflammation and strengthens epithelial barrier integrity in a NEC-in-a-Dish model through a variety of mechanisms. These findings support further investigation of dietary-derived metabolites as potential preventative strategies for NEC in preterm infants.

Impact

  • Urolithin A (UroA), a microbiota-derived metabolite, reduces inflammation and enhances epithelial barrier integrity in a human neonatal in vitro model of necrotizing enterocolitis (NEC).

  • This study is the first to demonstrate the protective effects of UroA in the neonatal small intestinal epithelium using a NEC-in-a-Dish model system. It also expands on the current understanding of the mechanism of action of UroA by revealing modulation of key inflammatory and oxidative stress pathways in neonatal tissue.

  • These findings identify UroA as a promising candidate for dietary-based prevention strategies targeting epithelial injury in NEC.

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Fig. 1: The NEC-in-a-Dish platform recapitulates hallmark features of human NEC.
The alternative text for this image may have been generated using AI.
Fig. 2: Pretreatment with urolithin A attenuates the inflammatory response in experimental NEC-in-a-Dish.
The alternative text for this image may have been generated using AI.
Fig. 3: Pretreatment with urolithin A enhances intestinal epithelial barrier function.
The alternative text for this image may have been generated using AI.
Fig. 4: RNA sequencing reveals that urolithin A downregulates key inflammatory pathways.
The alternative text for this image may have been generated using AI.
Fig. 5: Urolithin A pretreatment in the NEC-in-a-Dish model did not reveal enrichment in either canonical or non-canonical aryl hydrocarbon receptor (AhR) signaling pathways.
The alternative text for this image may have been generated using AI.

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Data availability

The datasets generated during the current study are available from the corresponding author on reasonable request. The sequencing data can be found at Gene Expression Omnibus (Geo), [https://www.ncbi.nlm.nih.gov/geo/], Series GSE312490.

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Funding

L.C.F. is supported by the National Institutes of Health (NIH) grant K08DK140604 and the UNC School of Medicine Physician Scientist Training Program Faculty Award. M.G. is supported by NIH grants R01DK124614, R01HD105301, R44HD110306, DP1DK140012, the Chan Zuckerberg Initiative Grant number 2022-316749, the Yang Biomedical Scholar Award, and the University of North Carolina at Chapel Hill Department of Pediatrics.

Author information

Authors and Affiliations

  1. Division of Pediatric Gastroenterology, Department of Pediatrics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Ahmad Salah Sami

  2. Division of Neonatal-Perinatal Medicine, Department of Pediatrics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Gergely Mozes, Corey M. Jania, Vikram Puri, Lauren Bolzan, Yukihiro Yamaguchi, Lauren C. Frazer, Stephen Mackay, Natalia S. Akopyants & Misty Good

  3. Bioinformatics and Analytics Research Collaborative (BARC), University of North Carolina, Chapel Hill, NC, USA

    Erika Maldonado-Rosado & Austin J. Hepperla

  4. Department of Genetics, University of North Carolina, Chapel Hill, NC, USA

    Austin J. Hepperla

  5. Neuroscience Center, University of North Carolina, Chapel Hill, NC, USA

    Austin J. Hepperla

  6. Carnegie Mellon University, Pittsburgh, PA, USA

    Nadia Ghetie

  7. Department of Microbiology and Immunology, University of Louisville, Louisville, KY, USA

    Venkatakrishna Jala

  8. James Graham Brown Cancer Center, University of Louisville, Louisville, KY, USA

    Venkatakrishna Jala

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Contributions

Conceptualization: A.S.S., N.S.A., and M.G.; data and sample acquisition: all authors; drafting the manuscript: A.S.S., G.M., E.M.R., A.H., N.S.A., and M.G.; critical manuscript review: all authors; funding acquisition for this study: M.G.

Corresponding author

Correspondence to Misty Good.

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The authors declare no competing interests.

Consent statement

Intestinal tissue was obtained in a de-identified manner from a premature infant in the St. Louis Children’s Hospital Neonatal Intensive Care Unit (NICU) who required surgery for intestinal atresia with waiver of consent after approval by the Washington University in St. Louis School of Medicine Institutional Review Board for discarded tissue.

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Sami, A.S., Mozes, G., Jania, C.M. et al. Urolithin A attenuates inflammation and enhances barrier integrity in an experimental NEC-in-a-Dish model. Pediatr Res (2026). https://doi.org/10.1038/s41390-026-05124-y

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