A translational investigation integrating human intestinal gene expression profiling with rodent cell cultures and neonatal mouse experiments has linked interleukin-1β signaling to lipid metabolic suppression and epithelial barrier disruption in necrotizing enterocolitis. Researchers from Wuxi Maternity and Child Health Care Hospital and Affiliated Children’s Hospital of Jiangnan University demonstrated that inflammatory stress in the intestinal epithelium coincides with suppressed fatty acid oxidation machinery alongside heightened accumulation of tissue lipids and markers of oxidative degradation. By testing targeted interventions, the research team observed that genetic knockdown of the cytokine-encoding gene Il1b and pharmacological activation of peroxisome proliferator-activated receptor alpha via fenofibrate each partially alleviated intestinal tissue damage, preserved tight-junction architecture, and limited lipid metabolic disturbances in experimental settings.

Necrotizing enterocolitis represents one of the most critical gastrointestinal crises encountered in premature infants, manifesting through severe gut inflammation, mucosal tissue death, loss of epithelial barrier integrity, and bacterial infiltration into systemic circulation. Standard understandings of the condition highlight excessive innate immune activation, particularly via Toll-like receptor 4 pathways, as a central mechanism converting the vulnerability of an immature intestine into progressive destruction. However, the intestinal lining does not operate merely as a passive barrier; it serves as a metabolically active interface tasked with nutrient handling, epithelial restitution, and the maintenance of mucosal immunity. The investigators noted that premature infants encounter severe inflammatory challenges in an environment defined by immature lipid processing and restricted antioxidant capacity, suggesting that metabolic failure and inflammatory cascades may exacerbate one another during tissue necrosis.

To investigate the transcriptional networks operating within clinical disease, the authors analyzed public microarray profiles from the NCBI Gene Expression Omnibus dataset GSE46619, comparing bowel samples from five infants diagnosed with necrotizing enterocolitis against four non-inflammatory surgical controls. Differential expression screening identified substantial upregulation of inflammatory mediators, including IL1B, IL1A, CXCL8, AQP9, and SERPINB2, alongside marked downregulation of metabolic and differentiation markers such as ALDOB, CYP3A4, SI, MUC13, and CLCA1. Pathway and functional enrichment analyses highlighted over-representation of differentially expressed genes within inflammatory responses, lipid binding, carboxylic acid metabolic processes, fat digestion and absorption, and peroxisome proliferator-activated receptor signaling cascades.

Further computational profiling utilizing gene set variation analysis revealed that fatty acid metabolism was the most prominently depressed metabolic program in human necrotizing enterocolitis samples, yielding a substantial decline compared to surgical controls with a p-value of 1.09 × 10⁻⁶. Cholesterol homeostasis signatures were also significantly depressed, whereas glycolysis exhibited a nonsignificant downward shift. Notably, correlation analysis across all nine surgical and disease specimens identified a strong inverse association between IL1B expression and fatty acid metabolism enrichment scores, characterized by a Pearson correlation coefficient of -0.93. Gene set enrichment analysis also indicated an exploratory negative enrichment trend for the BIOCARTA peroxisome proliferator-activated receptor alpha pathway, although this specific analysis did not reach conventional false discovery rate significance.

To evaluate these observations under controlled experimental conditions, the researchers challenged rat intestinal epithelial IEC-6 cells with either lipopolysaccharide or recombinant rat interleukin-1β. Both inflammatory exposures significantly diminished the messenger RNA levels of key tight-junction components, namely Tjp1 encoding ZO-1 and Ocln encoding occludin, alongside marked reductions in transepithelial electrical resistance. Simultaneously, inflammatory challenge suppressed transcript and protein levels of peroxisome proliferator-activated receptor alpha and carnitine palmitoyltransferase 1A, the rate-limiting mitochondrial enzyme regulating long-chain fatty acid beta-oxidation. At the same time, cultured cells exhibited elevated concentrations of malondialdehyde, an index of lipid peroxidation, accompanied by elevated total cholesterol and triglyceride content within cellular lysates.

Intervention assays in cell culture demonstrated that counteracting interleukin-1β expression helped rescue epithelial integrity. Applying a targeted small interfering RNA designated siIl1b-3 achieved significant knockdown of Il1b expression in IEC-6 cells. This intervention partially restored the messenger RNA expression of Tjp1, Ocln, Ppara, and Cpt1a, while significantly recovering Occludin and CPT1A protein levels and transepithelial electrical resistance values after lipopolysaccharide challenge. Furthermore, the targeted genetic knockdown substantially reduced malondialdehyde generation, total cellular cholesterol, and triglyceride accumulation, indicating that endogenous interleukin-1β participates actively in inflammatory epithelial barrier breakdown and metabolic disruption.

In parallel cell experiments, the authors assessed the therapeutic potential of fenofibrate, a clinically established peroxisome proliferator-activated receptor alpha agonist. Pretreatment of IEC-6 cells with fifty micromolar fenofibrate prior to exposure to either lipopolysaccharide or recombinant interleukin-1β partially prevented the decline in Tjp1, Ppara, and Cpt1a transcript abundance. Western blot analyses corroborated that fenofibrate preserved Occludin and CPT1A protein expression in the presence of inflammatory stimulants, which correlated with higher transepithelial electrical resistance across cultured cell monolayers. Because fenofibrate can exert broad antioxidant and anti-inflammatory actions, the investigators emphasized that these findings align with a peroxisome proliferator-activated receptor alpha-associated response rather than establishing absolute receptor dependency.

To test these findings in an intact physiological system, the team evaluated a neonatal mouse model using twenty-four male and female C57BL/6 pups distributed across four balanced experimental groups. On postnatal day two, pups received superficial temporal vein injections containing either an adeno-associated virus serotype nine expressing short hairpin RNA targeting murine Il1b or a negative control scrambled vector. Following a five-day viral transduction window, experimental enterocolitis was induced from postnatal day seven to ten through daily intragastric gavage of hypertonic formula combined with ten-minute exposures to five percent oxygen hypoxia twice daily, while a separate diseased cohort received daily oral fenofibrate suspensions at fifty milligrams per kilogram.

Analysis of distal ileal segments collected at postnatal day eleven revealed that the formula and hypoxia regimen provoked severe mucosal disruption, marked villus shortening, crypt distortion, and elevated Caplan histological injury scores. Molecular testing confirmed that disease induction elevated ileal Il1b transcription while depleting Tjp1, Ocln, Ppara, and Cpt1a transcripts alongside Occludin and CPT1A protein abundance. In parallel, intestinal tissue homogenates exhibited pronounced increases in malondialdehyde, total cholesterol, and triglyceride concentrations, accompanied by reduced epithelial occludin immunoreactivity in histological sections.

Both therapeutic interventions conferred measurable tissue protection in the neonatal mice. Targeted in vivo knockdown of Il1b and systemic fenofibrate treatment each independently attenuated histological damage, improved villus length and crypt depth, and lowered cumulative pathological scores. Both interventions partially restored ileal tight-junction transcripts, occludin immunohistochemical staining intensity, and CPT1A protein expression, while significantly curtailing tissue malondialdehyde levels, total cholesterol, and triglyceride accumulation. The researchers noted that these findings delineate a supportive working model in which interleukin-1β-driven inflammation, impaired fatty acid oxidation machinery, and lipid stress converge during epithelial failure.

The study authors carefully outlined important experimental limitations governing the interpretation of their data. The transcriptomic findings were derived from a small, bulk-tissue human cohort that cannot separate epithelial-specific changes from immune cell infiltration or tissue necrosis, and the inverse association observed across disease and control samples might reflect broader group contrasts. Methodologically, RNA integrity numbers were not recorded, and quantitative Western blotting in animal tissues was conducted using three independent biological replicates per group. Furthermore, dynamic metabolic flux, direct mitochondrial adenosine triphosphate production, and receptor-deficient genetic knockouts were not examined, meaning that strict causal linearity between interleukin-1β and peroxisome proliferator-activated receptor alpha, as well as the potential synergy of combined therapies, remains an untested hypothesis.

To preserve disease-specific signals, the bioinformatic workflow excluded spontaneous intestinal perforation cases present in the original GSE46619 microarray dataset, restricting comparisons strictly between five confirmed necrotizing enterocolitis specimens and four non-inflammatory surgical controls. The authors addressed an apparent discrepancy between human transcriptional profiles and experimental rodent tissues regarding lipid biology: while human necrotizing enterocolitis tissues exhibited decreased cholesterol homeostasis pathway scores, cell lysates and murine ileal homogenates showed marked accumulations of total cholesterol. The investigators clarified that gene set variation analysis quantifies messenger RNA expression signatures rather than metabolic flux, whereas biochemical assays reflect physical lipid content extracted directly from cellular and tissue matrices. Additionally, although the study tracked occludin and ZO-1 regulation, other critical tight-junction proteins like claudin-2 and claudin-15 were unmeasured, leaving their roles in paracellular sodium recycling and nutrient absorption uncharacterized.

The experimental mouse protocol also incorporated distinct design parameters, relying solely on intragastric administration of hypertonic formula, composed of a two-to-one mixture of Wyeth Stage 1 infant formula and Abbott Birth Puppy Formula, paired with systemic hypoxia, without introducing exogenous lipopolysaccharide or live bacterial inoculums. All twenty-four neonatal pups completed the protocol across the four balanced cohorts without reaching predefined humane endpoints or experiencing procedure-related mortality. Nevertheless, the researchers cautioned that this acute injury paradigm simulates formula- and hypoxia-induced gut damage rather than true premature gestation or the complex clinical heterogeneity seen in fragile infants. Cross-species differences also warrant consideration, as in vitro evaluations utilized rat-derived IEC-6 epithelial cells whereas living validations were restricted to neonatal mice.

Because fenofibrate possesses documented pleiotropic antioxidant and anti-inflammatory activities independent of nuclear receptor engagement, the authors stressed that the experimental recovery remains a peroxisome proliferator-activated receptor alpha-associated response rather than proven receptor dependency. Furthermore, dynamic parameters including mitochondrial respiration rates, fatty acid oxidation flux, adenosine triphosphate synthesis, and lipid droplet trafficking were not directly quantified in either model. The team emphasized that resolving these mechanistic questions will require future investigations employing epithelial-specific loss-of-function genetics, metabolic flux tracing, neonatal intestinal organoid cultures, and high-resolution spatial transcriptomics in clinical human tissue specimens.

Subject of Research: Pathogenic links between interleukin-1β inflammation, lipid metabolic suppression, and epithelial barrier damage in necrotizing enterocolitis

Article Title: IL-1β-associated inflammation, lipid metabolic alterations and epithelial barrier injury in necrotizing enterocolitis: partial protection by fenofibrate

Article References: Zhang, Y., Zhou, Y., Yu, R., & Mei, Y. (2026). IL-1β-associated inflammation, lipid metabolic alterations and epithelial barrier injury in necrotizing enterocolitis: partial protection by fenofibrate. Frontiers in Physiology, 17, Article 1979880. https://doi.org/10.3389/fphys.2026.1979880

Image Credits: AI Generated

DOI: 10.3389/fphys.2026.1979880

Keywords: necrotizing enterocolitis, interleukin-1 beta, lipid metabolism, fatty acid oxidation, epithelial barrier, fenofibrate, PPAR alpha, CPT1A, tight junctions, occludin, lipid peroxidation, malondialdehyde, neonatal mouse model, IEC-6 cells, transepithelial electrical resistance, GSE46619