Untargeted Metabolomic Profiling Reveals a Distinct Intestinal Metabolic Signature in Necrotizing Enterocolitis

Necrotizing enterocolitis (NEC) is a devastating gastrointestinal emergency affecting up to 10% of infants < 1500 g. Although prior metabolomic studies have profiled metabolite derangements, few have focused on intestinal tissue, the primary site of injury. In this study, we performed metabolomic profiling of intestinal tissue from patients with NEC and surgical controls to identify metabolic alterations associated with NEC. Tissue was collected during surgery from twenty-one infants (8 controls, 13 NEC). Samples underwent liquid chromatography-tandem mass spectrometry. Raw data were extracted, peak-identified, and quality-control-processed, and compounds were identified by comparison with a purified standards library. Following normalization and log transformation, a mixed-effects model accounting for repeated samples from individual infants was used to identify metabolites that differed between NEC and control tissue. There were no significant differences in sex, gestational age, days of life, antibiotic use, or type of feeds between groups. Principal component analysis and hierarchical clustering demonstrated partial separation of NEC and control samples. Metabolomic profiling identified 1183 biochemical features, with 308 demonstrating nominal differential abundance between NEC and control tissue. Indole-related metabolites were increased in NEC tissue, including the nominally significant 3-indoxyl sulfate, a host-microbial co-metabolite associated with epithelial barrier dysfunction. Caffeine-related metabolites were also increased in NEC tissue, while docosahexaenoate demonstrated a trend toward increased relative abundance that did not reach statistical significance. Enrichment of retinol metabolism was observed in NEC tissue, suggesting alterations in retinoid-associated metabolic pathways. Hydroxybutyrate demonstrated a nonsignificant trend toward decreased abundance in NEC. Overall, NEC intestinal tissue demonstrated broad metabolic alterations compared with surgical controls, including differences in lipid, retinol, and host-microbial metabolic pathways. These findings characterize the intestinal metabolic environment associated with NEC and highlight the value of tissue-based metabolomics for understanding disease-associated intestinal alterations.