Role of pyroptosis-related hub genes in necrotizing enterocolitis: a study based on bioinformatics analysis and clinical cohort validation

OBJECTIVE: To characterize the expression patterns and clinical significance of pyroptosis-related key genes in necrotizing enterocolitis (NEC), and to identify potential biomarkers with adjunctive diagnostic value. METHODS: Pyroptosis-related candidate genes associated with NEC were screened through transcriptomic differential expression analysis, weighted gene co-expression network analysis (WGCNA), and immune infiltration analysis, followed by validation in an NEC mouse model and an LPS-induced IEC-6 cell model. Furthermore, a clinical cohort study was conducted to compare serum ADIPOQ (adiponectin) levels and related laboratory parameters, and logistic regression together with receiver operating characteristic (ROC) curve analysis was used to assess its diagnostic performance. RESULTS: Transcriptomic analysis identified 1,693 differentially expressed genes, which were predominantly enriched in metabolism- and immunity-related pathways. By integrating WGCNA and protein-protein interaction (PPI) network analysis, four NEC-associated pyroptosis candidate genes were identified: Ccr5, Tlr9, Lcn2, and Adipoq. Both in vivo and in vitro experiments confirmed that Adipoq and Tlr9 were significantly downregulated in NEC in a time-dependent manner (P < 0.05). Clinical analyses demonstrated that serum ADIPOQ levels were significantly reduced in infants with NEC, showing significant negative correlations with C-reactive protein (CRP), lactate dehydrogenase (LDH), and the Aspartate aminotransferase/Alanine aminotransferase (AST/ALT) ratio, and significant positive correlations with glycocholic acid and total bilirubin (TBIL) (P < 0.05). Multivariable logistic regression analysis revealed that serum ADIPOQ, glycocholic acid, and CRP were independently associated with NEC. ROC curve analysis further indicated that ADIPOQ alone exhibited favorable predictive performance for NEC (AUC = 0.887, 95% CI: 0.806-0.952), while its combination with glycocholic acid and CRP further improved predictive efficacy (AUC = 0.932, 95% CI: 0.870-0.980). CONCLUSION: Adipoq and Tlr9 participate in immune and inflammatory dysregulation in NEC and may be involved in pyroptosis-related pathological processes, representing promising novel targets for the prevention and treatment of NEC.