(P04-025-25) Sulfites Dysregulate Amino Acid Metabolites and Energy Metabolism During Epithelial Differentiation in Pediatric Colon Organoids

Objectives: Mitochondrial metabolism is critical to intestinal epithelial regeneration and epithelial mitochondriopathy is evident in pediatric ulcerative colitis. Dietary sulfite additives are potentially toxic inhibitors of metabolism with a likely disproportionate exposure to children. Hence, we determined whether sulfite exposure impacts human intestinal epithelial metabolism and differentiation at a dose below the recommended limits.

Methods: We obtained biopsy-derived colon epithelial organoids (colonoids) from pediatric patients (n=3/group) with active ulcerative colitis (aUC, Mayo score >1), inactive disease (iUC), and non-IBD controls (control) and exposed them to sodium sulfite (50 µM) followed by 3- or 6-day differentiation. We analyzed the cellular metabolic state using extracellular flux analysis and untargeted metabolomics, and measured lineage-specific gene expression with single-cell RNA sequencing.

Results: Sulfite exposure during colonoid differentiation significantly reduced basal respiration and ATP production (P < 0.05) with enhanced susceptibility in iUC colonoids. Among the key macromolecules vital for cellular homeostasis, untargeted metabolomics revealed a preferential reduction in amino acid metabolites in sulfite-exposed control colonoids. In the Lgr5-rich stem cell clusters, gene ontology (GO) analysis of downregulated genes (Log2FC < -1) in untreated aUC vs control showed significant enrichment of amino acid transport and the signal recognition particle (SRP) which are critical for mRNA translation and maturation of membrane proteins. Upon sulfite exposure, stem cell clusters of control and iUC colonoids mirrored untreated aUC colonoids as GO analysis of downregulated genes showed significant enrichment of SRP genes (Rn7sl1 and Rn7sl2). Among the human colon epithelial lineages in control, only genes in the enteroendocrine cell cluster were significantly dysregulated (P < 0.05,15 up and 10 down), suggesting that epithelial sulfite exposure may impact enteroendocrine cell function.

Conclusions: Using patient-derived colon epithelial organoids, our study suggests that dietary sulfite exposure could have detrimental metabolic implications and dysregulate colon epithelial differentiation.

Funding Sources: 1. The National Institutes of Health
2. Stanford Center for Pediatric IBD and Celiac Disease