P0030Fecal Lipidomics Identifies Lactosylceramide as a Disease-Associated Glycosphingolipid with Diagnostic and Mechanistic Relevance in Crohn’s Disease

Abstract Background Lactosylceramide (LacCer) is a bioactive glycosphingolipid enriched in plasma membrane lipid rafts and implicated in immune activation and epithelial injury. Despite its central role in glycosphingolipid signaling, LacCer has not been systematically investigated in inflammatory bowel disease (IBD). This study aimed to characterise fecal lipidomic alterations in treatment-naïve Crohn’s disease (CD), determine the clinical relevance of LacCer, and explore glycosphingolipid metabolic pathways underlying its dysregulation. Methods A total of 148 newly diagnosed, treatment-naïve CD patients and 77 healthy controls (HC) were enrolled. Targeted fecal lipidomics quantified 560 lipid species. Differential lipids were identified using PCA, univariate and multivariate analyses (P 0.05, |log2FC| ≥ 0, VIP 1), followed by feature selection using Boruta, Random Forest (RF), and Support Vector Machine (SVM). Clinical correlations were evaluated with disease location, behavior, perianal involvement, and inflammatory biomarkers. Expression of key LacCer-related glycosphingolipid enzymes (B4GALT5/6, ST3GAL5, A4GALT, B4GALNT1, B3GNT5) was analyzed using host transcriptomics. Results Among 560 detected lipids, 308 were significantly upregulated and 103 downregulated in CD. Multi-model integration identified 186 candidate biomarkers, with LacCer being the most consistently elevated lipid species across all statistical and machine-learning analyses. Fecal LacCer levels were markedly higher in CD vs HC (P 0.0001). Clinically, LacCer was significantly elevated in patients with ileocolonic disease (L3) compared to isolated ileal disease (L1), and in those with perianal manifestations. LacCer correlated with CRP, ESR and SAA (Spearman r 0.4, all P 0.0001), but not with disease behaviour or CDAI activity. Transcriptomics revealed upregulation of LacCer-synthetic enzymes B4GALT5 (P 0.0001) and B4GALT6 (P 0.0001), alongside an increased B4GALT5/B4GALT6 ratio, suggesting enhanced LacCer biosynthetic flux.Downstream catabolic enzymes were minimally altered, indicating accumulation primarily driven by increased synthesis. Conclusion This study identifies LacCer as a robust fecal biomarker of Crohn’s disease with clear diagnostic, phenotypic, and inflammatory relevance. The elevation of LacCer is mechanistically supported by transcriptional activation of its biosynthetic pathway, highlighting glycosphingolipid metabolism as an overlooked contributor to CD pathogenesis. LacCer holds promise for non-invasive diagnosis, phenotype stratification, and inflammation assessment. Further mechanistic studies and therapeutic targeting of LacCer metabolism warrant investigation. Reference: Salihovic S, Nyström N, Mathisen CB, et al. Identification and validation of a blood- based diagnostic lipidomic signature of pediatric inflammatory bowel disease. Nat Commun. 2024;15(1):4567. Published 2024 Jun 3. doi:10.1038/s41467-024-48763-7 Conflict of interest: Ms. Lai, Wenhua: No conflict of interest Zhu, Xiaochen: No conflict of interest Yi, Youcai: No conflict of interest Zhu, Yijun: No conflict of interest Hu, Shixian: No conflict of interest Mao, Ren: No conflict of interest Chen, Minhu: No conflict of interest Feng, Rui: No conflict of interest