Abstract Background Intestinal fibrosis, a hallmark complication of Crohn’s disease (CD), frequently progresses to stricture formation and surgical intervention. Fibroblast metabolic reprogramming is important in organ fibrosis. However, its role in intestinal fibrogenesis of CD remain elusive. We aim to explore the metabolic reprogramming of fibroblasts and its upstream regulators during intestinal fibrosis of CD. Methods We collected paired mucosal and submucosal tissue from both stenotic and adjacent non-stenotic regions of the same CD patient. These samples underwent non-targeted metabolome, transcriptome, single-cell RNA sequencing (scRNA-seq), and spatial transcriptomic analysis. Primary human intestinal mucosal fibroblasts (HIMFs) were isolated, and a dextran sulfate sodium (DSS)-induced mouse model of intestinal fibrosis was established for functional validation of screened key metabolites and metabolic enzymes. Finally, we employed an integrated approach including targeted metabolomics, transcriptomics, metabolic flux analysis, Seahorse extracellular flux analysis, ChIP-seq, and ATAC-seq in HIMFs and the mouse intestinal fibrosis model to verify the upstream regulatory mechanisms of the identified metabolites and enzymes. Results Integrated multi-omics analysis revealed that activation of the pentose phosphate pathway (PPP) in fibroblasts is a key metabolic feature of CD-associated intestinal strictures. The PPP metabolite xylulose-5-phosphate (Xu5P) was significantly upregulated in CD stenotic tissues, showed the most prominent differential expression, and strongly correlated with clinical parameters of intestinal stricturing. Both in vitro and in vivo experiments confirmed that Xu5P promotes extracellular matrix synthesis and exacerbates intestinal fibrosis by upregulating collagen transcription via epigenetic mechanisms. Furthermore, the upstream metabolic enzyme xylulokinase (XYLB) was found to activate the PPP, enhance glycolysis, increase NADPH production, and thereby intensify the intestinal fibrotic process. Conclusion This study elucidates that the XYLB/Xu5P axis in fibroblasts exacerbates intestinal fibrosis in CD by activating the pentose phosphate pathway, providing a novel therapeutic target for CD-associated intestinal strictures. Conflict of interest: Dr. Zhou, Longyuan: All authors declare no competing interests. Chen, Minhu: I have received speaker honoraria from Takeda China, Xian Janssen, and AbbVie China, as well as research funding from Takeda (China). Mao, Ren: None
Zhou et al. (Thu,) studied this question.