P0134CHRDL2 secreted by the intestinal stem cell niche promotes intestinal epithelial repair and alleviates colitis by targeting EGFR in intestinal epithelium.

Abstract Background • IBD is a chronic recurrent gastrointestinal disorder including Crohn’s disease (CD) and ulcerative colitis (UC), with a rapidly increasing incidence in China1. • Its pathogenesis involves intestinal mucosal barrier damage, ISC dysfunction, and ISC niche dysregulation, making mucosal barrier repair a key therapeutic target2. • The ISC niche regulates ISC self-renewal via paracrine signals, and Bone morphogenetic protein (BMP) antagonists have emerged as potential regulators in IBD3-4. • CHRDL2, an ISC-related BMP antagonist, is significantly upregulated in inflamed intestinal tissues of IBD patients and colitis mice. Methods 1. Screened differentially expressed CHRDL2 via transcriptome sequencing of IBD patient biopsies and colitis mouse colon tissues, verifying its mRNA and protein expression. 2. Identified CHRDL2’s secretory sources (intestinal subepithelial myofibroblasts and smooth muscle cells) using RNAscope and cell culture. 3. Evaluated the effects of recombinant CHRDL2 (rCHRDL2) and CHRDL2 knockdown (via AAV) on DSS-induced colitis in mice, and assessed intestinal barrier function (TEER, FD4) and ISC activity (CCK-8, EdU, qPCR) in vitro and in organoids. 4. Validated CHRDL2-EGFR interaction via proteomics, Co-IP MS, and Pulldown, and confirmed the axis using EGFR inhibitors (AG1478, Gefitinib). 5. Explored downstream pathways via database analysis (STRING, GEO, GSEA) and verified the CHRDL2-EGFR-MUC5B/β-catenin axis experimentally. Results 1. CHRDL2 expression is significantly upregulated in intestinal tissues of IBD patients and colitis mice, with increased circulating abundance. 2. CHRDL2 is secreted by myofibroblasts and smooth muscle cells in the ISC niche. 3. rCHRDL2 alleviates DSS-induced mice colitis and protects the mucosal barrier, while CHRDL2 knockdown exacerbates inflammation; CHRDL2 promotes ISC proliferation and regeneration. 4. CHRDL2 binds and activates EGFR, promoting downstream MAPK pathways, and EGFR inhibitors block CHRDL2’s protective effects. 5. CHRDL2 upregulates MUC5B via EGFR phosphorylation, activating the β-catenin pathway to drive impaired ISC proliferation and regeneration. Conclusion CHRDL2 secreted by the ISC niche is a potential serological indicator of IBD. By targeting EGFR to activate the MUC5B/β-catenin pathway, it promotes damaged intestinal epithelial repair via ISC proliferation and regeneration, alleviating colitis and providing important scientific and clinical value for IBD management. References: 1 Honap S, Jairath V, Danese S, et al. Navigating the complexities of drug development for inflammatory bowel disease. Nature reviews Drug discovery. 2024;23(7):546-62. 2 Kraiczy J, McCarthy N, Malagola E, et al. Graded BMP signaling within intestinal crypt architecture directs self-organization of the Wnt-secreting stem cell niche. Cell Stem Cell. 2023;30(4):433-49.e8. 3 Koppens M A J, Davis H, Valbuena G N, et al. Bone Morphogenetic Protein Pathway Antagonism by Grem1 Regulates Epithelial Cell Fate in Intestinal Regeneration. Gastroenterology. 2021; 161, 239-254.e239. 4 McCarthy N, Tie G, Madha S, et al. Smooth muscle contributes to the development and function of a layered intestinal stem cell niche. Dev Cell. 2023;58(7):550-64.e6. Conflict of interest: Ms. Xie, Zhuo: No conflict of interest Huang, Shanshan: No conflict of interest Shenghong, Zhang: No conflict of interest