IDDF2024-ABS-0216 Gli1+ mesenchymal cells participate in crohn’s disease intestinal fibrogenesis through the matrix protein SMOC2

<h3>Background</h3> The activation of mesenchymal cells resulting in substantial extracellular matrix (ECM) deposition emerges as a pivotal factor in its pathogenesis of Crohn’s disease (CD). This study aims to identify potential targets by investigating key Gli1+ mesenchymal cell populations implicated in intestinal fibrogenesis. <h3>Methods</h3> Single-cell RNA sequencing was performed on surgical samples from CD patients, encompassing both nonfibrotic and fibrotic sites, as well as on a chronic colitis mouse model induced by dextran sulfate sodium (DSS) alongside controls. Gli1-CreERT2; R26-tdTomato mice were generated for lineage tracing and chronic colitis modeling, with fibrosis evaluated using flow cytometry, immunofluorescence staining, and other experiments. Cell sorting combined with RNA sequencing was utilized to isolate Gli1+ mesenchymal cells, revealing their pathogenic determinants. <h3>Results</h3> Single-cell RNA sequencing data and immunofluorescence staining from surgical patients demonstrated the enrichment of Gli1+ mesenchymal cells at sites of intestinal fibrosis. (IDDF2024-ABS-0216 Figure 1 (A)) Lineage-traced Gli1+ mesenchymal cells proliferated and acquired a myofibroblast phenotype after chronic colitis. (IDDF2024-ABS-0216 Figure 1 (B)) <i>In vivo,</i> ablation of Gli1+ mesenchymal cells alleviated the severity of intestinal fibrosis. (IDDF2024-ABS-0216 Figure 1 (C)) Compared to Gli1- mesenchymal cells, Gli1+ mesenchymal cells expressed a distinct profile of genes, including genes that are markers of ECM-related genes and Smoc2 (SPARC Related Modular Calcium Binding 2, encodes an extracellular matrix protein). (IDDF2024-ABS-0216 Figure 1 (D)) Elevated Smoc2 expression was confirmed in both humans and mice, with further Smoc2 knockdown substantiating its promotion of intestinal fibrosis pathogenesis. (IDDF2024-ABS-0216 Figure 1 (E)) Furthermore, a predictive and diagnostic model was established by detecting SMOC2 levels in both tissue and plasma samples from CD patients with intestinal fibrosis, coupled with protein structure predictions to identify potential therapeutic small molecule binding sites. (IDDF2024-ABS-0216 Figure 1 (F)) <h3>Conclusions</h3> The significant expansion of Gli1+ mesenchymal cells in fibrotic intestines led to extensive ECM deposition and exacerbated fibrosis. Specifically upregulated in Gli1+ mesenchymal cells, Smoc2 encodes a secreted protein with promising potential as both a biomarker and therapeutic target for intestinal fibrosis.