Fibroblast-Mediated MMP2 Contribution to Nonresponse in Anti-TNFα Therapy for Crohn’s Disease

Abstract Background Despite the transformative impact of biologics on the treatment of inflammatory bowel disease (IBD) in recent years, a substantial proportion of patients remain unresponsive to these advanced therapies. Overcoming resistance remains a significant clinical challenge, largely due to the incomplete understanding of its underlying mechanisms. This study explores the role of extracellular matrix (ECM) remodeling in the development of resistance to anti-TNFα therapy. Methods Using public single-cell and bulk transcriptomic datasets, combined with in vitro primary cell experiments and multiplex immunofluorescence validation, we investigated key factors contributing to therapy failure. Results Transcriptomic analysis revealed that ECM-related pathways were enriched in Crohn’s disease patients and associated with resistance to anti-TNFα therapy. Single-cell analysis identified Fibroblast 2 (CD81+ fibroblasts) as the major ECM-related stromal subpopulation, exhibiting the highest capacity for ECM secretion and degradation. Matrix metalloproteinase-2 (MMP2) was identified as a key protease highly expressed in this subset, showing close interaction with macrophages. Co-culture of primary fibroblasts and macrophages led to increased release of inflammatory mediators such as TNFα and IL-6, which was partially reduced by MMP2 inhibition, suggesting a potential regulatory role of MMP2 in fibroblast-macrophage crosstalk. Spatial transcriptomics and multiplex immunofluorescence further supported the spatial colocalization and interaction of fibroblasts, macrophages, and MMP2 within the tissue microenvironment. Conclusions This study highlights the association of pathogenic fibroblasts and ECM remodeling in anti-TNFα therapy failure, identifies MMP2 as a potential target, and suggests that combination therapy may offer a potential strategy for patients with treatment resistance.