The continuously remodeled extracellular matrix (ECM) plays a pivotal role in gastrointestinal health and disease, yet its precise functions remain elusive. In this project, we employed laser capture microdissection of cecal mucosa combined with low-input proteomics to investigate ECM remodeling during Salmonella-driven inflammation. To complement this, we probed how fibronectin fiber tension is altered using a mechanosensitive peptide probe. While fibronectin fibers in healthy intestinal tissue are typically stretched, fibronectin fiber relaxation occurred exclusively during late-stage infection at 72 hours and was localized to already existing clusters of infiltrated neutrophils in the cecal mucosa of Salmonella-infected mice.