Dupuytren’s contracture belongs to a large group of fibrotic diseases that share similar mechanisms but lack effective treatment and prevention options. Unlike fibrotic diseases affecting internal organs such as the heart, lung or liver which are typically diagnosed at later stages when the functions of the organ are irreversibly impaired, DD tissue can be diagnosed early and is readily accessible for scientific research after surgical excision. This enables us to analyse the mechanism of active fibrotic process in the relatively early-stage nodular tissue ex vivo. Here we described the changes in ECM-associated proteome of DD tissue and identified the components that potentially activate or promote the fibrosis in DD. Furthermore, our results indicate that the pathological changes in diseased ECM composition orchestrate a feedback loop through macrophage activation which promotes myofibroblast activation and differentiation.