Extracellular matrix-destructive enzymes, like matrix metalloproteinases (MMP), have been recognized in the process of inflammation and tissue remodeling and repair. The affected tissues often contain markedly increased numbers of mast cells. Although mast cells are capable of activating latent collagenase and proMMP, it has so far been unknown whether human mast cells themselves produce and secrete MMP9. In this study, MMP9 production by cord blood-derived cultured human mast cells and HMC-1 human mast cells was examined by reverse-transcriptase PCR, gelatin zymography and Western blot analysis using an antibody against MMP9. Cultured mast cells and HMC-1 cells treated with phorbol 12-myristate 13-acetate were shown to express MMP9 mRNA, and the cultured conditioned media from these cells showed gelatinolytic activity, identical with MMP9. Immunohistochemical examination was performed to detect MMP9 in tissue mast cells; mast cells localized in the skin, lung and synovial tissue showed strongly positive reactions for MMP9. Thus, these findings indicate that human mast cells can produce MMP9, which might contribute to extracellular matrix degradation and absorption in the process of allergic and nonallergic responses.