Remodeling of the bronchial wall is a major determinant of morbidity in asthma. An increased number of myofibroblasts beneath the bronchial epithelial basement membrane has been described in asthma. The production of mediators by epithelial cells in close proximity to myofibroblasts during epithelial repair after repeated damage is one of the possible mechanisms for airway remodeling. In this study, we established a three-dimensional co-culture system in which myofibroblasts derived from human bronchial wall were maintained in collagen gels and a human bronchial epithelial cell line, 16HBE14o-, was grown on the surface of the gels. The epithelial cells were chemically injured by exposure to poly-L-arginine as a surrogate for eosinophil granule cationic protein and the proliferative response of the fibroblasts was examined. Conditioned medium from mechanically damaged epithelial cells was also tested for its effect on fibroblast proliferation. Myofibroblasts in the co-cultures showed significantly enhanced proliferation after poly-L-arginine-induced epithelial damage. Conditioned medium from mechanically damaged epithelial cells also increased fibroblast-proliferation. After epithelial perturbation, basic fibroblast growth factor, platelet-derived growth factor, IGF-1, transforming growth factor-beta2, and endothelin-1 levels increased in culture supernatants. Blockade of these growth factors inhibited fibroblast proliferation by 76% after epithelial injury. This study demonstrates that epithelial cells are an important regulator of airway remodeling by means of paracrine control of bronchial myofibroblasts in response to cell damage and repair.