Several members of the matrix metalloproteinase (MMP) family function in various processes of innate immunity, particularly in controlling leukocyte influx. Epilysin (MMP-28) is expressed in numerous tissues and, in adult mice, it has the highest expression in lung, where it is detected in bronchial epithelial cells (Clara cells). Epilysin is also expressed by bone marrow-derived macrophages, but not by alveolar macrophages, suggesting that its expression by macrophages is dependent on localization and differentiation. To assess the role of this MMP, we generated epilysin-null (Mmp28) mice. Although epilysin is constitutively expressed in normal tissues, Mmp28 mice have no overt phenotype. However, using a murine model of Pseudomonas aeruginosa pneumonia, we found that Mmp28 mice had an early increase in macrophage recruitment into the lungs, as well as enhanced bacterial clearance and reduced pulmonary neutrophilia, which we predicted were due to accelerated macrophage influx. Macrophage depletion in WT and Mmp28 mice confirmed a role for macrophages in clearing P. aeruginosa and regulating neutrophil recruitment. Furthermore, we observed that macrophages derived from Mmp28 mice migrated faster than did wildtype cells to bronchoalveolar lavage fluid from P. aeruginosa-treated mice of either genotype. These observations indicate that epilysin functions as an intrinsic negative regulator of macrophage recruitment by retarding the chemotaxis of these cells. Copyright © 2009 by The American Association of Immunologists, Inc.