Proteases influence various leukocyte effector functions. We investigated the specific binding activity of neutrophil elastase (NE) on rat peritoneal macrophages to stimulate chemokine production in vitro. NE enhanced macrophage production of monocyte chemo-attractant protein-1 (MCP-1) (CC-chemokine) and cytokine-induced neutrophil chemo-attractant (CINC) (CXC-chemokine). However, the serine protease inhibitor, phenylmethylsulfonyl fluoride (PMSF), significantly reduced these chemokine productions by macrophages stimulated with NE. 125I-NE was significantly bound to macrophages, but 125I-NE binding was inhibited by addition of unlabeled NE. In addition, NE significantly stimulated [3H]thymidine incorporation into DNA on 3T3 fibroblasts in a dose-dependent manner. These results suggest that NE stimulates chemokine production by macrophages. This may be a receptor-mediated reaction. Thus, NE not only degrades extra-cellular matrix but also stimulates chemokine production by macrophages.