Lung neutrophilia is common to a variety of lung diseases. The production of reactive oxygen and nitrogen species during neutrophil oxidative burst has been associated with protein and DNA damage. Myeloperoxidase (MPO) is an enzyme stored in the azurophilic granula of neutrophils. It is important in host defense because it generates the reactive oxidant hypochlorous acid and has been described to play a role in the activation of neutrophils during extravasation. We hypothesized that MPO contributes directly to the development of acute lung neutrophilia via stimulation of neutrophil extravasation and indirectly to the subsequent production of cytokines and chemokines in the lung. To test this hypothesis, wild-type (WT) and Mpo-/- mice were given a single LPS instillation, after which the development of neutrophil-dominated lung inflammation, oxidative stress, and cytokine and chemokine levels were examined. Mpo-/- mice demonstrated a decreased lung neutrophilia that peaked earlier than neutrophilia in WT mice, which can be explained by decreased neutrophil chemoattractant levels in LPS-exposed Mpo-/- compared with WT mice. However, oxidative stress levels were not different in LPS-exposed WT and Mpo-/- mice. Furthermore, in vivo findings were confirmed by in vitro studies, using isolated neutrophils. These results indicate that MPO promotes the development of lung neutrophilia and indirectly influences subsequent chemokine and cytokine production by other cell types in the lung. Copyright © 2009 by The American Association of Immunologists, Inc.