We investigated the effects of G-CSF in a granulocytopenic mouse model of Pseudomonas aeruginosa lung infection. The model was prepared by intratracheal instillation of the bacteria, while granulocytopenia was induced by intraperitoneal injection of 4.0 mg of cyclophosphamide (CPA). There was no difference in the survival rate between G-CSF-treated animals and the normal group, and the number of neutrophils in the blood and lung recovered to normal in the former group. However, the phagocytic and killing activities of neutrophils were lower in G-CSF-treated mice than in controls. Interestingly, the mortality rate increased significantly when anti-TNF-alpha antibody was combined with G-CSF, although it was intermediate between CPA alone and CPA-G-CSF-treated mice. However, the improved mortality was not associated with a change in the number of neutrophils in the circulation and lung. Administration of anti-TNF-alpha antibody resulted in a significant suppression of TNF-alpha in bronchoalveolar lavage fluid and of enhanced alveolar macrophage function (phagocytic and bactericidal activity) against P. aeruginosa in G-CSF-treated granulocytopenic mice. We showed also increased TNF-alpha mRNA expression and TNF-alpha production in vitro using G-CSF-pretreated alveolar macrophages compared with control untreated macrophages. Our results are the first evidence to suggest that G-CSF provides a synergistic protective effect against lethal P. aeruginosa lung infection in the granulocytopenic host. This effect is probably due to enhancement of alveolar macrophage function through endogenous TNF-alpha production, in addition to increasing the number of circulating neutrophils.