Preclinical studies with first-generation adenovirus (Ad) vectors administered in vivo to the respiratory tract have demonstrated a nonspecific host response consisting, in part, of parenchymal neutrophil accumulation followed by mononuclear cell and macrophage accumulation. We hypothesized that the mechanism for this host response might be the elaboration of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) from the airway epithelium following the exposure to Ad. To evaluate this hypothesis, we infected A549 cells (a human-derived lung epithelial cell line) in vitro with an adenovirus type 5 (Ad5)-based vector expressing a nuclear targeted beta-galactosidase enzyme (Av1LacZ4). We found that cellular transduction was efficient, resulting in gene delivery to 85.5% +/- 3.9% of the cell monolayer after 96 hr. Importantly, IL-8 mRNA transcript levels in Av1LacZ4-transduced cells were significantly higher than uninfected controls by 24 hr and remained elevated for 96 hr. IL-8 protein secretion from Av1LacZ4-transduced cells was increased for the same period. The Av1LacZ4-transduced A549 cells also showed a neutrophil chemoattractant activity higher than control cells, measurable at 24 hr, and persisting for 96 hr. The chemoattractant activity could be neutralized by a specific monoclonal antibody to IL-8. Whereas Av1LacZ4 transduction induced IL-8 gene expression, there was a lack of expression of MCP-1 by A549 cells. These observations demonstrate that the gene delivery to the airway epithelium using the Ad5-based expression vector results in IL-8 gene activation in these cells, which may contribute to the described inflammatory host response.