Tracheal aspirate IL-8 concentration and airway epithelial cell IL-8 expression are each increased in premature infants undergoing mechanical ventilation. We sought to determine the cytokines responsible for IL-8 expression in this context. Tracheal aspirates were collected from 18 mechanically ventilated premature infants. IL-8 protein abundance was high in tracheal aspirates from ventilated premature infants (mean, 5806 Ϯ 4923 pg/mL). IL-1␣ (mean, 20 Ϯ 6 pg/mL), IL-1␤ (mean 67 Ϯ 46 pg/mL), and tumor necrosis factor (TNF)-␣ (mean, 8 Ϯ 2 pg/mL) were also found. Incubation of tracheal aspirates with 16HBE14o-human bronchial epithelial cells increased IL-8 protein in both cell lysates and supernatants, as well as transcription from the IL-8 promoter. Aspirates also induced nuclear factor (NF)-B activation. Mutation of the IL-8 promoter NF-B site abolished aspirate-induced IL-8 transcription. Endotoxin concentrations in the tracheal aspirates were negligible and incapable of inducing IL-8 promoter activity. Finally, incubation of tracheal aspirates with a neutralizing antibody against IL-1␤ reduced epithelial cell IL-8 production, whereas neutralizing antibodies against IL-1␣ and TNF-␣ had no effect. We conclude that airway fluid from mechanically ventilated premature infants contains soluble factors capable of inducing airway epithelial cell IL-8 expression via a NF-B-dependent pathway, and that IL-1␤ plays a specific role in this process. Despite advances in neonatal care, BPD continues to be a major cause of morbidity and mortality in premature neonates. Accordingly, interest remains high in the pathogenesis of this chronic lung disease. Recent evidence suggests the importance of inflammation in the pathogenesis of BPD. During the first days of postnatal life, an influx of polymorphonuclear leukocytes and macrophages is seen in the tracheal aspirates of mechanically ventilated premature infants who later develop BPD (1). Elevations of pro-inflammatory cytokines such as IL-8, IL-6, IL-1␤, and TNF-␣ are also seen (2-7). Similar findings are present in animal models of BPD (8) and hyperoxic exposure (9). Of the cytokines implicated in BPD, IL-8 may be of special importance, as it is the most potent neutrophil chemotactic factor.IL-8 may be elaborated by macrophages, T-lymphocytes, neutrophils, and respiratory epithelial cells (4). In a recent study examining IL-8 expression in the lung tissue of neonates with hyaline membrane disease, the majority of cases demonstrated IL-8 immunoreactivity in fetal and neonatal neutrophils, and in almost half of these cases, airway epithelial cell IL-8 expression (10). An earlier study in ventilated preterm infants detected IL-8 in bronchoalveolar macrophages, neutrophils, and exfoliated epithelial cells (6). In newborn rats, hyperoxic exposure induced the expression of cytokineinduced neutrophil chemoattractant-1, an IL-8 homologue, in alveolar macrophages and epithelial cells (9). Numerous studies have shown that cultured airway epithelial cells may produce IL-8 in response to a...