Previous studies in our laboratory demonstrated that the synthetic surfactant Exosurf (Burroughs Wellcome Co.) inhibited endotoxin-stimulated cytokine secretion from human alveolar macrophages in vitro. The purpose of the present study was to further characterize the suppressive effects of Exosurf, which consists of dipalmitoylphosphatidylcholine (DPPC), cetyl alcohol (spreading agent), and tyloxapol (nonionic dispersing agent). Suppression was not stimulus specific in that Exosurf also significantly reduced cytokine production elicited by either Staphylococcus aureus or recombinant interleukin-1. Suppression was also mediated by a modified bovine surfactant (Survanta), which, in contrast to Exosurf, contains the surfactant-associated proteins B and C, and several different phospholipids, but no cetyl alcohol or tyloxapol. This suggests that suppression of macrophage cytokines is not specific to Exosurf. Both cell associated and secreted tumor necrosis factor and interleukin-1 were reduced by Exosurf, indicating that Exosurf is not simply blocking cytokine release. At 3 h, cytokine mRNA levels were not different between Exosurf-treated and untreated cells. However, at 8 and 24 h, cytokine mRNA levels were lower in Exosurf-treated cells. The observations that mRNA levels were decreased at 8 and 24 h and that cellular cytokine release was not blocked suggest that Exosurf's effect may in part be pretranslationally mediated. Collectively, these data add to previous work indicating that pulmonary surfactant may play a critical role in reducing inflammatory cytokine production associated with the adult respiratory distress syndrome and similar disorders.