Cytokines affect pseudomonas binding to tracheal cells via a neutrophil-mediated process.

Critical illness is often associated with gram-negative bacterial colonization of the airways, increasing the risk of nosocomial pneumonia. Cytokines, released in response to endotoxin, might contribute to this phenomenon by causing changes in epithelial cell binding of bacteria. To investigate this possibility, human monocytes and hamster pulmonary macrophages were cultured without or with Escherichia coli endotoxin (10 micrograms/ml) for 4 and 24 h. Hamster and human tracheal epithelial cells were treated with supernates from monocyte cultures for 24 h, and subsequent binding of 14C-labeled Pseudomonas aeruginosa to the epithelial cells was measured (percent adherence). In separate experiments, recombinant human (rh) tumor necrosis factor-alpha (TNF-alpha) (25 to 100 ng/ml) and interleukin-1 beta (IL-1 beta) (2,000 to 8,000 pg/ml) were added to hamster monolayers. Neither monocyte supernates nor purified cytokines were toxic to the epithelial cells for up to 48 h. There was no significant change in P. aeruginosa adherence to either hamster or human tracheal epithelial cells after 24 h of exposure to culture supernates from either endotoxin-stimulated human monocytes or hamster macrophages. Similarly, purified rhTNF and rhIL-1 exposure did not increase bacterial adherence. However, when polymorphonuclear leukocytes were coincubated with the monocyte supernates and epithelial cells, P. aeruginosa adherence was significantly increased. Moreover, this effect was enhanced by an epithelial cell-derived substance. Thus, while inflammatory cytokines may participate in enhancing bacterial colonization of the lung in vivo, they do not do so by a direct action on tracheal epithelial cells but can act via a neutrophil-dependent mechanism.

Characterization of Pseudomonas aeruginosa adherence to cultured hamster tracheal epithelial cells.

This study reports an in vitro system that allows the convenient study of both microenvironmental and bacterial factors affecting adherence of Pseudomonas aeruginosa to tracheal epithelium. Primary cultures of mixed ciliated and nonciliated epithelial cells isolated from hamster tracheas were grown on collagen-coated multiwell plates containing 10(5) epithelial cells/well at confluence. When 10(7) 14C-labeled P. aeruginosa (nonmucoid, strain Y-4) suspensions were added to each well, 8.13 +/- 2.6% (mean +/- SD) of the initial inoculum bound to the cultured cells, an amount comparable to that measured using suspensions of human tracheal epithelial cells and the same bacteria. The bacteria adhered preferentially to the cultured cells rather than to an acellular collagen matrix. Five additional nonmucoid strains of P. aeruginosa also bound well to the cultured cells, while two mucoid strains were less adherent. Strains of two other gram-negative bacteria, Pseudomonas maltophilia and Klebsiella pneumoniae, did not bind significantly, emphasizing the bacterial species specificity of the adherence interaction being measured. The binding interaction with P. aeruginosa was both pH-sensitive and altered by the presence of the divalent cation calcium. Thus, the in vitro assay system described provides a consistent surface of tracheal epithelial cells that binds P. aeruginosa in a specific manner and can be used to examine the effects of bacterial variables and microenvironmental conditions that may be present in the human airway.