Pseudomonas-induced neutrophil recruitment in the dog airway in vivo is mediated in part by IL-8 and inhibited by a leumedin.

A bacteria-free supernatant of Pseudomonas aeruginosa induces the production of neutrophil chemotactic activity in human bronchial epithelial cells in vitro that is due to the potent chemotactic factor, interleukin-8 (IL-8). Because P. aeruginosa supernatant itself is not chemotactic, we hypothesized that intratracheal P. aeruginosa induces the production of neutrophil chemotactic factors, including IL-8, in vivo. Because neutrophils play a key role in cystic fibrosis, inhibition of neutrophil recruitment might be therapeutic. We studied the effect of P. aeruginosa supernatant in the isolated tracheal segment of dogs in vivo, and we measured neutrophil chemotactic activity in vitro in the tracheal fluid. We also determined the local effect of intratracheal administration of leumedin NPC 15669, an inhibitor of neutrophil recruitment, on IL-8- and Pseudomonas-induced neutrophil accumulation. P. aeruginosa supernatant and IL-8 both caused time-dependent accumulation of neutrophils in the tracheal fluid. Tracheal fluid obtained after P. aeruginosa administration had neutrophil chemotactic activity in vitro that was significantly inhibited by the IL-8 antibody. Intratracheal NPC 15669 prevented both IL-8- and Pseudomonas-induced accumulation of neutrophils. We conclude that P. aeruginosa supernatant recruits neutrophils into the airway indirectly by inducing the production of chemotactic factors, including IL-8. Our results suggest a potential therapeutic role for leumedins in chronic airway diseases.

Leumedin NPC 15669 inhibits antigen-induced recruitment of inflammatory cells into the canine airways.

We studied the effects of NPC 15669, a member of a new class of anti-inflammatory drugs called leumedins, on chemotaxis of both human eosinophils and neutrophils and on Mac-1 receptor upregulation on stimulated eosinophils in vitro. Then, we examined the effect of NPC 15669 on antigen-induced eosinophil and neutrophil recruitment and subsequent airway hypersecretion (indicated by an increase in lysozyme concentration) in the allergic dog trachea in vivo. NPC 15669 inhibited eosinophil chemotaxis in vitro at a drug concentration of 10(-5) M (mean inhibition, 48.2%) without affecting Mac-1 receptor upregulation on stimulated eosinophils. NPC 15669 also inhibited neutrophil chemotaxis: at 10(-5) M, NPC 15669 inhibited neutrophil chemotaxis by a mean of 29.7%. In allergic dogs in vivo, NPC 15669 (10(-5) M) prevented antigen-induced recruitment of eosinophils and neutrophils and prevented the increase in elastase and lysozyme concentrations. We conclude that NPC 15669 is an effective inhibitor of antigen-induced leukocyte recruitment and elastase release and subsequent hypersecretion in airways.

Interleukin-8: an important chemoattractant in sputum of patients with chronic inflammatory airway diseases.

Sputum from patients with cystic fibrosis, bronchiectasis, and chronic bronchitis contains neutrophils and neutrophil proteases, which have been implicated in the pathophysiology of mucus hypersecretion in airways. We asked whether interleukin-8 (IL-8), a potent neutrophil chemoattractant, might be involved in recruiting neutrophils into airways of patients with cystic fibrosis, bronchiectasis, and chronic bronchitis. We found significant neutrophil chemotactic activity in sputum obtained from these patients. The IL-8 concentrations that we measured in sputum of patients with cystic fibrosis (7.1 +/- 1.0 x 10(-9) M, mean +/- SE), bronchiectasis (9.6 +/- 2.9 x 10(-9) M), and chronic bronchitis (2.8 +/- 1.0 x 10(-9) M) have been reported to cause significant chemotaxis in vitro and in airways in vivo, whereas concentrations measured in induced sputum from healthy subjects (1.1 +/- 0.3 x 10(-10) M) do not. A monoclonal antibody to IL-8 significantly inhibited the chemotactic activity in patients' sputum by 75-98%, but not in induced sputum from healthy subjects (9%). We conclude that IL-8 is an important chemotactic factor in sputum of patients with cystic fibrosis, bronchiectasis, and chronic bronchitis, and we suggest that IL-8 accounts, at least in part, for neutrophil recruitment into airways of patients with these diseases.

Interleukin-8 induces neutrophil accumulation but not protease secretion in the canine trachea.

The neutrophil enzyme elastase is a potent secretagogue of airway secretory cells, and elastase is present in high concentrations in sputum of patients with hypersecretion (e.g., cystic fibrosis, bronchiectasis). Interleukin-8 (IL-8), a recently discovered cytokine with potent neutrophil chemotactic properties in vitro, is also found in the sputum of these patients. We used an isolated tracheal segment in dogs in vivo to study the effect of IL-8 in causing neutrophil accumulation, elastase release, and secretion (by measuring lysozyme concentrations) in the luminal superfusate. IL-8 caused a potent time-dependent neutrophil accumulation at between 3 and 6 h. The effect was significant at 10(-9) and maximum at 10(-8) M. No increase in free elastase, cathepsin G, or lysozyme was detected in the superfusate. Thus, in contrast to previous studies showing that ragweed antigen causes the accumulation of neutrophil elastase which in turn causes lysozyme secretion, IL-8 causes neutrophil accumulation without granule secretion (or subsequent secretagogue activity). The findings were confirmed with dog and human neutrophils in vitro.