Effect of interleukin-8 and granulocyte colony-stimulating factor on priming and activation of bovine neutrophils

Neutrophils are important effector cells in innate and acquired immunity, but the magnitude and character of their phagocytic and bactericidal responses depend on cues derived from mediators in the local microenvironment. This study investigated the effect of bovine interleukin-8 (IL-8) and granulocyte colony-stimulating factor (G-CSF) on priming and activation of bovine neutrophils in vitro and in vivo. Neutrophils were isolated from blood and cultured for up to 18 h, with or without cytokines, and then Mannheimia haemolytica-induced oxidative burst and phagocytosis of Staphylococcus aureus were measured by flow cytometry. Neither IL-8 nor G-CSF directly triggered an oxidative burst, but incubation with these cytokines for 18 h primed neutrophils for a greater oxidative burst triggered by M. haemolytica and for enhanced uptake of S. aureus. The maximal response was observed when neutrophils were incubated with both cytokines together, at concentrations of 200 ng/ml for G-CSF and 400 ng/ml for IL-8. The IL-8-induced priming effect was reduced by treatment with a neutralizing antibody to IL-8, and was not attributed to endotoxin contamination. Instillation of IL-8 into the lung using a bronchoscope induced neutrophil recruitment within 18 h. Neutrophils from IL-8-treated lung showed dose-dependent enhancement of the oxidative burst triggered by M. haemolytica. Histologically, neutrophils filled alveoli and bronchioles, and scattered macrophages contained neutrophils with morphological features of apoptosis. Thus, prolonged in vitro or in vivo exposure to IL-8 and/or G-CSF enhances the subsequent oxidative burst and phagocytic responses of bovine neutrophils.