Eosinophil peroxidase stimulates the release of granulocyte-macrophage colony-stimulating factor from bronchial epithelial cells

Background: Asthma is characterized by an accumulation of activated eosinophils in the airway. Eosinophil viability-enhancing activity is present in the sputum of patients with asthma, largely because of granulocyte-macrophage colony-stimulating factor (GM-CSF). Bronchial epithelial cells have been shown to release cytokines including GM-CSF when stimulated with IL-1 beta or tumor necrosis factor-alpha. Objective: the study was designed to determine whether eosinophil peroxidase (EPO) stimulates the release of GM-CSF from bronchial epithelial cells. Methods: Epithelial cells (BEAS-2B) were cultured in serum free HD-F12 medium in a 24-well tissue culture plated until they became confluent. the cells were then exposed to EPO (5.9 x 10(-8) to 5.9 x 10(-7) mol/L) for 15 minutes, washed twice, and cultured in 1 ml of HD-F12. The supernatants were harvested at 3, 6 or 24 hours, and GM-CSF concentration was measured by ELISA. BEAS-2B cells were also treated with a system comprising EPO (1.9 x 10(-9) to 5.9 x 10(-8) mol/L) + 10(-5) mol/L H2O2 + 10(-4) mol/L Br for 24 hours. Results: The GM-CSF concentration in the supernatant pretreated with EPO increased in a time- and concentration-dependent manner compared with control The release of GM-CSF was not inhibited by catalase but was inhibited by cyclohexamide and by mixing of EPO with heparin, suggesting that the action is due to the cationic property of EPO. When EPO was combined with H2O2 and Br, 5.9 x 10(-9) mol/L EPO + 10(-5) mol/L H2O2 released two times more GM-CSF into the supernatants compared with control. Conclusion: These results suggest that EPO stimulates epithelial cells to release GM-CSF and forms a self-stimulatory cycle.