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Keywords:

  • eosinophil cationic protein;
  • human eosinophils;
  • N-acetylcysteine;
  • p47phox–p67phox;
  • reactive oxygen species

Summary

Background Oxidative stress appears to be relevant in the pathogenesis of inflammation in allergic diseases like bronchial asthma. Eosinophils are oxidant-sensitive cells considered as key effectors in allergic inflammation.

Objective The aim of this work was to study the effects of the clinically used antioxidant N-acetyl-l-cysteine (NAC) on the functional responses of human-isolated eosinophils.

Methods Human eosinophils were purified from the blood of healthy donors by a magnetic bead separation system. The effects of NAC were investigated on the generation of reactive oxygen species (chemiluminescence and flow cytometry), Ca2+ signal (fluorimetry), intracellular glutathione (GSH; flow cytometry), p47phox–p67phox translocation (Western blot) and eosinophil cationic protein (ECP) release (radioimmunoassay).

Results NAC (0.1–1 mm) inhibited the extracellular generation of oxygen species induced by N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP) and eotaxin (in the presence of IL-5) with −logIC50 values of 3.61±0.03 and 3.36±0.09, respectively. Also, the intracellular generation of hydrogen peroxide was virtually abolished by NAC (0.5–1 mm). NAC (1 mm) did not alter the fMLP-induced Ca2+ signal but augmented the eosinophil content of reduced GSH and inhibited p47phox–p67phox translocation. NAC inhibited the release of ECP (∼90% inhibition at 1 mm) from fMLP-activated eosinophils.

Conclusion Inhibition by NAC of human eosinophil functions in vitro is potentially useful in the treatment of allergic inflammation.