Eosinophil peroxidase mediates protein nitration in allergic airway inflammation in mice

The eosinophilic inflammatory response in asthma is associated with protein nitration, detected as immunostaining for 3-nitrotyrosine (3NT). As the pr esence of 3NT is strongly correlated with upregulation of the inducible for m of nitric oxide synthase (NOS II), it has been hypothesized that 3NT form ation results from the action of peroxynitrite (ONOO-), a highly reactive N O derivative produced from the reaction of molecular NO and O-2(-). However , recent observations have suggested that the action of peroxidases, includ ing eosinophil peroxidase (EPO), may be responsible for protein nitration. In this study, we used murine models of allergic asthma to address the rela tive contribution of EPO and NOS II to protein nitration. We studied EPO-de ficient New Zealand White (NZW) mice, which were sensitized and challenged intranasally with oval-bumin (OVA). Despite comparable levels of eosinophil ia, NO, and superoxide production, NZW mice exhibited markedly decreased 3N T staining around the airways after OVA challenge when compared with two ot her strains (A/J and C57BL/6J). Immunocytochemical analysis of bronchoalveo lar lavage (BAL) cells and lung sections suggested that 3NT staining was la rgely confined to eosinophils. This was confirmed by Western Blot analysis of proteins from different subsets of BAL cells that demonstrated a marked decrease in 3NT formation in eosinophils from NZW mice. These results contr ast with those obtained in OVA-sensitized and -challenged NOS It deficient mice, which despite decreased NO production, exhibited similar 3NT staining in the airways after OVA challenge as in wild-type control mice. In this m odel, protein nitration was thus not a function of NO production by NOS II. We conclude that in the mouse, 3NT formation after specific allergen chall enge is dependent on EPO activity, particularly in eosinophils themselves. In contrast, 3NT formation is not driven by upregulation of NOS II expressi on in this model and does not appear to depend on increases in the level of NO production.