Extensive Eosinophil Degranulation and Peroxidase-Mediated Oxidation of Airway Proteins Do Not Occur in a Mouse Ovalbumin-Challenge Model of Pulmonary Inflammation

Abstract

Paradigms of eosinophil effector function in the lungs of asthma patients invariably depend on activities mediated by cationic proteins released from secondary granules during a process collectively referred to as degranulation. In this study, we generated knockout mice deficient for eosinophil peroxidase (EPO) to assess the role(s) of this abundant secondary granule protein in an OVA-challenge model. The loss of EPO had no effect on the development of OVA-induced pathologies in the mouse. The absence of phenotypic consequences in these knockout animals extended beyond pulmonary histopathologies and airway changes, as EPO-deficient animals also displayed OVA-induced airway hyperresponsiveness after provocation with methacholine. In addition, EPO-mediated oxidative damage of proteins (e.g., bromination of tyrosine residues) recovered in bronchoalveolar lavage from OVA-treated wild-type mice was <10% of the levels observed in bronchoalveolar lavage recovered from asthma patients. These data demonstrate that EPO activities are inconsequential to the development of allergic pulmonary pathologies in the mouse and suggest that degranulation of eosinophils recruited to the lung in this model does not occur at levels comparable to those observed in humans with asthma.

Coexpression of IL-5 and Eotaxin-2 in Mice Creates an Eosinophil-Dependent Model of Respiratory Inflammation with Characteristics of Severe Asthma

“…9 -11) linked with marked methacholine sensitivity and significant airway epithelial/smooth muscle remodeling. In contrast, similar observations are lacking in the mouse as exemplified in studies of knockout animals deficient of the abundant eosinophil secondary granule proteins, major basic protein-1 (MBP-1) 3 (12) or eosinophil peroxidase (EPO) (13). Thus, the lack of extensive degranulation in the mouse has limited insights regarding the extent by which eosinophils contribute to allergic pulmonary disease.…”

“…C57BL/6J mice (8 -16 wk of age) were sensitized and challenged with chicken (OVA) as described earlier (13). Mice were assessed for pulmonary cellular infiltrates, eosinophil degranulation, and histopathologies on day 28 (i.e., 2 days following the last OVA challenge) of this protocol.…”

Eosinophil Peroxidase Catalyzed Protein Carbamylation Participates in Asthma

“…Mice were challenged on days 24, 25, and 26 by 20-min inhalations of an aerosol (nebulized 1% OVA in normal saline; control mice received a 20-min aerosol of nebulized normal saline). Mice on day 28 were assessed for pulmonary histopathologies, tissue harvest, cellular infiltrates, or lung function as described (49). Lungs from EPO ϩ/ϩ and EPO Ϫ/Ϫ mice (n ϭ 6 per group), challenged with OVA and normal saline (NS) as a control, were also provided by the Mayo Clinic.…”

“…Initially we used a well characterized model where large numbers of eosinophils (Ͼ10 6 , 20 -30%) are induced into the peritoneal cavity of mice via a sensitization/challenge protocol using a whole-protein extract of the helminth Mesocestoides corti. Subsequent intraperitoneal injection of zymosan, a yeast cell wall glucan, triggers activation of the recruited peritoneal eosinophils, resulting in eosinophil degranulation and EPO release, coupled with H 2 O 2 generation from the oxidant burst (20,49). As illustrated in Fig.…”

Evidence for eosinophil recruitment, leukotriene B4 production and mast cell hyperplasia following Toxocara canis infection in rats

“…The supernatants were carefully removed, 100 µL substrate solution (2.4 mM o-phenylenediamine dihydrochloride in 50 mM Tris-HCl, pH 8.0, with 6.6 mM H 2 O 2 ) was added to each well, and the plates were incubated at room temperature for 15 min. This substrate is specific for EPO and does not recognize myeloperoxidase (23). The reaction was stopped by addition of 50 µL 4 M H 2 SO 4 , and the absorbance of the samples was determined at 490 nm.…”