Cieslewicz, Grzegorz

Abstract Allergen provocation of allergic asthma patients is often characterized by an initial period of bronchoconstriction, or early phase reaction (EPR), that leads to maximal airway narrowing within 15–30 min, followed by a recovery period returning airway function to baseline within 1–2 h. In this study, we used a defined OVA provocation model and mice deficient for specific leukocyte populations to investigate the cellular/molecular origins of the EPR. OVA-sensitized/challenged wild-type (C57BL/6J) mice displayed an EPR following OVA provocation. However, this response was absent in gene knockout animals deficient of either B or T cells. Moreover, transfer of OVA-specific IgG, but not IgE, before the OVA provocation, was capable of inducing the EPR in both strains of lymphocyte-deficient mice. Interestingly, an EPR was also observed in sensitized/challenged mast cell-deficient mice following an OVA provocation. These data show that the EPR in the mouse is an immunologically based pathophysiological response that requires allergen-specific IgG but occurs independent of mast cell activities. Thus, in the mouse the initial period of bronchoconstriction following allergen exposure may involve neither mast cells nor IgE-mediated events.

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.

Abstract The relationship between eosinophils and the development of Ag-induced pulmonary pathologies, including airway hyper-responsiveness, was investigated using mice deficient for the secondary granule component, major basic protein-1 (mMBP-1). The loss of mMBP-1 had no effect on OVA-induced airway histopathologies or inflammatory cell recruitment. Lung function measurements of knockout mice demonstrated a generalized hyporeactivity to methacholine-induced airflow changes (relative to wild type); however, this baseline phenotype was observable only with methacholine; no relative airflow changes were observed in response to another nonspecific stimulus (serotonin). Moreover, OVA sensitization/aerosol challenge of wild-type and mMBP-1−/− mice resulted in identical dose-response changes to either methacholine or serotonin. Thus, the airway hyper-responsiveness in murine models of asthma occurs in the absence of mMBP-1.

Abstract The temporal association between airway inflammation and airway hyperresponsiveness (AHR) has been analyzed in BALB/c mice sensitized to, and subsequently exposed to, a single intranasal challenge of ovalbumin (OVA). In OVA-sensitized/challenged animals only a small increase in responsiveness to methacholine (MCh) was seen at 8 h, peaked at 24 to 48 h, and resolved by 96 h. An early bronchoalveolar lavage fluid (BALF) neutrophil infiltrate (peaking at 8 h postchallenge; ∼ 72% total cells was observed) that returned to baseline by 48 h. BALF eosinophil numbers did not increase until 48 h (∼ 32% of total cells), peaked at 96 h (∼ 38% total cells), and remained elevated at 8 d (∼ 27% total cells). Airway tissue eosinophilia preceded changes in BALF. Eosinophil peroxidase (EPO) levels in BALF were elevated in OVA-sensitized/challenged mice at 48 h only. BALF TNF- α levels peaked at 8 h, whereas IL-5 and IL-4 levels peaked at 24 h. IL-13 levels were increased at both 24 and 48 h. Mucus-positive cells were not observed in the airway epithelium until 48 h. Administration of IL-5 or VLA-4 antibody prior to OVA challenge prevented the development of AHR in sensitized mice as well as BALF and tissue eosinophilia. These data identify a temporal association between Th2 cytokine production, tissue eosinophil infiltration and activation, and, importantly, both the development and resolution kinetics of AHR. Moreover, the antibody studies further support the association of eosinophilia with the pathogenesis of AHR.