Kirsten A. Larson

The late, but not early, asthmatic response is dependent on IL-5 and correlates with eosinophil infiltration

Early-phase reactions (EPRs) and late-phase reactions (LPRs) are characteristic features of bronchial asthma, although the pathogenetic mechanisms responsible for each of the responses are not fully defined. A murine model of EPRs and LPRs was developed to investigate the role of IL-5 and eosinophils in development of both responses. After initial intraperitoneal sensitization and airway challenge to ovalbumin (OVA), mice were provoked by additional exposure to OVA. An EPR, characterized by a transient increase in airway responsiveness, was observed 5-30 minutes after antigen provocation. This response was followed by an LPR that reached its maximum at 6 hours after challenge and was characterized by increased airway responsiveness and significant lung eosinophilia. The EPR was blocked by cromoglycate and albuterol, whereas the LPR was abolished by cromoglycate and hydrocortisone. Before provocation with allergen, administration of anti-IL-5 antibody prevented the influx of eosinophils into the lung tissue and abolished the LPR but not EPR. These results suggest that IL-5 and eosinophils are essential for development of the LPR, but not EPR, in this model.

Eosinophil Major Basic Protein-1 Does Not Contribute to Allergen-Induced Airway Pathologies in Mouse Models of Asthma

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.

Mouse eosinophil-associated ribonucleases: a unique subfamily expressed during hematopoiesis

Abstract. A unique family of ribonucleases was identified by exhaustive screening of genomic and cDNA libraries using a probe derived from a gene encoding a ribonuclease stored in the mouse eosinophil secondary granule. This family contains at least 13 genes, which encode ribonucleases, and two potential pseudogenes. The conserved sequence identity among these genes (∼70%), as well as the isolation/purification of these ribonucleases from eosinophil secondary granules, has led us to conclude that these genes form a unique clade in the mouse that we have identified as the Ear (Eosinophil-associated ribonuclease) gene family. Analyses of the nucleotide substitutions that have occurred among these ribonuclease genes reveal that duplication events within this family have been episodic, occurring within three unique periods during the past 18 × 106 years. Moreover, comparisons of non-synonymous (Ka) vs. synonymous (Ks) rates of nucleotide substitution show that although these genes conserve residues necessary for RNase activity, selective evolutionary pressure(s) exist such that acquired amino acid changes appear to be advantageous. The selective advantage of these amino acid changes is currently unclear, but the occurrence of this phenomenon in both the mouse and the human highlights the importance of these changes for Ear and, therefore, eosinophil effector function(s).

Identification of a new murine eosinophil major basic protein (mMBP) gene: Cloning and characterization of mMBP-2

We have identified a new eosinophil major basic protein gene family member in the mouse and have given it the designation murine major basic protein‐2 (mMBP‐2). The gene was initially characterized as a unique expressed sequence tag (EST) clone having significant identity to the previously recognized member of this gene family, mMBP‐1. The EST was used to screen and isolate mMBP‐2 from a bone marrow cDNA library. In addition, a genomic clone of mMBP‐2 was isolated and this gene was shown to be physically linked to within 100 kb of mMBP‐1 on the central region of mouse chromosome 2. Progressive similarity alignment of the deduced mMBP‐2 open reading frame demonstrates the apparent conservation of the “pre‐pro‐mature” protein structure found in the other known mammalian MBPs. Mature mMBP‐2 maintains the cationic nature associated with these proteins with a predicted pI of 9.95. However, unlike the human MBPs, which display a three orders of magnitude charge difference [hMBP‐1 (pI11.4) vs. hMBP‐2 (pI8.7)], mMBP‐2 is only slightly less cationic than mMBP‐1 (pI 10.5). Expression studies demonstrate that transcription of the mMBP‐2 gene parallels mMBP‐1 and is confined to hematopoietic compartments engaged in eosinophilopoiesis. Moreover, using mMBP‐1 knockout mice and immunohistochemistry with an antisera that recognizes both mMBP‐1 and ‐2, we demonstrate that mMBP‐2 protein expression is restricted to eosinophil lineage‐committed cells. J. Leukoc. Biol. 67: 567–576; 2000.