Interleukin-5 (IL-5) is important in the control of differentiation, migration, and activation of eosinophils. In order to study the role of IL-5 in the development of eosinophilic inflammation of the airways, we have used a monoclonal antibody to murine IL-5 (TRFK-5) in a murine model of allergic pulmonary inflammation. B6D2F1 mice were sensitized with alum-precipitated ovalbumin and were challenged with aerosolized ovalbumin on day 12 after sensitization. Samples of bronchoalveolar lavage (BAL) fluid, lung tissue, blood, and bone marrow aspirate were collected at different times after ovalbumin challenge. Twenty-four hours after challenge there were significant increases in the number of eosinophils in the BAL fluid, lung tissue, and blood while bone marrow eosinophils were decreased. Treatment of sensitized mice with TRFK-5 (0.01-1 mg/kg, i.p.) 2 h before ovalbumin challenge reduced the numbers of eosinophils in the BAL fluid and lung tissue and prevented the decrease in bone marrow eosinophils in a dose-dependent fashion. The number of eosinophils in the BAL fluid, peribronchial and alveolar regions of the lung was also reduced when TRFK-5 (2 mg/kg, i.p.) was given up to 5 d after ovalbumin challenge. Furthermore, there was no evidence of increased epithelial damage, edema, or the presence of mucus that could have resulted from eosinophil apoptosis and release of toxic proteins after neutralization of IL-5. These results demonstrate an important role for IL-5 in the development of eosinophilic inflammation of the airways and for the migration of eosinophils from the bone marrow into blood in response to antigen challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Mast cells are important effector cells in IgE-mediated acute allergic reactions. Mast cells also produce cytokines such as interleukin (IL)-3, IL-4, IL-5, tumor necrosis factor (TNF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) that regulate the function of eosinophils and the development of a late-phase inflammatory response to antigen challenge. To evaluate the role of mast cells on the development of IgE-mediated allergic pulmonary eosinophilia in vivo, we compared the eosinophil infiltration into lungs of mast cell deficient mice (WBB6F1/J-W/Wv) with their congenic normal littermates (W/W+). Mice were sensitized with alum-precipitated ovalbumin and challenged with aerosolized ovalbumin on day 12 after sensitization. Bronchoalveolar lavage (BAL) fluid, lung tissue biopsies, and blood samples were collected after ovalbumin challenge. Eosinophil numbers in the BAL and lung tissue, lung eosinophil peroxidase (EPO) activity and serum levels of IgE and IgG1 were measured. In sensitized W/W+ mice, there were increased numbers of eosinophils in the BAL fluid and lung tissue, and EPO levels were increased after ovalbumin challenge. Ovalbumin challenge of sensitized mast-cell-deficient mice produced fewer numbers of eosinophils in the BAL fluid and lungs, and EPO levels were also reduced compared with their challenged congenic littermates. On the other hand, levels of serum IgE and IgG1 were not different between W/Wv mice and their congenic littermates.(ABSTRACT TRUNCATED AT 250 WORDS)

Nitric oxide (NO) is an important mediator of inflammatory reactions and may contribute to the lung inflammation in allergic pulmonary diseases. To assess the role of NO in pulmonary inflammation, we studied the effect of four nitric oxide synthase (NOS) inhibitors, N-nitro-L-arginine methyl ester (L-NAME), aminoguanidine, N(G)-monomethyl-L-arginine (NMMA) and L-N6-(1-Iminoethyl) lysine (L-NIL), on the influx of eosinophils into the bronchoalveolar lavage (BAL) fluid and lung tissue of antigen-challenged allergic mice. We also analyzed lung tissues for the presence of steady state mRNA for inducible nitric oxide synthase (iNOS) and iNOS protein. Furthermore, BAL fluid and serum were analyzed for their nitrite content. B6D2F1/J mice were sensitized to ovalbumin (OVA) and challenged with aerosolized OVA. The NOS inhibitors were given 0.5 h before and 4 h after the antigen challenge. OVA challenge induced a marked eosinophilia in the BAL fluid and lung tissue 24 h after challenge. The OVA-induced pulmonary eosinophilia was significantly reduced by L-NAME (10 and 50 mg/kg, intraperitoneally [i.p.]). The inactive isomer, D-NAME (50 mg/kg, i.p.) had no effect. When mice were treated with L-NAME (20 mg/kg, i.p.) and an excess of NOS substrate, L-arginine (200 mg/kg, i.p.), the OVA-induced pulmonary eosinophilia was restored. Treatment with aminoguanidine (0.4-50 mg/kg, i.p.) also reduced the pulmonary eosinophilia. Treatment with NMMA (2-50 mg/kg, i.p.) partially reduced the eosinophilia, but L-NIL (10-50 mg/kg, i.p.), a selective iNOS inhibitor, had no effect. L-NAME had no effect on the reduction of eosinophils in the bone marrow following OVA challenge to sensitized mice. OVA challenge to sensitized mice had no effect on iNOS protein expression or iNOS mRNA in the lungs or on the levels of nitrite in the BAL fluid. These results suggest that NO is involved in the development of pulmonary eosinophilia in allergic mice. The NO contributing to the eosinophilia is not generated through the activity of iNOS nor does NO contribute to the efflux of eosinophils from the bone marrow in response to antigen challenge. It is speculated that after antigen challenge, the localized production of NO, possibly from pulmonary vascular endothelial cells, is involved in the extravasation of eosinophils from the circulation into the lung tissue.

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This report describes the development and the biology of Sch 55700, a humanized monoclonal antibody to human IL-5 (hIL-5). Sch 55700 was synthesized using CDR (complementarity determining regions) grafting technology by incorporating the antigen recognition sites for hIL-5 onto consensus regions of a human IgG4 framework. In vitro, Sch 55700 displays high affinity (Kd = 20 pmol/l) binding to hIL-5, inhibits the binding of hIL-5 to Ba/F3 cells (IC50 = 0.5 nmol/l) and blocks IL-5 mediated proliferation of human erythroleukemic TF-1 cells. In allergic mice, Sch 55700 (0.1-10 mg/kg, i.p. or i.m.) inhibits the influx of eosinophils in the lungs, demonstrates long duration of activity and the anti-inflammatory activity of this compound is additive with oral prednisolone. In allergic guinea pigs, Sch 55700 (0.03-30 mg/kg i.p.) inhibits both the pulmonary eosinophilia and airway hyperresponsiveness and at 30 mg/kg, i.p. inhibited allergic, but not histamine-induced bronchoconstriction. In allergic rabbits, Sch 55700 blocks cutaneous eosinophilia. Sch 55700 (0.1-1 mg/kg i.p.) also blocks the pulmonary eosinophilia and neutrophilia caused by tracheal injection of hIL-5 in guinea pigs. In allergic cynomolgus monkeys, a single dose of Sch 55700 (0.3 mg/kg i.v.) blocks the pulmonary eosinophilia caused by antigen challenge for up to six months. Sch 55700 is, therefore, a potent antibody against IL-5 in vitro and in a variety of species in vivo that could be used to establish the role of IL-5 in human eosinophilic diseases such as asthma.