The goal of this proposal is to better understand the mechanisms and control of immediate hypersensitivity disorders and asthma. Studies will focus on the critical interaction between human lung mast cells and eosinophils. Considerable evidence points to both mast cells and eosinophils playing a central role in asthma, though details of their interaction have never been completely studied in humans. Using the information gained from these studies, our long term goal is to apply this knowledge to the development of new interventions for the treatment of asthma. The studies involve isolating eosinophils from the blood of allergic and non-allergic donors by standard gradient techniques or by a negative- selection immunomagnetic bead technique. Purified human lung mast cells (HLMC) are isolated from the normal tissue in lungs resected for carcinoma, using techniques developed in our laboratory. Using these purified cell preparations, we will examine several measures of eosinophil activation in response to HLMC or HLMC mediators, focusing on the critical role of the extracellular matrix (ECM) in this interaction. The hypothesis to be tested is that the ECM provides critical signals (co-stimulatory or inhibitory) which modify the responsiveness of eosinophils to mast cell mediators. Preliminary data is presented showing that anti-IgE activated HLMC induce the formation of """"""""hypodense"""""""" eosinophils, eosinophils which are activated in comparison with normal eosinophils. Data is also presented demonstrating the potent eosinophil-stimulatory effects of crude supernatants from immunologically activated HLMC, which has lead us to further investigate the ability of mast cells to directly stimulate eosinophils when mast cells and eosinophils are cocultured. Finally, data is presented related to studies examining the important role of extracellular matrix (ECM) proteins in HLMC mediator-induced eosinophil activation. We propose to: 1) Determine the adhesion receptors responsible for eosinophil adherence to ECM, and the functional consequences of adherence. 2) Characterize the combined effects of adherence to ECM with individual HLMC mediator stimulation on the in vitro responsiveness of eosinophils obtained from both normal and atopic donors. 3) Using co-culture techniques, examine the direct interaction of anti-IgE activated HLMC with ECM-adherent eosinophils, and determine the functional consequences of this interaction.
| Mohanty, J G; Raible, D G; McDermott, L J et al. (2001) Effects of purine and pyrimidine nucleotides on intracellular Ca2+ in human eosinophils: activation of purinergic P2Y receptors. J Allergy Clin Immunol 107:849-55 |
| Raible, D G; Mohanty, J G; Jaffe, J S et al. (2000) Hydrogen peroxide release from human eosinophils on fibronectin: scopoletin enhances eosinophil activation. Free Radic Biol Med 28:1652-60 |
| Mohanty, J G; Jaffe, J S; Schulman, E S et al. (1997) A highly sensitive fluorescent micro-assay of H2O2 release from activated human leukocytes using a dihydroxyphenoxazine derivative. J Immunol Methods 202:133-41 |
