This application focuses on the use of human umbilical cord blood-derived mast cells and a mast cell line, HMC-1 (serving as a control), to understand the role of these cells in the cellular networks regulating inflammation. Mast cells and helper T-cells often co-localize to areas of inflammation, as in the airways of patients with bronchial asthma. The investigator's primary hypothesis is that activated human mast cells are capable of expressing a repertoire of inflammatory cytokines and/or chemokines and that this process could be regulated by interaction with co-stimulatory molecules expressed, or cytokines secreted, by T-cells.
The aims of the project are to: 1. Generate human mast cells from cord blood stem cells and to characterize these human cord blood-derived mast cells (CBDMC) in terms of morphology, protease expression, co-stimulatory molecule expression, and cytokine/chemokine repertoire in response to IgE-dependent and IgE-independent activation. 2. To assess the regulation of nuclear factor kappa-B (NF-kB) and IkBa in response to IgE receptor cross-linking in CBDMC, transcription factors critical to the induction of several inflammatory cytokines. 3. To determine whether supernatants from Th1/Th2 cells and select Th1 (IL-2, g-IFN) and Th2 (IL-4/IL-13) cytokines modulate expression of mediators (histamine, tryptase, PGD2) and key cytokines, IL-4, IL-8, IL-13, MCP-1, MIP-1a from HMC-1 and CBDMC either independently or in response to immunological activation. 4. To determine if cognate signals from activated T-lymphocytes (by addition of membranes from resting or activated T-cells) can trigger CBDMC activation independently or in synergy with IgE-mediated signaling. The investigator proposes to determine whether this results in the nuclear translocation of NF-kB. He will also study expression of mediators (histamine, tryptase, PGD2) and of genes encoding key cytokines, IL-4, IL-8, IL-13, MCP-1, MIP-1a. If cytokine signaling occurs, the investigator will attempt to determine whether this occurs via a CD40-CD40L, B7-CD28 or LFA-1/ICAM-1-dependent interaction by the use of appropriate monoclonal antibodies and blocking studies. These studies may shed light on the molecular pathogenesis of asthma, an inflammatory disease of human airways.
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| Fitzgerald, S Matthew; Chi, David S; Lee, Steven A et al. (2004) Inhibition of GM-CSF production in fibroblast-monocyte coculture by prednisone and effects of rhGM-CSF on human lung fibroblasts. Front Biosci 9:342-8 |
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