Flow cytometry-based monitoring of mast cell activation.

Mast cell activation is a central process in the initiation of allergic disorders. As described elsewhere in this volume, this process can be readily monitored by biochemical, antibody-based, and enzyme-based formats when the cell population examined is homogenous. When dealing with mixed and transfected cell populations however, such approaches may not be appropriate.

Knockout of the Trpc1 gene reveals that TRPC1 can promote recovery from anaphylaxis by negatively regulating mast cell TNF-α production.

Antigen-mediated mast cell (MC) degranulation is the critical early event in the induction of allergic reactions. Transient receptor potential channels (TRPC), particularly TRPC1, are thought to contribute to such MC activation. To explore the contribution of TRPC1 in MC-driven allergic reactions, we examined antigen-mediated anaphylaxis in Trpc1⁻/⁻ and WT mice, and TRPC1 involvement in the activation of MCs derived from the bone marrow (BMMCs) of these mice.

Btk-dependent Rac activation and actin rearrangement following FcepsilonRI aggregation promotes enhanced chemotactic responses of mast cells.

Mast cells infiltrate the sites of inflammation associated with chronic atopic disease and during helminth and bacterial infection. This process requires receptor-mediated cell chemotaxis across a concentration gradient of their chemotactic ligands. In vivo, mast cells are likely to be exposed to several such agents, which can cooperate in a synergistic manner to regulate mast cell homing.

Amplification mechanisms for the enhancement of antigen-mediated mast cell activation.

Activation of mast cells in the allergic inflammatory response occurs via the high affinity receptor for IgE (FcepsilonRI) following receptor aggregation induced by antigen-mediated cross-linking of IgE-occupied FcepsilonRI. Recent observations suggest this response is profoundly influenced by other factors that reduce the threshold for, and increase the extent of, mast cell activation. For example, under experimental conditions, cell surface receptors such as KIT and specific G protein-coupled receptors synergistically enhance FcepsilonRI-mediated mast cell degranulation and cytokine production.

Synergistic activation of phospholipases Cgamma and Cbeta: a novel mechanism for PI3K-independent enhancement of FcepsilonRI-induced mast cell mediator release.

Antigen/IgE-mediated mast cell activation via FcvarepsilonRI can be markedly enhanced by the activation of other receptors expressed on mast cells and these receptors may thus contribute to the allergic response in vivo. One such receptor family is the G protein-coupled receptors (GPCRs). Although the signaling cascade linking FcvarepsilonRI aggregation to mast cell activation has been extensively investigated, the mechanisms by which GPCRs amplify this response are relatively unknown.

G protein-coupled receptors and the modification of FcepsilonRI-mediated mast cell activation.

By releasing multiple pro-inflammatory mediators upon activation, mast cells are critical effector cells in the pathogenesis of allergic inflammation. The traditional viewpoint of antigen-dependent mast cell activation is that of a Th(2)-driven process whereby antigen-specific IgE molecules are produced by B cells followed by binding of the IgE to high affinity IgE receptors (FcepsilonRI) expressed on mast cells. Subsequent antigen-dependent aggregation of the FcepsilonRI initiates an intracellular signalling cascade that culminates in mediator release.

Targeting kit activation: a potential therapeutic approach in the treatment of allergic inflammation.

The prevalence of allergic diseases is increasing worldwide. Hence, there is continued need for novel pharmacological therapies for the treatment of these disorders. As the mast cell is one of the essential cells that contributes to the inflammation associated with allergic diseases, this cell type remains an attractive target for such pharmacological intervention.

Comparison of Fc epsilon RI- and Fc gamma RI-mediated degranulation and TNF-alpha synthesis in human mast cells: selective utilization of phosphatidylinositol-3-kinase for Fc gamma RI-induced degranulation.

We have demonstrated that CD34(+) IFN-gamma-treated human mast cells (HuMC) express functional Fc gamma RI and that aggregation of these receptors leads to mediator release. As the signaling pathways linking Fc gamma RI aggregation to mediator release are unknown, we examined Fc gamma RI-dependent activation of specific signal transduction molecules and determined the relative involvement of these events in HuMC degranulation and TNF-alpha production following both Fc gamma RI and Fc epsilon RI aggregation. Fc gamma RI aggregation resulted in the phosphorylation/activation of src kinases and p72(syk) and subsequent tyrosine phosphorylation of multiple substrates.

Activation of human mast cells through the high affinity IgG receptor.

Mast cells are known to participate in the induction of inflammation through interaction of antigen with specific IgE bound to the high affinity receptor for IgE (FcepsilonRI). Human mast cells, derived from CD34(+) hematopoietic precursors, not only express FcepsilonRI but also express high affinity receptors for IgG (FcgammaRI), the latter only after IFN-gamma exposure. Human mast cells that express FcgammaRI are activated following FcgammaRI aggregation, either using antibodies directed to the receptor or through IgG bound to the receptor.

Determination of protein phosphorylation in Fc epsilon RI-activated human mast cells by immunoblot analysis requires protein extraction under denaturing conditions.

The advent of activation state antibodies has greatly facilitated studies aimed at understanding the intracellular signaling cascade following occupancy and/or aggregation of surface receptors. As part of an ongoing study investigating the signal transduction cascade initiated following aggregation of the high affinity receptor for IgE (Fc epsilon RI) in human mast cells, we observed substantial differences in responses monitored by these antibodies when cells were extracted either under nonreducing or reducing conditions. This was true even in the presence of high concentrations of protease inhibitors.

Mastocytosis: molecular mechanisms and clinical disease heterogeneity.

Systemic mastocytosis has one unifying feature: an unexplained and pathologic increase in mast cells in specific tissues. This observation, along with clinical disease heterogeneity has long suggested that mastocytosis is a disease of complex etiology. At the same time, the last decade has witnessed significant progress in identifying the critical elements that regulate mast cell growth and development.