Mechanisms of mast cell signaling in anaphylaxis.

The recent development of a consensus definition and proposed diagnostic criteria for anaphylaxis offers promise for research efforts and a better understanding of the epidemiology and pathogenesis of this enigmatic and life-threatening disease. This review examines basic principles and recent research advances in the mechanisms of mast cell signaling believed to underlie anaphylaxis. The unfolding complexity of mast cell signaling suggests that the system is sensitive to regulation by any of several individual signaling pathways and intermediates and that complementary pathways regulate mast cell activation by amplified signals. The signaling events underlying anaphylactic reactions have largely been identified through experiments in genetically modified mice and supported by biochemical studies of mast cells derived from these mice. These studies have revealed that signaling pathways exist to both upregulate and downregulate mast cell responses. In this review we will thus describe the key molecular players in these pathways in the context of anaphylaxis.

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. Activating mutations in critical signaling molecules may also contribute to such responses. In this review, we describe our research exploring the mechanisms regulating these synergistic interactions and, furthermore, discuss the relevance of our observations in the context of clinical considerations.

Understanding the mechanisms of anaphylaxis.

PURPOSE OF REVIEW: The present review considers recent reports that identify the roles of key intermediate signaling components and mediators during and after mast cell activation and degranulation leading to anaphylaxis. RECENT FINDINGS: Mechanisms of anaphylaxis are becoming better understood as the interaction of several regulatory systems in the mast cell activation and degranulation signaling cascade. Multiple tyrosine kinases, activated after immunoglobulin E binding to the high-affinity receptors for immunoglobulin E (FcepsilonRI), exert both positive and negative regulation on the signaling cascade, which may vary with genetic background or mutations in signaling proteins. Calcium influx, the essential, proximal intracellular event leading to mast cell degranulation, is controlled also by both negative and positive regulation through calcium channels. Sphingosine-1-phosphate is emerging as a newly realized mediator of anaphylaxis, acting as a signaling component within the mast cell and as a circulating mediator. SUMMARY: Anaphylaxis is a systemic reaction involving multiple organ systems, but it is believed that it may be influenced by cellular events in mast cells and basophils resulting in the release of mediators. Therefore, understanding the mechanisms of mast cell activation and degranulation is critical to understanding the mechanisms of anaphylaxis. Recent reports have identified important regulatory components of the signaling cascade and, consequently, potential targets for therapeutic intervention.

Differential effects of Gq alpha, G14 alpha, and G15 alpha on vascular smooth muscle cell survival and gene expression profiles.

Gqalpha family members (Gqalpha, G11alpha, G14alpha, and G15/16alpha) stimulate phospholipase Cbeta (PLCbeta) and inositol lipid signaling but differ markedly in amino acid sequence and tissue distribution predicting unappreciated functional diversity. To examine functional differences, we compared the signaling properties of Gqalpha, G14alpha, and G15alpha and their cellular responses in vascular smooth muscle cells (VSMC). Constitutively active forms of Gqalpha, G14alpha, or G15alpha elicit markedly different responses when introduced to VSMC. Whereas each Galpha stimulated PLCbeta to similar extents when expressed at equal protein levels, Gqalpha and G14alpha but not G15alpha initiated profound cell death within 48 h. This response was the result of activation of apoptotic pathways, because Gqalpha and G14alpha, but not G15alpha, stimulated caspase-3 activation and did not alter phospho-Akt, a regulator of cell survival pathways. Gqalpha and G14alpha stimulate nuclear factor of activated T cell (NFAT) activation in VSMC, but Galpha-induced cell death seems independent of PKC, InsP(3)/Ca(2+), and NFAT, in that pharmacological inhibitors of these pathways did not block cell death. Gene expression analysis indicates that Gqalpha, G14alpha, and G15alpha each elicit markedly different profiles of altered gene sets in VSMC after 24 h. Whereas all three Galpha stimulated changes (> or =2-fold) in 50 shared mRNA, Gqalpha and G14alpha (but not G15alpha) stimulated changes in 221 shared mRNA, many of which are reported to be pro-apoptotic and/or involved with TNF-alpha signaling. We were surprised to find that each Galpha also stimulated changes in nonoverlapping Galpha-specific gene sets. These findings demonstrate that Gqalpha family members activate both overlapping and distinct signaling pathways and are more functionally diverse than previously thought.

Common signaling pathways link activation of murine PAR-1, LPA, and S1P receptors to proliferation of astrocytes.

Receptors for the serine protease thrombin and for lysophospholipids are coupled to G proteins and control a wide range of cellular functions, including mitogenesis. Activators of these receptors are present in blood, and can enter the brain during central nervous system (CNS) injury. Reactive astrogliosis, a prominent component of CNS injury with potentially harmful consequences, may involve proliferation of astrocytes. In this study, we have examined the expression and activation of protease activated receptors (PARs), lysophosphatidic acid (LPA) receptors, and sphingosine-1-phosphate (S1P) receptors on murine astrocytes. We show that activation of these three receptor classes can lead to astrogliosis in vivo and proliferation of astrocytes in vitro. Cultured murine cortical astrocytes express mRNA for multiple receptor subtypes of PAR (PAR-1-4), LPA (LPA-1-3) and S1P (S1P-1, -3, -4, and -5) receptors. Comparison of the intracellular signaling pathways of glial PAR-1, LPA, and S1P receptors indicates that each receptor class activates multiple downstream signaling pathways, including Gq/11-directed inositol lipid/Ca2+ signaling, Gi/o activation of mitogen-activated protein kinases (MAPK) (extracellular signal-regulated kinase 1/2 and stress activated protein kinase/c-jun N-terminal kinase, but not p38), and activation of Rho pathways. Furthermore, activation of these different receptor classes can differentially regulate two transcription factor pathways, serum response element and nuclear factor of activated T cells. Blockade of Gi/o signaling with pertussis toxin, MAPK activation with 1,4-diamino-2,3-dicyano-1,4-bis(2-aminophynyltio)butadiene (U0126), or Rho kinase signaling with R-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexane carboxamide (Y27632) can markedly reduce the proliferative response of glial cells to PAR-1, LPA, or S1P receptor activation, suggesting that each of these pathways is important in coupling of receptor activation to glial proliferation.

Differential regulation of metabotropic glutamate receptor 5-mediated phosphoinositide hydrolysis and extracellular signal-regulated kinase responses by protein kinase C in cultured astrocytes.

The metabotropic glutamate receptor 5 (mGluR5) exhibits a rapid loss of receptor responsiveness to prolonged or repeated agonist exposure. This receptor desensitization has been seen in a variety of native and recombinant systems, and is thought to result from receptor-mediated, protein kinase C (PKC)-dependent phosphorylation of the receptor, uncoupling it from the G protein in a negative feedback regulation. We have investigated the rapid PKC-mediated desensitization of mGluR5 in cortical cultured astrocytes by measuring downstream signals from activation of mGluR5. These include activation of phosphoinositide (PI) hydrolysis, intracellular calcium transients, and extracellular signal-regulated kinase 2 (ERK2) phosphorylation. We present evidence that PKC plays an important role in rapid desensitization of PI hydrolysis and calcium signaling, but not in ERK2 phosphorylation. This differential regulation of mGluR5-mediated responses suggests divergent signaling and regulatory pathways which may be important mechanisms for dynamic integration of signal cascades.