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The leaves of the Piper betle Linn. (Piperaceae) are used in traditional medicine and possess anti-oxidant, anti-bacterial, anti-fungal, anti-diabetic and radioprotective activities. However, little is known about their anti-allergic activity. Therefore, the effects of P. betle ethanolic extract (PE) on the production of histamine and granulocyte macrophage-colony-stimulating factor (GM-CSF) by murine bone marrow mast cells (BMMCs) and on the secretion of eotaxin and IL-8 by the human lung epithelial cell line, BEAS-2B, were investigated in vitro. PE significantly decreased histamine and GM-CSF produced by an IgE-mediated hypersensitive reaction, and inhibited eotaxin and IL-8 secretion in a TNF-alpha and IL-4-induced allergic reaction. The results suggest that P. betle may offer a new therapeutic approach for the control of allergic diseases through inhibition of production of allergic mediators.

We investigated the effect of several chlorinated organic solvents on antigen-induced histamine release and inflammatory mediator production. Non-purified rat peritoneal mast cells (NPMC) and rat basophilic leukemia (RBL-2H3) cells were sensitized with anti-dinitrophenol (DNP) monoclonal IgE antibody, and then stimulated with DNP-conjugated bovine serum albumin (DNP-BSA) and several chlorinated organic solvents. Trichloroethylene (TCE) and tetrachloroethylene (PCE) enhanced histamine release from antigen-stimulated NPMC and RBL-2H3 in a dose-dependent manner. In addition, TCE and PCE increased IL-4 and TNF-alpha production from antigen-stimulated RBL-2H3. In an in vivo study, we investigated the effect of TCE and PCE on passive cutaneous anaphylaxis (PCA) reaction. TCE and PCE enhanced PCA reaction markedly. These results suggest that TCE and PCE increase histamine release and inflammatory mediator production from antigen-stimulated mast cells via the modulation of immune responses. In addition, exposure to TCE and PCE may lead to the augmentation of allergic diseases.

Background: The clinical efficacy of combination therapy comprising a long acting beta(2)-agonist (LABA) and corticosteroid is widely recognized for the treatment of adult asthma. Here we examine the effect of salmeterol xinafoate (SX) and fluticasone propionate (FP) alone and in combination on the immunological activation of human cultured mast cells (HCMC)in vitro., Methods: HCMC were passively sensitized with IgE antibody and then activated by challenging with anti-IgE antibody. The effect of drugs on the activation of mast cells was examined by measuring the amount of released chemical mediators (histamine, leukotrienes (LT) and prostaglandin D(2) (PGD(2))) and granulocyte macrophage colony stimulating factor (GM-CSF)., Results: The release of each chemical mediator was inhibited by 10-9-10-8M SX but not by 10-10-10-7M FP. The production of GM-CSF was inhibited by a concentration of 10-8M in both drugs and the inhibition was augmented by combined treatment with 10-11M of each drug., Conclusions: The immunological release of chemical mediators (histamine, LT, PGD(2)) from HCMC was inhibited by SX but not by FP. SX and FP inhibited the production of GM-CSF by HCMC and both drug showed synergistic inhibition in the production of GM-CSF.

Background: Matrix metalloproteinases (MMPs) have been reported to play crucial roles in the migration of inflammatory cells through basement membrane components. To confirm the role of mast cells as a source of MMPs, we investigated the production of MMP and its pathway in human cultured mast cells (HCMC). We also investigated the production of tissue inhibitors of metalloproteinase (TIMPs)., Methods: HCMC was stimulated with phorbor 12-miristate 13-acetate (PMA) and/or calcium ionophore A23187 (A23187), and the resulting MMP production was evaluated by gelatin zymography and western blotting. Expression of MMP and TIMP mRNA was also examined. Granulocyte macrophage-colony stimulating factor (GM-CSF) was measured by ELISA and activation of extracellular signal-regulated kinase (ERK) was evaluated by western blotting., Results: We detected the de novo synthesis of MMP-9 in HCMC after stimulation with PMA and found that the synthesis was mediated through protein kinase C-mitogen activated protein kinase kinase (MEK)-ERK pathway. The MMP-9 production induced by PMA was suppressed by simultaneous treatment with A23187, whereas GM-CSF production was potentiated. We also detected the expression of mRNA for membrane-type 1 (MT1)-MMP, TIMP-1 and TIMP-2 after stimulation with PMA. Glucocorticoids and flavonoids inhibited MMP-9 production, and TIMPs and MMP inhibitors inhibited the gelatinolytic activity of mast cell-derived MMP-9. Furthermore, phenylmethylsulfonyl fluoride, a protease inhibitor, inhibited the conversion from proMMP-9 to active MMP-9., Conclusions: These results suggest that the human mast cell is a leading member of MMP production, and the production, activation and activity are controllable by pharmacological agents.

Integrin alphaIIb, a well-known marker of megakaryocyte-platelet lineage, has been recently recognized on hemopoietic progenitors. We now demonstrate that integrin alphaIIbbeta3 is highly expressed on mouse and human mast cells including mouse bone marrow-derived mast cells, peritoneal mast cells, and human cord blood-derived mast cells, and that its binding to extracellular matrix proteins leads to enhancement of biological functions of mast cells in concert with various stimuli. With exposure to various stimuli, including cross-linking of FcepsilonRI and stem cell factor, mast cells adhered to extracellular matrix proteins such as fibrinogen and von Willebrand factor in an integrin alphaIIbbeta3-dependent manner. In addition, the binding of mast cells to fibrinogen enhanced proliferation, cytokine production, and migration and induced uptake of soluble fibrinogen in response to stem cell factor stimulation, implicating integrin alphaIIbbeta3 in a variety of mast cell functions. In conclusion, mouse and human mast cells express functional integrin alphaIIbbeta3.

In the present study, we examined the inhibitory effects of the beta2-adrenoceptor agonists isoproterenol, salbutamol, fenoterol, and clenbuterol, on the release of chemical mediators from cultured human mast cells after prolonged treatment with the agonists. Although preincubation of sensitized mast cells for 10 min with beta2-adrenoceptor agonists potently inhibited mediator release, prolongation of the preincubation period up to 240 min attenuated the inhibition. The attenuation of histamine release inhibition was potent when compared with that of prostaglandin D2 (PGD2) and cysteinyl leukotriene (LT) release inhibition. In contrast, forskolin inhibited mediator release and the inhibition increased gradually in proportion to the preincubation period. The reduced inhibition by the beta2-adrenoceptor agonists was compensated for by simultaneous treatment with cholera toxin. The beta2-adrenoceptor agonists elevated intracellular cAMP levels after 10-min incubation and the elevated levels were almost comparable to those after 240-min incubation. Forskolin elevated the intracellular cAMP levels more potently after incubation for 240 min than after 10 min. When mast cells were incubated for 3 d with the beta2-adrenoceptor agonists, similar attenuation of mediator release inhibition was observed. Elevation of intracellular cAMP levels was also attenuated, although beta2-adrenoceptor mRNA expression was potentiated. The present results collectively indicate that the attenuation of mediator release inhibition by beta2-adrenoceptor agonists under the present experimental conditions involves uncoupling between beta2-adrenoceptors and Gs proteins. Furthermore, the beta2-adrenoceptor desensitization causes differential attenuating effects on the inhibition of histamine, PGD2, and LT release, suggesting that downstream events involved in each inhibitory pathway have different sensitivity to receptor desensitization.

Tea contains a variety of bioactive compounds. In this study, we show that two O-methylated catechins, (-)-epigallocatechin-3-O-(3-O-methyl) gallate and (-)-epigallocatechin-3-O-(4-O-methyl) gallate, inhibit in vivo mast cell-dependent allergic reactions more potently than their nonmethylated form, (-)-epigallocatechin-3-O-gallate. Consistent with this, these O-methylated catechins inhibit IgE/Ag-induced activation of mouse mast cells: histamine release, leukotriene release, and cytokine production and secretion were all inhibited. As a molecular basis for the catechin-mediated inhibition of mast cell activation, Lyn, Syk, and Bruton's tyrosine kinase, the protein tyrosine kinases, known to be critical for early activation events, are shown to be inhibited by the O-methylated catechins. In vitro kinase assays using purified proteins show that the O-methylated catechins can directly inhibit the above protein tyrosine kinases. These catechins inhibit IgE/Ag-induced calcium response as well as the activation of downstream serine/threonine kinases such as Akt and c-Jun N-terminal kinase. These observations for the first time have revealed the molecular mechanisms of antiallergic effects of tea-derived catechins.

Resveratrol is a natural phytoalexin occurring in grapes, vines and peanuts and possesses both antitumor and antioxidation capabilities. Its chemoprotective actions are attributed partially to its anti-inflammation effect. The present study is aimed at checking the inhibitory actions of resveratrol on the release of mediators from bone marrow-derived mouse mast cells (BMMC) in vitro. Mast cells were prepared by isolating bone marrow cells from intact mice femora and culturing them for 4 weeks in the presence of IL-3 and FCS. The release reaction was triggered by IgE or calcium ionophore (A23187). Mediated by IgE, the release of histamine and tumor necrosis factor-alpha was significantly inhibited by resveratrol at a concentration of 100 microM; IgE-mediated release of leukotrienes and prostaglandin D (2) was also strongly suppressed at concentrations of 100 and 10 microM. Also, A23187-mediated release of histamine and leukotrienes release was strongly reduced by resveratrol at concentrations of 100 and 10 microM, respectively. Resveratrol exhibited its behavior without a significant cytotoxic activity against mast cells. In conclusion, resveratrol is a potent non-selective inhibitor of mediator release from activated mast cells and deserves further investigation of its biological modulations.

To investigate the effect of flavonoids on the activation of p72(syk) (Syk) protein tyrosine kinase which plays a pivotal role in the high affinity IgE receptor-mediated degranulation of mast cells, we picked out 10 flavonoids, classified them into 4 series, and examined their effects on the activation of Syk and on the degranulation of human mast cells. Flavones and flavonols showed clear inhibition, whereas flavanones and isoflavones had either weak or no effect on Syk enzymatic activity induced by amino acid peptide corresponding to the activation loop domain and on IgE-dependent degranulation of human cultured mast cells (HCMC). On the basis of calculated logP (ClogP) values as a prediction of compound lipophilicity, some flavonoids were speculated to have low lipophilicity, the reason for poor cell permeability. A significant relationship was observed between the inhibition of Syk activity and HCMC degranulation attributable to flavonoids when the ClogP values of the compounds were taken into account (r(2)=0.89). These results suggested that the impairment of mast cell degranulation by several flavonoids classified into flavones and flavonols might be mediated via inhibition of the intracellular activation of Syk.

Coculture of mouse bone marrow-derived immature mast cells (BMMC) with Swiss 3T3 fibroblasts in the presence of stem cell factor (SCF) promotes morphological and functional maturation toward a connective tissue mast cell (CTMC)-like phenotype, which is accompanied by increased expression of several unique genes. Here we report the molecular identification of one of them, mast cell maturation-associated inducible gene (MMIG)-1. The MMIG-1 cDNA encodes a 117-kDa cytosolic protein that comprises an N-terminal PYRIN domain, a central nucleotide-binding domain, and nine C-terminal leucine-rich repeats. MMIG-1 shows >85% sequence similarity to human cryopyrin/PYPAF1, a causal gene for familial cold urticaria and Muckle-Wells syndrome. MMIG-1 was distributed in the cytosol of CTMC-like differentiated BMMC. MMIG-1 underwent alternative splicing in the leucine-rich repeats and each variant was induced differently in BMMC during coculture. Moreover, its expression was increased in the ears of mice with experimental atopic dermatitis. Thus, MMIG-1, a likely mouse PYPAF1 ortholog, may play a role in mast cell-directed inflammatory diseases.

Spleen tyrosine kinase (Syk) tyrosine kinase plays essential roles in receptors for Fc portion of immunoglobulins and B cell receptor complex signaling in various inflammatory cells; therefore, inhibitors of Syk kinase may show potential as antiasthmatic/allergic therapeutics. We identified 2-[7-(3,4-dimethoxyphenyl)-imidazo[1,2-c]pyrimidin-5-ylamino]-nicotinamide dihydrochloride (BAY 61-3606), a potent (Ki = 7.5 nM) and selective inhibitor of Syk kinase. BAY 61-3606 inhibited not only degranulation (IC50 values between 5 and 46 nM) but also lipid mediator and cytokine synthesis in mast cells. BAY 61-3606 was highly efficacious in basophils obtained from healthy human subjects (IC50 = 10 nM) and seems to be at least as potent in basophils obtained from atopic (high serum IgE) subjects (IC50 = 8.1 nM). B cell receptor activation and receptors for Fc portion of IgG signaling in eosinophils and monocytes were also potently suppressed by BAY 61-3606. Oral administration of BAY 61-3606 to rats significantly suppressed antigen-induced passive cutaneous anaphylactic reaction, bronchoconstriction, and bronchial edema at 3 mg/kg. Furthermore, BAY 61-3606 attenuated antigen-induced airway inflammation in rats. Based on these anti-inflammatory effects of BAY 61-3606 both in vitro and in vivo, it was demonstrated that Syk may play a very critical role in the pathogenesis of allergic reactions.

Rodent mast cells (MCs) are common experimental tools but are somewhat different from their human counterparts in their responses to certain cytokines and drugs. We examined the expression of more than 10 000 distinct genes in human and mouse cultured MCs using high-density oligonucleotide probe arrays to find molecules similarly regulated and expressed by the 2 MC types. After stimulation via high-affinity Fcepsilon receptor I (FcepsilonRI), the transcriptional levels of several CC chemokines were markedly increased, and I-309 (CCL1), macrophage inflammatory protein-1alpha (MIP-1alpha) (CCL3) and MIP-1beta (CCL4) were found among the 10 most increased human and mouse transcripts from approximately 12 000 genes (including some expressed sequence tags). In addition, a costimulatory molecule that was originally found on the membrane of activated T cells, 4-1BB (CD137), was found among the 10 most increased transcripts. The FcepsilonRI-induced expression of CC chemokines and 4-1BB was also detected at the protein level in both MC types. The conservation of these responses suggests that MCs play a crucial role in recruitment of various CCR-expressing cells into the tissue in a manner dependent on immunoglobin E, and that FcepsilonRI-mediated induction of several CC chemokines and 4-1BB is highly conserved between human and mouse. Interspecies comparison studies at the whole genome expression level should be useful for the interpretation of experimental data obtained in animal models of human pathobiology.

Group IID secretory phospholipase A(2) (sPLA(2)-IID), a heparin-binding sPLA(2) that is closely related to sPLA(2)-IIA, augments stimulus-induced cellular arachidonate release in a manner similar to sPLA(2)-IIA. Here we identified the residues of sPLA(2)-IID that are responsible for heparanoid binding, are and therefore essential for cellular function. Mutating four cationic residues in the C-terminal portion of sPLA(2)-IID resulted in abolition of its ability to associate with cell surface heparan sulfate and to enhance stimulus-induced delayed arachidonate release, cyclooxygenase-2 induction, and prostaglandin generation in 293 cell transfectants. As compared with several other group II subfamily sPLA(2)s, which were equally active on A23187- and IL-1-primed cellular membranes, sPLA(2)-IID showed apparent preference for A23187-primed membranes. Several human colon carcinoma cell lines expressed sPLA(2)-IID and sPLA(2)-X constitutively, the former of which was negatively regulated by IL-1. sPLA(2)-IID, but not other sPLA(2) isozymes, was expressed in human cord blood-derived mast cells. The expression of sPLA(2)-IID was significantly altered in several tissues of mice with experimental inflammation. These results indicate that sPLA(2)-IID may be involved in inflammation in cell- and tissue-specific manners under particular conditions.

We investigated the role of Bruton's tyrosine kinase (Btk) in FcepsilonRI-dependent activation of mouse mast cells, using xid and btk null mutant mice. Unlike B cell development, mast cell development is apparently normal in these btk mutant mice. However, mast cells derived from these mice exhibited significant abnormalities in FcepsilonRI-dependent function. xid mice primed with anti-dinitrophenyl monoclonal IgE antibody exhibited mildly diminished early-phase and severely blunted late-phase anaphylactic reactions in response to antigen challenge in vivo. Consistent with this finding, cultured mast cells derived from the bone marrow cells of xid or btk null mice exhibited mild impairments in degranulation, and more profound defects in the production of several cytokines, upon FcepsilonRI cross-linking. Moreover, the transcriptional activities of these cytokine genes were severely reduced in FcepsilonRI-stimulated btk mutant mast cells. The specificity of these effects of btk mutations was confirmed by the improvement in the ability of btk mutant mast cells to degranulate and to secrete cytokines after the retroviral transfer of wild-type btk cDNA, but not of vector or kinase-dead btk cDNA. Retroviral transfer of Emt (= Itk/Tsk), Btk's closest relative, also partially improved the ability of btk mutant mast cells to secrete mediators. Taken together, these results demonstrate an important role for Btk in the full expression of FcepsilonRI signal transduction in mast cells.

The purpose of this review is to summarize the role of prostaglandins (PGs) in allergic inflammation and to know the value of PGs, as a target molecule for an anti-allergic drug. PGD2 is the major PG produced by the cyclooxygenase pathway in mast cells. Our and others findings indicate that PGD2 is one of the potent allergic inflammatory mediators and must be a target molecule of anti-allergic agent. From our data, one of PGD2 receptor antagonists show clear inhibition of airway hypersensitivity caused by allergic reaction. Concerning the role of PGE2 in allergic inflammation, conflicting results have been reported. Many experimental data suggest an individual role of each PGE2 receptor, EP1, EP2, EP3 and EP4 in allergic reaction. Our results indicate the protective action of PGE2 on allergic reaction via EP3. In addition, one of EP3 agonists clearly inhibits the allergic airway inflammation. These findings indicate the value of EP3 agonists as an anti-allergic agent. In addition, some investigators including us reported that PGI2 plays an important role for the protection of the onset of allergic reaction. However, the efficacy of PGI2 analogue as an anti-allergic agent is not yet fully investigated. Finally, the role of thromboxane A2 (TxA2) in allergic reaction is discussed. Our experimental results suggest a different participation of TxA2 in allergic reaction of airway and skin. In this review, the role of PGs in allergic inflammation is summarized and the value of PGs as a target molecule for developing a new anti-allergic agent will be discussed. [ABSTRACT FROM AUTHOR]

Group IID secretory phospholipase A2 (sPLA2 -IID), a heparin-binding sPLA2 that is closely related to sPLA2 -IIA, augments stimulus-induced cellular arachidonate release in a manner similar to sPLA2 -IIA. Here we identified the residues of sPLA2 -IID that are responsible for heparanoid binding, are and therefore essential for cellular function. Mutating four cationic residues in the C-terminal portion of sPLA2 -IID resulted in abolition of its ability to associate with cell surface heparan sulfate and to enhance stimulus-induced delayed arachidonate release, cyclooxygenase-2 induction, and prostaglandin generation in 293 cell transfectants. As compared with several other group II subfamily sPLA2 s, which were equally active on A23187- and IL-1-primed cellular membranes, sPLA2 -IID showed apparent preference for A23187-primed membranes. Several human colon carcinoma cell lines expressed sPLA2 -IID and sPLA2 -X constitutively, the former of which was negatively regulated by IL-1. sPLA2 -IID, but not other sPLA2 isozymes, was expressed in human cord blood-derived mast cells. The expression of sPLA2 -IID was significantly altered in several tissues of mice with experimental inflammation. These results indicate that sPLA2 -IID may be involved in inflammation in cell- and tissue-specific manners under particular conditions. [ABSTRACT FROM AUTHOR]

Epicutaneous antigen challenge in passively sensitized mice with IgE produces a biphasic cutaneous response which peaks 1 h (immediate-phase reaction) and 24 h (late-phase reaction; LPR) after the antigen challenge. In this model, anaphylactic degranulation and interleukin 6 (IL-6) expression between 4 and 8 h are observed in resident mast cells as the preceding stage of LPR. Prednisolone at a dose of 3 mg kg[sup –1] clearly inhibited the LPR when administered 2 h before and 4 h after antigen challenge. Slight or no inhibition of LPR was observed by prednisolone administered 6–12 h after challenge. Histologically, prednisolone treatment 2 h before antigen challenge completely inhibited edema and inflammatory cell infiltration, while treatment at 6 h did not at all. In order to investigate the relationship between inhibition of LPR by prednisolone and mast cell activation, the effects of prednisolone on degranulation of mast cells and IL-6 expression in mast cells were investigated. 8 h after antigen challenge, prednisolone clearly inhibited the increase in the number of anaphylactic degranulated and IL-6-positive mast cells by administration 2 h before challenge, but did not affect it by administration 6 h after challenge. These data indicate that the inhibitory mechanism of prednisolone on LPR, at least, involves the inhibition of mast cell activation before LPR.Copyright © 2001 S. Karger AG, Basel. [ABSTRACT FROM AUTHOR]

Background: Human cultured mast cells release histamine, sulfidoleukotrienes and prostaglandin D[sub 2] , and produce some cytokines after an allergic reaction. As concerns the mitogen-activated protein kinase family, extracellular signal-regulated kinases, c-Jun NH[sub 2] -terminal kinases and p38 mitogen-activated protein kinase are known. The present study has been conducted to investigate the role of mitogen-activated protein kinase in the mediator release from human cultured mast cells. Material and Methods: To investigate the role of these kinase pathways, we examined the participation of this kinase activation for mediator release by making use of U0126, a signal transduction, extracellular signal-regulated kinase pathway inhibitor, and SB203580, a p38 mitogen-activated protein kinase pathway inhibitor. Results: Histamine release is not related to mitogen-activated protein kinase pathways. Sulfidoleukotrienes and prostaglandin D[sub 2] release are mediated by signal transduction, extracellular signal-regulated kinase pathway. Granulocyte macrophage-colony-stimulating factor production may be induced by both signal transduction, extracellular signal-regulated kinase and c-Jun NH[sub 2] -terminal kinase pathways. Conclusion: Mitogen-activated protein kinase pathways play an important role in mediator release from human mast cells.Copyright © 2001 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Abstract: In previous report, we have shown that trichloroethylene (TCE) increases histamine release and inflammatory cytokine production from antigen-stimulated mast cells. In this study, we examined the enhancing effect of a small amount of TCE ingestion from drinking water on antigen-stimulated allergic responses. After exposure of Wistar rats to TCE ingestion for 2 or 4 weeks, we performed a passive cutaneous anaphylaxis (PCA) reaction. TCE ingestion for 2 and 4 weeks enhanced PCA reaction in a dose-dependent manner. On histological examination, TCE ingestion for 2 weeks exacerbated inflammation characterized by infiltration of lymphocytes and accumulation of mast cells around the vessel in the skin. After TCE ingestion for 4 weeks, the mesenteric lymph nodes (MLNs) showed increase of the size and wet weight, and germinal centers changed distinctly. The interleukin-4 (IL-4) mRNA levels on spleen, MLNs and leukocytes were increased. Moreover, serum total IgE levels of TCE ingestion increased in a time-dependent manner. Our results suggest that TCE ingestion induces pro-inflammatory responses and causes Th1/Th2-type helper T-cell imbalance. And more, a small amount of TCE ingestion may lead to the initiation and acceleration of type I allergic reaction. [Copyright &y& Elsevier]

Abstract: Previously, we observed that tetrachloroethylene (perchloroethylene, PCE) increased histamine release and inflammatory mediator production from antigen-stimulated mast cells. In this study, we examined the enhancing effect of low concentrations of PCE in drinking water on antigen-stimulated allergic responses. After exposure of Wistar rats to PCE in drinking water for 2 or 4 weeks, we performed a passive cutaneous anaphylaxis (PCA) reaction. PCE exposure for 4 weeks enhanced PCA reaction in a dose-dependent manner. In pathological studies, PCE exposure for 2 weeks exacerbated inflammation characterized by infiltration of lymphocytes and accumulation of mast cells around the vessel. Non-purified mast cells (NPMCs) from rats treated with 1mg/L PCE in drinking water for 2 weeks increased antigen-stimulated histamine release. Furthermore, the leukocytes of rats treated with PCE in drinking water for 4 weeks showed increased interleukin (IL)-4 expression. The mechanism of enhancing the PCA reaction is assumed to be that PCE increases IL-4 production and PCE causes T helper (Th) 1/Th2-type helper T-cell imbalance and increases histamine release from excessively accumulated mast cells. The results suggest that the intake of PCE in drinking water, even at a low concentration, leads to the initiation and acceleration of allergic diseases. [Copyright &y& Elsevier]

Abstract Background: Although β-adrenergic receptor agonists inhibit antigen-induced release of histamine, leukotrienes and prostaglandin D2 from human lung fragments, dispersed human lung mast cells and human skin mast cells, subcellular mechanisms for the inhibition of histamine release by β-adrenergic receptor agonists are not well delineated. The aim of the present study was to investigate the inhibitory mechanisms of β-adrenergic receptor agonists for human mast cell histamine release using human cultured mast cells. Methods: Human mast cells were obtained by culturing umbilical cord blood CD34+ cells in the presence of stem cell factor and interleukin-6. Cultured mast cells were sensitized with human myeloma IgE and stimulated with antihuman IgE. Results: Stimulation of mast cells induced the elevation of intracellular cytosolic free Ca2+ concentrations ([Ca 2+ ] i ) and the translocation of protein kinase C (PKC) from the cytosol to the cell membrane, followed by the release of stored histamine. Isoproterenol, sal­butamol and dibutyryl cAMP inhibited both the histamine release and PKC translocation, whereas they failed to affect the elevation of [Ca 2+ ] i . H-89, a protein kinase A (PKA) inhibitor, abrogated the inhibition. Conclusions: The present results suggest that PKA activation induced by β-adrenergic receptor agonists plays a crucial role in inhibiting IgE-mediated histamine release from human cultured mast cells through suppressing PKC translocation. [ABSTRACT FROM AUTHOR]

Compound 48/80 induced scratching behavior in BALB/c mice, and the role of mast cell mediators in this behavior was examined. Mouse scratching behavior was detected and evaluated using a new apparatus, MicroAct. Compound 48/80 increased the incidence of scratching behavior and scratching time in a dose-dependent manner, accompanied by a potent activation of mast cells and a potent increase in vascular permeability. Dibucaine and μ-opioid receptor antagonists inhibited the scratching behavior. Although histamine H1 receptor antagonists potently inhibited the vascular permeability increase, they did not affect the scratching behavior. Methysergide inhibited the scratching behavior slightly without affecting the vascular permeability increase, whereas cyproheptadine inhibited both. A cyclooxygenase inhibitor, a 5-lipoxygenase-activating protein inhibitor and a PAF receptor antagonist did not affect the scratching behavior. High doses of serotonin induced scratching behavior less frequently than did compound 48/80. Furthermore, mast cell-deficient WBB6F1-W/Wv mice exhibited frequent scratching behavior after injection of compound 48/80. These results clearly indicate that compound 48/80 can induce scratching behavior in mice independent of mast cell mediators. [Copyright &y& Elsevier]