Degranulation In RBL-2H3 Cells Research Articles

Inhibitory Activity of Citrus Madurensis Ripe Fruits Extract on Antigen-induced Degranulation in RBL-2H3 Cells

The purpose of this study was to search edible ripe Citrus fruits which are applicable for functional food materials as juice, tea and/or jam with sweet taste and rich aroma. A fifty percent ethanolic extract (CMR-ext) obtained from the edible ripe fruit of Citrus madurensis exhibited an inhibitory activity of antigen-induced degranulation in anti-dinitrophenyl (DNP) IgE antibody sensitized rat basophilic leukemia (RBL) -2H3 cells. The inhibitory effect of the CMR-ext on degranulation in RBL-2H3 cells was attributable to 3’,5’-di-C-β-glucopyranosylphloretin (1) which is a constituent of C. madurensis. The effect of 1 on Akt and mitogen-activated protein kinases (MAPK) phosphorylation was examined in RBL-2H3 cells. Western blot analysis revealed that 1 (50 μM) inhibited the degranulation by suppression of Akt and p38 phosphorylation.

Imidacloprid inhibits IgE-mediated RBL-2H3 cell degranulation and passive cutaneous anaphylaxis.

BackgroundImidacloprid has been commonly used as a pesticide for crop protection and acts as nicotinic acetylcholine receptor agonists. Little information about the relationship between imidacloprid and allergy is available.ObjectiveThis study aims to examine the effects of imidacoprid on IgE-mediated mast cell activation.MethodsThe rat basophilic leukemia cell line RBL-2H3 (RBL-2H3 cells) were treated with 10-3 – 10-12 mol/L imidacloprid, followed by measuring the mediator production, influx of Ca2+ in IgE-activated RBL-2H3 cells, and the possible effects of imidacoprid on anti-dinitrophenyl IgE-induced passive cutaneous anaphylaxis (PCA).ResultsIt was shown that imidacoprid suppressed the production of histamine, β-hexosaminidase, leukotriene C4, interleukin-6, tumor necrosis factor-α, and Ca2+ mobilization in IgE-activated RBL-2H3 cells and decreased vascular extravasation in IgE-induced PCA.ConclusionIt is the first time to show that imidacloprid suppressed the activation of RBL-2H3 cells.

Vam3, a Compound Derived from Vitis amurensis Rupr., Attenuated Colitis-Related Tumorigenesis by Inhibiting NF-κB Signaling Pathway.

Background: Chronic inflammation is one of the important mediators of colitis-related colon cancer (CRC). Abundant mast cells (MCs) were observed in the tumor microenvironment and mediators released upon MC activation play an important role in the process of chronic inflammation. Previously, we found that activation of intestine mucosal MCs recruited and modulated the inflammatory CD11b+Gr1+ cells to promote the CRC development. In the current study we investigated the effects of Vam3, a resveratrol dimer with potent anti-inflammatory effects, on CRC development.Methods: RBL-2H3 cells, a basophilic leukemia cell line, were pretreated with 2.5 or 5 µM Vam3 and then stimulated with dinitrophenol-conjugated bovine serum albumin (DNP-BSA) plus lipopolysaccharide (LPS). The MC degranulation was determined by measuring β-hexosaminidase release. Generation of TNF-α and IL-6 in RBL-2H3 cells or in peritoneal macrophages was determined by ELISA and real-time qPCR. NF-κB p65 and phospho-NF-κB p65 expression was determined by Western blotting. NF-κB activity in RAW264.7 cells was determined by luciferase reporter assay. CRC was induced in C57BL/6 mice by intraperitoneal injection of azoxymethane (AOM), followed by oral exposure to dextran sodium sulfate (DSS). Vam3 at 50 mg/kg, or disodium cromoglycate (DSCG, MC stabilizer) at 100 mg/kg, or vehicle were administrated to the mice 4 weeks after DSS withdrawal. Levels of TNF-α, IL-6, and mouse MC protease-1 were determined by ELISA. Infiltration of CD11b+Gr1+ cells was determined by flow cytometry analysis. One-way ANOVA was used to compare difference between groups.Results: Pretreatment with Vam3 significantly inhibited RBL-2H3 cell degranulation and inflammatory cytokine production from RBL-2H3 cells and from peritoneal macrophages. After Vam3 treatment, NF-κB activity in RAW264.7 cells, and expressions of phospho-NF-κB p65 in RBL-2H3 cells and in peritoneal macrophages were significantly down-regulated. In the AOM plus DSS-induced CRC murine model, the Vam3 and DSCG-treated mice had less tumor numbers than those treated with vehicle. Expression of phospho-NF-κB p65, production of inflammatory cytokines, and infiltration of MCs and CD11b+Gr1+ cells were attenuated in the Vam3-treated mice.Conclusion: Vam3 treatment could attenuate the CRC development. This effect may be due to its inhibition on NF-κB signaling pathway in MCs and macrophages of the inflamed intestines.

Inhibitory Effects of Echinochrome A, Isolated from Shell of the Sea Urchin Anthocidaris crassispina, on Antigen-Stimulated Degranulation in Rat Basophilic Leukemia RBL-2H3 Cells through Suppression of Lyn Activation

Echinochrome A (Echi-A) was isolated from the sea urchin Anthocidaris crassispina and its structure determined using ID and 2D-NMR. In the present study, we examined the inhibitory effect of Echi-A on antigen-stimulated degranulation in rat basophilic leukemia RBL-2H3 cells, which were suppressed in a dose dependent manner.. The antigens bind to the high affinity immunoglobulin E receptor, which is expressed on the surface of mast cells and basophils and activate intracellular signal transduction, resulting in the release of biologically active mediators such as histamine. In order to disclose the inhibitory mechanisms of degranulation by Echi-A, we examined the elevation in intracellular C²⁺ concentration ([Ca²⁺]i), production levels of intracellular reactive oxygen species (ROS) and early intracellular signaling events. Both elevation of [Ca²⁺] and intracellular ROS production were markedly suppressed in cells treated with Echi-A. Echi-A also suppressed the activation of Lyn, Syk, and PLCyl/2 in antigen-stimulated cells. These results indicated that inhibition of antigen-stimulated degranulation in RBL-2H3 cells by Echi-A is mainly due to the inactivation of Lyn/Syk/PLCy signaling pathways. Our findings suggest that Echi-A could be a beneficial agent for alleviating the symptoms of type I allergy.

Yeast Ca2 +-signal transduction inhibitors isolated from Dominican amber prevent the degranulation of RBL-2H3 cells through the inhibition of Ca2 +-influx

A new norlabdane compound, named kujigamberol has previously been isolated from Kuji amber (but not from Baltic amber) by activity guided fractionation. However, there has been no study of biological compounds in Dominican amber. Biological activities were examined using the hypersensitive mutant yeast (zds1Δ erg3Δ pdr1Δ pdr3Δ) with respect to Ca2+-signal transduction, enzymes and rat basophilic leukemia (RBL)-2H3 cells. The structures were elucidated on the basis of spectral analysis including high resolution (HR)-EI-MS, 1D NMR and 2D NMR. Three diterpenoid compounds, 5(10)-halimen-15-oic acid (1), 3-cleroden-15-oic acid (2) and 8-labden-15-oic acid (3), which are different from the bioactive compounds in Kuji and Baltic ambers, were isolated from Dominican amber. They inhibited both calcineurin (CN) (IC50=40.0, 21.2 and 34.2μM) and glycogen synthase kinase-3β (GSK-3β) (IC50=48.9, 43.8 and 41.1μM) which are involved in the growth restored activity against the mutant yeast. The most abundant compound 2 showed inhibitory activity against both degranulation and Ca2+-influx in RBL-2H3 cells. The compounds having the growth restoring activity against the mutant yeast have potential as anti-allergic compounds.

Cinnamaldehyde derivatives inhibit degranulation and inflammatory mediator production in rat basophilic leukemia cells

Mast cells play a critical role in allergic diseases. Therefore, development of new therapeutic agents that suppresses the activation of mast cells may help prevent or treat allergic diseases. Here, we investigated the anti-allergic effects of 4-chloro-cinnamaldehyde and 4-trifluoro-cinnamaldehyde in RBL-2H3 cells. β-Hexosaminidase assays revealed that degranulation of RBL-2H3 cells was decreased following treatment with 60μM 4-chloro-cinnamaldehyde or 4-trifluoro-cinnamaldehyde. Moreover, quantitative real-time reverse transcription polymerase chain reaction showed that the relative expression levels of tumor necrosis factor-α, interleukin-4, and cyclooxygenase-2 mRNAs were decreased in RBL-2H3 cells treated with 4-chloro-cinnamaldehyde and 4-trifluoro-cinnamaldehyde in a concentration-dependent manner. Additionally, 4-chloro-cinnamaldehyde blocked the phosphorylation of MKKs and MAPKs. These data clearly suggested that 4-chloro-cinnamaldehyde and 4-trifluoro-cinnamaldehyde had inhibitory effects on the inflammatory responses of mast cells and may have potential as novel therapeutic agents for the prevention or treatment of allergic diseases.

Structure–activity relationships of vanillic acid ester analogs in inhibitory effect of antigen-mediated degranulation in rat basophilic leukemia RBL-2H3 cells

Methyl vanillate (1) showed strong degranulation inhibitory activity among vanillin derivatives tested. In order to find structure–activity relationships for developing anti-allergic agents with simple structures and potent activity, we synthesized several vanillic acid (VA) ester derivatives with C1–C4 and C8 alkyl chains and evaluated their degranulation inhibitory activities. The most active compound of VA ester derivatives was derivative 5 with a C4 straight alkyl chain, and derivative 5 exhibited approximately three-fold greater inhibitory activity than that of 1. Moreover, we designed 8 types of analogs based on 5, and we found that the minimum structure for potent degranulation inhibitory activity requires direct connection of the butyl ester moiety on the benzene ring and at least one hydroxyl group on the benzene ring. Butyl meta or para hydroxyl benzoate (10 or 11) has a simpler structure than that of 5 and exhibited more potent degranulation inhibitory activity than that of 5.

Effects of Viola yedoensis Makino anti-itching compound on degranulation and cytokine generation in RBL-2H3 mast cells

Ethnopharmacological relevanceThe Chinese herb compound prescription Viola yedoensis Makino Anti-itching Compound (VYAC), which consists of Viola yedoensis Makino, herb, Sophora flavescens Aiton, root, and Dictamnus dasycarpus Turcz, root and rhizome, has been traditionally used to treat various skin allergic inflammatory diseases in clinic. Aim of the studyThe aim of this study is to investigate the effects of VYAC on degranulation and to determine its anti-inflammatory mechanism in RBL-2H3 mast cells. Materials and methodsVYAC was extracted with water-coction extraction (Shufen et al., 2012). The aqueous extracts were concentrated in vacuum under reduced pressure and lyophilized using a freeze dryer, and lyophilized powder was obtained. MTT was used to evaluate the cytotoxic of VYAC on RBL-2H3 cells. Degranulation was carried out with RBL-2H3 cell model, which was stimulated with A23187 plus PMA. β-Hexosaminidase and histamine were measured to evaluate degranulation. The mRNA levels of inflammation cytokines (IL-1β, TNF-α, IL-6, and iNOS) were investigated by RT-PCR to explain the anti-inflammatory mechanism of VYAC. ResultsVYAC did not show cytotoxic effect on RBL-2H3 cells in the range of 25–400μg/mL. A higher dose of VYAC (800μg/mL) showed significant cytotoxicity (P<0.05). VYAC could significantly inhibit β-hexosaminidase and histamine release when treated with 100, 200, and 400μg/mL (P<0.05), but could not significantly inhibit β-Hexosaminidase and histamine release when treated with 25 and 50μg/mL (p>0.05). The mRNA levels of inflammatory cytokines (TNF-α, IL-1β, IL-6, and iNOS) could significantly decrease when treated with 200 and 400μg/mL (P<0.05) of VYAC, which were associated with the development of inflammation. ConclusionsResults showed that VYAC inhibited β-hexosaminidase and histamine release, which was inhibit A23187 plus PMA stimulated RBL-2H3 cell degranulation and downregulated inflammatory cytokines (IL-1β, TNF-α, IL-6, and iNOS) expression to block inflammatory development.

A high content screening method for detection of anaphylactoid reaction of injection formulations

Anaphylactoid reaction (AR) is the most common adverse reaction of injection formulations, however, there are obvious drawbacks in available methods for AR detection. A novel in vitro detection method for AR was established based on fluorescent labeling and high content screen (HCS) system in present study. With the use of RBL-2H3 cells degranulation model, positive cell count was determined with specific cellular membrane fluorescent dye FM4-64 labeling vesicle recycle, and total cells count was determined with specific nucleus fluorescent dye Hochest 3334, and then the ratio of cells degranulation after drug stimulation was calculated. In order to verify the reliability of this HCS method, positive drug Compound 48/80 was first used to confirm the consistence of HCS method with the traditional β-hexosaminidase release test and the Evans blue staining ears test in mice. The results showed high consistence between HCS method and traditional testing methods, and the HCS method showed higher sensitivity than the other two tests. Then 30 samples of Danhong injection (DHI) with clinical allergy symptoms further were used to confirm the reliability of this HCS method. The HCS results showed high consistence with the clinical report, and the HCS method had the advantage in reducing the interference by drug color. Therefore, this HCS method is reliable, sensitive, simple and high-throughput method in detection of AR, applicable for the AR evaluation of injection formulations, and can provide guidance for safety of clinical application in clinical practice.

A novel in vitro co-culture model comprised of Caco-2/RBL-2H3 cells to evaluate anti-allergic effects of food factors through the intestine

The prevalence of type I allergic diseases such as food allergy and allergic rhinitis has increased. Therefore, many studies have focused on food factors with anti-allergic activities in recent years. In order to investigate the effect of food factors on mast cell activation, a RBL-2H3 cell monoculture system has been widely used, in which various food factors have been reported to inhibit degranulation of RBL-2H3 cells. However, some orally administered food factors do not interact directly with immune cells but do so indirectly through intestinal epithelial cells. In this report, we established a novel in vitro co-culture model to evaluate anti-allergic effects of orally administered food factors. The co-culture system, comprised of Caco-2 cells (apical component) and RBL-2H3 cells (basolateral component), was able to evaluate the effects of two flavonoids that are known to have the inhibitory effects on mast cell degranulation. Moreover, we evaluated the anti-allergic effects of Enterococcus faecalis strains that are not absorbed through the intestine. We identified two strains of lactic acid bacteria that had inhibitory effects on mast cell degranulation using this co-culture system and possessed anti-allergic properties in a passive cutaneous anaphylaxis model mouse. This novel in vitro co-culture model was applicable for finding food factors with anti-allergic effects and might be useful for examining its anti-allergic mechanisms.

Effect of concurrent administration of nobiletin and β-lactoglobulin on the symptoms of Japanese cedar pollinosis models in mice

Nobiletin and β-lactoglobulin were previously demonstrated to suppress mast cell de-granulation in vitro and in vivo. In this study, the multiple anti-allergic effects of both substances by simultaneous treatment were examined in order to determine if they could further mitigate allergic symptoms. Concurrent treatment with nobiletin and β-lactoglobulin effectively suppressed de-granulation of RBL-2H3 cells by inhibiting increases in intracellular Ca2+ concentration. Immunoblot analysis showed that phosphorylation of phosphoinositide 3-kinase, Akt, and Syk in RBL-2H3 cells was all inhibited by the concurrent presence of nobiletin and β-lactoglobulin. Moreover, concurrent administration of nobiletin and β-lactoglobulin reduced passive cutaneous anaphylaxis (PCA) reaction in mice compared with nobiletin or β-lactoglobulin administered alone. In addition, the concurrent administration of freeze-dried Citrus unshiu peel powder and yogurt whey containing these bioactive substances also reduced PCA reaction and allergic symptoms of a murine model of Japanese cedar pollinosis.

Atomic force microscopy study of ionomycin-induced degranulation in RBL-2H3 cells.

Mast cell degranulation is the typical anaphylaxis process of mast cells associated with the release of cytokines, eicosanoids and their secretory granules, which play very important roles in the allergic inflammatory response of the human body upon anaphylactogen stimulation. The calcium ionophore ionomycin is widely used as a degranulation induction agent for mast cell degranulation studies. In the present work, ionomycin-induced degranulation of RBL-2H3 basophilic leukemia cell line cells was investigated in vitro by high resolution atomic force microscopy (AFM). Ionomycin, which could increase the intracellular free Ca2+ level and β-Hexosaminidase release, was found to induce the formation of a kind of peculiar vesicles in the cytoplasm area of RBL-2H3 cells. Those vesicles induced by ionomycin would desintegrate to release a larger amount of granules surrounding RBL-2H3 cells by the controlling of F-actin. These results provide the precise morphological information of ionomycin-induced mast cell degranulation at nanoscale, which could benefit our understanding of ionomycin-induced mast cell anaphylaxis model and also validate the applicability of AFM for the detection of allergic inflammatory response in mast cells. SCANNING 38:525-534, 2016. © 2016 Wiley Periodicals, Inc.

A novel eremophilane lactone inhibits FcεRI-dependent release of pro-inflammatory mediators: structure-dependent bioactivity.

Allergic inflammation is primarily mediated by immune effector cells such as mast cells and basophils that release proinflammatory cytokines. Both mast cells and basophils are activated via their high affinity IgE receptor (FcεRI) which initiates the release of proinflammatory mediators such as histamine and tumor necrosis factor (TNF). Considerable effort has been focused on finding an effective basophil stabilizer that inhibits the activation of FcεRI-activated mediator release. Recently, eremophilane lactones, a novel family of sesquiterpene compound originally isolated from Petasites japonicas (Sieb. et Zucc.), have been described, and it has been postulated that they may have anti-inflammatory activity, particularly in allergic disease. Our objective was to determine the effect of two eremophilane lactones derived from 6β-angeloyloxy-3β,8-dihydroxyeremophil-7(11)-en-12,8β-olide (F-1) on immunoglobulin E (IgE)-dependent release of pro-inflammatory mediators by a basophil cell line, RBL-2H3, a model system for FcεRI-mediated activation of pro-inflammatory mediator release. The parent compound (F-1) was chemically modified to produce F-1a [6β-angeloyloxy-3β-benzoyloxy-8-hydroxyeremophil-7(11)-en-12,8β-olide] and F-1b [6β-angeloyloxy-3β,8-diacetoxyeremophil-7(11)-en-12,8β-olide]. RBL-2H3 cells were sensitized with DNP-specific IgE and then activated with DNP-BSA. The effect of these compounds on IgE-dependent basophil degranulation was assessed by measuring the release of β-hexosaminidase (b-hex). In addition, TNF release was measured via ELISA. The phenylacetyl reaction modified C-8 and produced F-1a whereas acetylation of F-1 produced F-1b. F-1a was not cytotoxic to RBL-2H3 cells even at 50 μM, but F-1b was slightly cytotoxic at 50 μM, reducing viability of the cells by approximately 15 %. Neither F-1a nor F-1b inhibited FcεRI-dependent activation of RBL-2H3 cells when the cells were pretreated for only 30 min with the compounds. However, 24 h pretreatment with F-1a inhibited antigen-dependent degranulation by as much as 60 % and TNF production by as much as 90 %. F-1b had no effect on RBL-2H3 activation via FcεRI. These results indicate that F-1a inhibits degranulation of RBL-2H3 cells activated via the high affinity IgE receptor, FcεRI, and that this effect is dependent upon hydroxylation of the third carbon.

Cooperative Interactions of Oligosaccharide and Peptide Moieties of a Glycopeptide Derived from IgE with Galectin-9

We previously showed that galectin-9 suppresses degranulation of mast cells through protein-glycan interaction with IgE. To elucidate the mechanism of the interaction in detail, we focused on identification and structural analysis of IgE glycans responsible for the galectin-9-induced suppression using mouse monoclonal IgE (TIB-141). TIB-141 in combination with the antigen induced degranulation of RBL-2H3 cells, which was almost completely inhibited by human and mouse galectin-9. Sequential digestion of TIB-141 with lysyl endopeptidase and trypsin resulted in the identification of a glycopeptide (H-Lys13-Try3; 48 amino acid residues) with a single N-linked oligosaccharide near the N terminus capable of neutralizing the effect of galectin-9 and another glycopeptide with two N-linked oligosaccharides (H-Lys13-Try1; 16 amino acid residues) having lower activity. Enzymatic elimination of the oligosaccharide chain from H-Lys13-Try3 and H-Lys13-Try1 completely abolished the activity. Removal of the C-terminal 38 amino acid residues of H-Lys13-Try3 with glutamyl endopeptidase, however, also resulted in loss of the activity. We determined the structures of N-linked oligosaccharides of H-Lys13-Try1. The galectin-9-binding fraction of pyridylaminated oligosaccharides contained asialo- and monosialylated bi/tri-antennary complex type oligosaccharides with a core fucose residue. The structures of the oligosaccharides were consistent with the sugar-binding specificity of galectin-9, whereas the nonbinding fraction contained monosialylated and disialylated biantennary complex type oligosaccharides with a core fucose residue. Although the oligosaccharides linked to H-Lys13-Try3 could not be fully characterized, these results indicate the possibility that cooperative binding of oligosaccharide and neighboring polypeptide structures of TIB-141 to galectin-9 affects the overall affinity and specificity of the IgE-lectin interaction.

Effects of Cymbidium Root Ethanol Extract on Atopic Dermatitis.

Cymbidium has known antibacterial and antiedema activity and has been used as an ingredient in cosmetics and fragrances. The effects of Cymbidium ethanol extract (CYM) on allergic response and the underlying mechanisms of action have not been reported. Therefore, the purpose of this study was to determine the effect of CYM on allergic responses. Topical application of CYM was effective against immunoglobulin E (IgE)/dinitrophenyl-conjugated bovine serum albumin- (DNP-BSA-) induced degranulation of RBL-2H3 cells and anaphylaxis in ICR mice. An allergic dermatitis-like mouse model was used to evaluate the therapeutic potential of CYM in vivo. Continuous application of 2,4-dinitrochlorobenzene (DNCB) not only induced dermatitis in ICR mice but also aggravated the skin lesioning. However, the application of CYM decreased skin lesion severity, scratching behavior, and IgE levels. In addition, CYM downregulated the expression of the proinflammatory cytokines interleukin- (IL-) 4, IL-13, and tumor necrosis factor- (TNF-) α. Studies of signal transduction pathways showed that CYM suppressed the phosphorylation of spleen tyrosine kinase (Syk), an upstream molecule. It also inhibited the phosphorylation of Akt, phospholipase C- (PLC-) γ, and mitogen-activated protein kinase kinase kinase (MEKK). These results indicate that CYM may be effective in preventing and reducing allergic response and may have therapeutic potential as an antiallergic agent in disorders such as atopic dermatitis.

Inhibitory effects of enzyme-treated dried sardine extract on IgE-mediated degranulation of RBL-2H3 cells and a murine model of Japanese cedar pollinosis

Protease-digested small dried sardine extract relieved allergic symptoms in mice through inhibiting microtubule dynamics.

Inhibition Activity of Plants on IgE-mediated Degranulation of RBL-2H3 Cells

In this study, the inhibitory activities of fifty plant extracts on IgE-mediated degranulation in the rat basophilic leukemia cell line (RBL-2H3 cells) were measured; the release of interleukin (IL)-4 and β-hexosaminidase from IgE-sensitized cells treated with the plant extracts was measured; and the effects of the plant extracts on cell viability were tested. The results of the analysis of plant extracts at 20 ㎍/㎖, including the aerial part of Magnolia sieboldii K. Koch, exhibited suppressive activities upon the release of IL-4. Furthermore, several plant extracts including methanol extracted from Lindera erythrocarpa Makino (aerial part) at the same concentration significantly inhibited the release of β-hexosaminidase. Twenty-six of the plant extracts, including methanol extract of Weigela subsessilis (Nakai) L. H. Bailey (branch), showed a cell proliferation effect of over 80% at 100 ㎍/㎖. In conclusion, the results suggest that the leaf/stem of Geum japonicum Thunb. and the stamen/ovary of Nelumbo nucifera Gaertn., which exhibited effective inhibition on β-hexosaminidase release and IL-4 release from mast cells and showed high cell viability, could be useful candidates as anti-allergy materials.

Establishment of a Mouse Anti-ovalbumin IgE Monoclonal Antibody That Induces FcɛRII (CD23)-dependent Activation Without FcɛRI-Dependent Activation.

IgE mainly activates cells via two receptors, FcɛRI and FcɛRII. Blocking antibodies against and animals genetically targeted for these receptors have been successfully used to distinguish between these two activating pathways. In the present study, we investigated whether our newly established anti-ovalbumin (OVA) monoclonal IgE OE-2 induced FcɛRII-dependent activation, but not FcɛRI-dependent activation in vivo and in vitro, in contrast to the previously established anti-OVA IgE OE-1, which stimulated FcɛRI and FcɛRII. The FcɛRI-mediated degranulation of RBL2H3 cells and passive systemic anaphylaxis in mice were induced by OE-1 but not OE-2. On the other hand, the production of nitric oxide by rat peritoneal macrophages and the primary antibody response in mice against co-injected OVA, which were mediated through FcɛRII, were induced and enhanced by OE-1 and OE-2. Differences in the epitopes recognized by OE-1 and OE-2 may partially explain why OE-1, but not OE-2, triggered FcɛRI-dependent activation. OE-1 bridged FcɛRI through effective aggregation with OVA, whereas OE-2 crosslinked the receptor strongly and only moderately upon the addition of an anti-kappa antibody and polymerized OVA, namely, an OVA-conjugated resin, respectively, resulting in degranulation. Our results offer a novel approach for determining the relative importance of FcɛRI and FcɛRII in various IgE-dependent responses by using OE-1 and OE-2.

The glycoprotein (18 kDa) isolated from Lactobacillus plantarum L67 suppressed ß-hexosaminidase, histamine, and the expression of TNF-α and IL-4 in the BPA-stimulated RBL-2H3 cells

This study investigated the inhibitory effect of the 18kDa glycoprotein isolated from Lactobacillus plantarum L67 on the bisphenol A-mediated inflammatory reaction in RBL-2H3 cells. The RBL-2H3 cells were treated with bisphenol A (50μmol) and the glycoprotein isolated from L. plantarum L67 (5–100μg/ml) for 30min. Subsequently, the release of histamine and β-hexosaminidase into the medium and the intracellular Ca2+ levels were measured. The activities of PKC and iNOS were measured by western blotting. In addition, the gene expression levels of IL-4 and TNF-α were measured by qRT-PCR. In conclusion, the results demonstrated that the degranulation of the BPA-stimulated RBL-2H3 cells and their histamine release were inhibited by the glycoprotein of L. plantarum L67; concurrently, the levels of intracellular Ca2+, the activity of iNOS, and the expression of TNF-α and IL-4, two allergy-related cytokines, were also suppressed by the glycoprotein. Taken together, these results indicate that the glycoprotein might help to prevent allergic diseases induced by environmental hormones, such as BPA.

Inhibitory Effects of Soyasaponins on Antigen-induced Degranulation in RBL-2H3 Cells

Abstract Soyasaponins are glycosylated, which gives rise to awide diversity of structures and functions. We evaluated forinhibitory effects of 4 soyasaponins on antigen-induced degranulationin RBL-2H3 Cells. 4 soyasaponins had shown dose-dependentlyinhibited histamine and β-hexosaminidase.Keywords allergic reaction · β-hexosaminidase · degranulation · RBL-2H3 Cells · soyasaponins 현대사회의 서구화된 식생활과 생활환경의 변화, 각종 환경 오염 증가 그리고 유전적 요인 등의 복합적인 원인에 의해 알레르기 질환이 해마다 증가하고 있다(Ebo 등, 2001; Alma 등,2008). IgE가 관여하여 일어나는 제 1형 과민반응은 즉시형 알레르기 반응으로 불리우며, 아토피 피부염, 천식, 알레르기 비염 등을 포함하는 가장 흔한 만성 면역계 질환이다. 비만세포는 대부분의 알레르기 질환에 중요한 역할을 하는 세포로 알려져 있다. 알레르기 반응은 IgE가 비만세포의 표면 수용체(high-affinity receptor for IgE, FceRI)와 결합되어 있어 알레르기 원인물질인 항원이 재차 유입되게 되면 비만세포 표면의 고친화성 IgE 수용체(FceRI)에 항원이 결합하게 된다. 이때 비만세포가 활성화가 되면 비만세포의 탈과립이 유도되어 과립내 존재하던 히스타민, 사이토카인 그리고 케모카인과 같은 알레르기유발 물질들이 세포 밖으로 분비되어 알레르기 반응을 일으키게 된다(Huang 등, 2008; Toshio, 2012).그러므로 탈과립 억제를 통한 비만세포 활성화 조절은 알레르기 반응의 예방과 치료에 중요한 자료로 활용될 수 있을 것이다.콩은 우리나라뿐만 아니라 전세계적으로 다양한 전통 및 가공식품으로 이용되고 있으며, 단백질, 지방 등의 함량이 높고isoflavone 및 soyasaponin 같은 생리활성물질들이 다량 함유되어 있다. 콩의 대표적 생리활성 물질로 isoflavone 이 많이 알려졌으나, 최근 콩에 함유된 soyasaponin의 혈중 콜레스테롤 감소효과, 항산화활성, 항바이러스, 간독성 물질제거, 항종양활성,면역증진 효과 등 여러 생리활성 기능이 밝혀지면서 신규 식의약 소재로 많은 주목을 받고 있다.콩 종실에 포함된 soyasaponin은 품종, 재배연도, 재배지역 및