Human Mast Cell Degranulation Research Articles

P2X7 Receptor-Induced Human Mast Cell Degranulation Is Enhanced by Interleukin 33.

MCs are tissue-resident immune cells that strategically reside in barrier organs and respond effectively to a wide range of stimuli, such as IL-33, a mediator released upon epithelial damage. Adenosine triphosphate (ATP) accumulates at sites of tissue injury and is known to modulate MC activities. This study investigated how an inflammatory tissue environment rich in IL-33 modulates the ATP-mediated activation of MCs. Human primary MCs primed with IL-33 displayed a strongly increased response to ATP but not ADP. This resulted in increased degranulation, IL-8 release, and pERK1/2 signalling. Such effects are unique to IL-33 stimulation and not shared by the epithelial alarmin, TSLP. MC exposure to IL-33 also increased membrane expression of purinergic and ATP-binding P2X receptors. The use of selective P2X receptor inhibitors identified P2X7 receptor as the key mediator of the enhanced ATP-induced ERK1/2 signalling and degranulation in IL-33-primed MCs. Whilst the inhibition of P2X1 and P2X4 receptors had no effect on MC degranulation, inhibiting these receptors together with P2X7 resulted in further decreased MC-mediated degranulation. These data therefore point toward the potential mechanisms by which IL-33 contributes to the modulation of ATP-mediated activation in human MCs.

An optimized method for IgE-mediated degranulation of human lung mast cells.

Mast cells are critically involved in IgE-mediated diseases, e.g., allergies and asthma. Human mast cells are heterogeneous, and mast cells from different anatomical sites have been shown to respond differently to certain stimuli and drugs. The origin of the mast cells is therefore of importance when setting up a model system, and human lung mast cells are highly relevant cells to study in the context of asthma. We therefore set out to optimize a protocol of IgE-mediated activation of human lung mast cells. Human lung mast cells were extracted from lung tissue obtained from patients undergoing pulmonary resection by enzyme digestion and mechanical disruption followed by CD117 magnetic-activated cell sorting (MACS) enrichment. Different culturing media and conditions for the IgE-mediated degranulation were tested to obtain an optimized method. IgE crosslinking of human lung mast cells cultured in serum-free media gave a stronger response compared to cells cultured with 10% serum. The addition of stem cell factor (SCF) did not enhance the degranulation. However, when the cells were put in fresh serum-free media 30 minutes prior to the addition of anti-IgE antibodies, the cells responded more vigorously. Maximum degranulation was reached 10 minutes after the addition of anti-IgE. Both CD63 and CD164 were identified as stable markers for the detection of degranulated mast cells over time, while the staining with anti-CD107a and avidin started to decline 10 minutes after activation. The levels of CD203c and CD13 did not change in activated cells and therefore cannot be used as degranulation markers of human lung mast cells. For an optimal degranulation response, human lung mast cells should be cultured and activated in serum-free media. With this method, a very strong and consistent degranulation response with a low donor-to-donor variation is obtained. Therefore, this model is useful for further investigations of IgE-mediated mast cell activation and exploring drugs that target human lung mast cells, for instance, in the context of asthma.

Cannabidiol Inhibits IgE-Mediated Mast Cell Degranulation and Anaphylaxis in Mice.

Cannabidiol (CBD), the most abundant non-psychoactive constituent of the plant Cannabis sativa, is known to possess immune modulatory properties. This study investigates the effects of CBD on mast cell degranulation in human and mouse primary mast cells and passive cutaneous anaphylaxis in mice. Mouse bone marrow-derived mast cells and human cord-blood derived mast cellsare generated. CBD suppressed antigen-stimulated mast cell degranulation in a concentration-dependent manner. Mechanistically, CBD inhibited both the phosphorylation of FcεRI downstream signaling molecules and calcium mobilization in mast cells, while exerting no effect on FcεRI expression and IgE binding to FcεRI. These suppressive effectsare preserved in the mast cells thatare depleted of type 1 (CB1) and type 2 (CB2) cannabinoid receptors, as well as in the presence of CB1 agonist, CB2 agonist, CB1 inverse agonist, and CB2 inverse agonist. CBD also inhibited the development of mast cells in a long-term culture. The intraperitoneal administration of CBD suppressed passive cutaneous anaphylaxis in mice as evidenced by a reduction in ear swelling and decrease in the number of degranulated mast cells. Based on these results, the administration of CBD is a new therapeutic intervention in mast cell-associated anaphylactic diseases.

68 EVO756 is a novel MRGPRX2 antagonist that potently inhibits human mast cell degranulation in response to multiple agonists – potential treatment for CSU and beyond

68 EVO756 is a novel MRGPRX2 antagonist that potently inhibits human mast cell degranulation in response to multiple agonists – potential treatment for CSU and beyond

Degranulation of human mast cells: modulation by P2 receptors' agonists.

Since the late 1970s, there has been an alarming increase in the incidence of asthma and its morbidity and mortality. Acute obstruction and inflammation of allergic asthmatic airways are frequently caused by inhalation of exogenous substances such as allergens cross-linking IgE receptors expressed on the surface of the human lung mast cells (HLMC). The degree of constriction of human airways produced by identical amounts of inhaled allergens may vary from day to day and even hour to hour. Endogenous factors in the human mast cell (HMC)'s microenvironment during allergen exposure may markedly modulate the degranulation response. An increase in allergic responsiveness may significantly enhance bronchoconstriction and breathlessness. This review focuses on the role that the ubiquitous endogenous purine nucleotide, extracellular adenosine 5'-triphosphate (ATP), which is a component of the damage-associated molecular patterns, plays in mast cells' physiology. ATP activates P2 purinergic cell-surface receptors (P2R) to trigger signaling cascades resulting in heightened inflammatory responses. ATP is the most potent enhancer of IgE-mediated HLMC degranulation described to date. Current knowledge of ATP as it relates to targeted receptor(s) on HMC along with most recent studies exploring HMC post-receptor activation pathways are discussed. In addition, the reviewed studies may explain why brief, minimal exposures to allergens (e.g., dust, cat, mouse, and grass) can unpredictably lead to intense clinical reactions. Furthermore, potential therapeutic approaches targeting ATP-related enhancement of allergic reactions are presented.

Widespread monoclonal IgE antibody convergence to an immunodominant, proanaphylactic Ara h 2 epitope in peanut allergy

Despite their central role in peanut allergy, human monoclonal IgE antibodies have eluded characterization. We sought to define the sequences, affinities, clonality, and functional properties of human monoclonal IgE antibodies in peanut allergy. We applied our single-cell RNA sequencing-based SEQ SIFTER discovery platform to samples from allergic individuals who varied by age, sex, ethnicity, and geographic location in order to understand commonalities in the human IgE response to peanut allergens. Select antibodies were then recombinantly expressed and characterized for their allergen and epitope specificity, affinity, and functional properties. We found striking convergent evolution of IgE monoclonal antibodies (mAbs) from several clonal families comprising both memory B cells and plasmablasts. These antibodies bound with subnanomolar affinity to the immunodominant peanut allergen Ara h 2, specifically a linear, repetitive motif. Further characterization of these mAbs revealed their ability to single-handedly cause affinity-dependent degranulation of human mast cells and systemic anaphylaxis on peanut allergen challenge in humanized mice. Finally, we demonstrated that these mAbs, reengineered as IgGs, inhibit significant, but variable, amounts of Ara h 2- and peanut-mediated degranulation of mast cells sensitized with allergic plasma. Convergent evolution of IgE mAbs in peanut allergy is a common phenomenon that can reveal immunodominant epitopes on major allergenic proteins. Understanding the functional properties of these molecules is key to developing therapeutics, such as competitive IgG inhibitors, that are able to stoichiometrically outcompete endogenous IgE for allergen and thereby prevent allergic cascade in cases of accidental allergen exposure.

A fully human anti-c-Kit monoclonal antibody 2G4 inhibits proliferation and degranulation of human mast cells

Given that mast cells are pivotal contributors to allergic diseases, various allergy treatments have been developed to inhibit them. Omalizumab, an anti-immunoglobulin E antibody, is a representative therapy that can alleviate allergy symptoms by inhibiting mast cell degranulation. However, omalizumab cannot reduce the proliferation and accumulation of mast cells, which is a fundamental cause of allergic diseases. c-Kit is essential for the proliferation, survival, and differentiation of mast cells. Excessive c-Kit activation triggers various mast cell diseases, such as asthma, chronic spontaneous urticaria, and mastocytosis. Herein, we generated 2G4, an anti-c-Kit antibody, to develop a therapeutic agent for mast cell diseases. The therapeutic efficacy of 2G4 antibody was evaluated in LAD2, a human mast cell line. 2G4 antibody completely inhibited c-Kit signaling by blocking the binding of stem cell factor, known as the c-Kit ligand. Inhibition of c-Kit signaling led to the suppression of proliferation, migration, and degranulation in LAD2 cells. Moreover, 2G4 antibody suppressed the secretion of pro-inflammatory cytokines, including granulocyte–macrophage colony-stimulating factor, vascular endothelial growth factor, C–C motif chemokine ligand 2, brain-derived neurotrophic factor, and complement component C5/C5a, which can exacerbate allergy symptoms. Taken together, these results suggest that 2G4 antibody has potential as a novel therapeutic agent for mast cell diseases.

Covalent heterobivalent inhibitor effectively inhibits anaphylaxis to peanut allergen in a humanized mouse model

Abstract Peanut-induced allergy is an IgE-mediated type I hypersensitivity reaction that manifests symptoms ranging from local edema to life-threatening anaphylaxis. While there are treatments for symptoms in allergic patients resulting from allergen exposure, there are few preventive therapies other than avoidance and no allergen specific therapies. We have previously designed a covalent heterobivalent inhibitor (cHBI) that binds in an allergen-specific manner as a preventive for allergic reactions. Building on previous in vitro testing, in this report we have developed a humanized mouse model to test cHBI efficacy in vivo. Humanized mice (NSG-SCF/GM-CSF/IL-3) developed mature functional human mast cells in various tissues and developed robust anaphylactic reactions when passively sensitized with human IgE monoclonal antibodies specific for peanut allergen. We found that the allergic response is IgE dose-dependent and is mediated by human mast cells indicated by elevated tryptase serum levels and the upregulation of mast cell degranulation markers. Using the validated humanized mouse model, we showed that cHBI inhibited IgE-mediated anaphylaxis and human mast cell degranulation. We demonstrated that cHBI effectively inhibits anaphylaxis for up to 14 days, and cHBI inhibition is specific to peanut allergen. Importantly, cHBI rescued the mice from lethal anaphylactic response during oral peanut-induced anaphylaxis. These findings suggest that cHBI has the potential to be an effective preventative for peanut food allergy in patients.

Fluoxetine-mediated suppression reveals a role for P2X receptors in mast cell function and allergic disease

Abstract There is a clinical need for new treatment options addressing allergic disease because current approaches are often suboptimal. Selective serotonin reuptake inhibitors (SSRIs) are a class of antidepressants that have been shown to possess anti-inflammatory properties, but the mechanism of action is unclear. Because mast cells are critical effectors of allergic inflammation, we tested SSRI effects on IgE-induced mast cell function. We show that the SSRI fluoxetine suppresses IgE-mediated degranulation and cytokine production in mouse and human mast cells. Fluoxetine was also effective in a model of allergic airway inflammation, where it reduced bronchoresponsiveness and inflammation. IgE crosslinkage elicited rapid ATP release from mast cells, suggesting ATP-P2X receptor signaling may potentiate IgE-driven responses. We subsequently found that fluoxetine inhibits ATP-induced mast cell function. We propose that fluoxetine acts by blocking an ATP-P2X autocrine or paracrine loop that augments mast cell-mediated inflammation. These data support the potential repurposing of fluoxetine in allergic disease. Supported by NIH/NIAID R01138495 R01164710

The first reptilian allergen and major allergen for fish-allergic patients: Crocodile β-parvalbumin.

BackgroundClinical cross‐reactivity between bony fish, cartilaginous fish, frog, and chicken muscle has previously been demonstrated in fish‐allergic patients. In indicative studies, two reports of anaphylaxis following the consumption of crocodile meat and IgE‐cross‐binding were linked to the major fish allergen parvalbumin (PV). This study investigates IgE‐binding proteins in crocodile meat with a focus on PV and their clinical relevance.MethodsProteins were extracted from muscle tissue of crocodile, three bony fish, and two cartilaginous fish. A cohort of fish‐allergic pediatric patients (n = 77) underwent allergen skin prick testing (SPT) to three fish preparations (n = 77) and crocodile (n = 12). IgE‐binding proteins were identified and quantified by SDS‐PAGE, mass spectrometric analyses, and immunoblotting using commercial and in‐house antibodies, as well as individual and pooled patients’ serum. PV isoforms were purified or recombinantly expressed before immunological analyses, including human mast cell degranulation assay.ResultsOf the tissues analyzed, PV was most abundant in heated crocodile preparation, triggering an SPT of ≥3 mm in 8 of 12 (67%) fish‐allergic patients. Seventy percent (31 of 44) of fish PV‐sensitized patients demonstrated IgE‐binding to crocodile PV. Crocodile β‐PV was the major IgE‐binding protein but 20‐fold less abundant than α‐PV. Cellular reactivity was demonstrated for β‐PV and epitopes predicted, explaining frequent IgE‐cross‐binding of β‐PVs. Both PV isoforms are now registered as the first reptile allergens with the WHO/IUIS (β‐PV as Cro p 1 and α‐PV as Cro p 2).ConclusionFish‐allergic individuals may be at risk of an allergy to crocodile and should seek specialist advice before consuming crocodilian meat.

MRGPRX2-Mediated Degranulation of Human Skin Mast Cells Requires the Operation of Gαi, Gαq, Ca++ Channels, ERK1/2 and PI3K-Interconnection between Early and Late Signaling.

The recent discovery of MRGPRX2 explains mast cell (MC)-dependent symptoms independently of FcεRI-activation. Because of its novelty, signaling cascades triggered by MRGPRX2 are rudimentarily understood, especially in cutaneous MCs, by which MRGPRX2 is chiefly expressed. Here, MCs purified from human skin were used following preculture or ex vivo and stimulated by FcεRI-aggregation or MRGPRX2 agonists (compound 48/80, Substance P) in the presence/absence of inhibitors. Degranulation was assessed by β-hexosaminidase or histamine release. Phosphorylation events were studied by immunoblotting. As a G protein-coupled receptor, MRGPRX2 signals by activating G proteins; however, their nature has remained controversial. In skin MCs, Gαi and Gαq were required for degranulation, but Gαi was clearly more relevant. Ca++ channels were likewise crucial. Downstream, PI3K was essential for granule discharge initiated by MRGPRX2 or FcεRI. ERK1/2 and JNK were additional participants, especially in the allergic route. Addressing possible points of intersection between early and later events, pERK1/2 and pAKT were found to depend on Gαi, further highlighting its significance. Gαq and Ca++ channels made some contributions to the phosphorylation of ERK. Ca++ differentially affected PI3K activation in FcεRI- vis-à-vis MRGPRX2-signaling, as channel inhibition increased pAKT only when triggered via FcεRI. Collectively, our study significantly extends our understanding of the molecular framework behind granule secretion from skin MCs.

Micronutritional supplementation with a holoBLG-based FSMP (food for special medical purposes)-lozenge alleviates allergic symptoms in BALB/c mice: Imitating the protective farm effect.

Previously, the protective farm effect was imitated using the whey protein beta-lactoglobulin (BLG) that is spiked with iron-flavonoid complexes. Here, we formulated for clinical translation a lozenge as food for special medical purposes (FSMP) using catechin-iron complexes as ligands for BLG. The lozenge was tested in vitro and in a therapeutical BALB/c mice model. Binding of iron-catechin into BLG was confirmed by spectroscopy and docking calculations. Serum IgE binding of children allergic or tolerating milk was assessed to loaded (holo-) versus empty (apo-) BLG and for human mast cell degranulation. BLG and Betv1 double-sensitized mice were orally treated with the holoBLG or placebo lozenge, and immunologically analysed after systemic allergen challenge. Human PBMCs of pollen allergic subjects were flow cytometrically assessed after stimulation with apoBLG or holoBLG using catechin-iron complexes as ligands. One major IgE and T cell epitope were masked by catechin-iron complexes, which impaired IgE binding of milk-allergic children and degranulation of mast cells. In mice, only supplementation with the holoBLG lozenge reduced clinical reactivity to BLG and Betv1, promoted Tregs, and suppressed antigen presentation. In allergic subjects, stimulation of PBMCs with holoBLG led to a significant increase of intracellular iron in circulating CD14+ cells with significantly lower expression of HLADR and CD86 compared to their stimulation with apoBLG. The FSMP lozenge targeted antigen presenting cells and dampened immune activation in human immune cells and allergic mice in an antigen-non-specific manner, thereby conferring immune resilience against allergic symptoms.

Novel Insights on MRGPRX2-Mediated Hypersensitivity to Neuromuscular Blocking Agents And Fluoroquinolones.

Neuromuscular blocking agents (NMBAs) like atracurium and rocuronium as well as fluoroquinolones (FQs) cause mast cell-mediated anaphylaxis by activating Mas-related G protein-coupled receptor X2 (MRGPRX2), but many questions remain unanswered. Here, we address three of them, namely whether primary human mast cells show similar activation by these drugs as murine mast cells and mast cell lines, how sugammadex protects from atracurium-induced MRGPRX2-mediated mast cell activation, and why some but not all patients treated with rocuronium develop anaphylaxis. We used peripheral blood-derived cultured mast cells from healthy donors and patients, assessed mast cell activation and degranulation by quantifying intracellular calcium and CD63 expression, respectively, and made use of MRGPRX2-silencing, via electroporation with Dicer-substrate small interfering RNAs, and single cell flow cytometric analyses. Atracurium, ciprofloxacin, and levofloxacin activated and degranulated primary human mast cells, but only MRGPRX2-positive and not MRGPRX2-negative or -silenced mast cells. Sugammadex attenuated the atracurium-induced and MRGPRX2-mediated activation and degranulation of human mast cells by reducing free atracurium levels. The mast cells of patients with IgE-independent anaphylaxis to rocuronium were similar, in their MRGPRX2 expression and function, to those of patients with IgE-mediated anaphylaxis. These findings further improve our understanding of the role and relevance of MRGPRX2-driven mast cell activation in anaphylactic reactions to NMBAs and FQs and may help to improve their prediction, prevention, and treatment.

MYO1F Regulates IgE and MRGPRX2-Dependent Mast Cell Exocytosis.

The activation and degranulation of mast cells is critical in the pathogenesis of allergic inflammation and modulation of inflammation. Recently, we demonstrated that the unconventional long-tailed myosin, MYO1F, localizes with cortical F-actin and mediates adhesion and migration of mast cells. In this study, we show that knockdown of MYO1F by short hairpin RNA reduces human mast cell degranulation induced by both IgE crosslinking and by stimulation of the Mas-related G protein-coupled receptor X2 (MRGPRX2), which has been associated with allergic and pseudoallergic drug reactions, respectively. Defective degranulation was accompanied by a reduced reassembly of the cortical actin ring after activation but reversed by inhibition of actin polymerization. Our data show that MYO1F is required for full Cdc42 GTPase activation, a critical step in exocytosis. Furthermore, MYO1F knockdown resulted in less granule localization in the cell membrane and fewer fissioned mitochondria along with deficient mitochondria translocation to exocytic sites. Consistent with that, AKT and DRP1 phosphorylation are diminished in MYO1F knockdown cells. Altogether, our data point to MYO1F as an important regulator of mast cell degranulation by contributing to the dynamics of the cortical actin ring and the distribution of both the secretory granules and mitochondria.

The Use of Bruton's Tyrosine Kinase Inhibitors to Treat Allergic Disorders.

Purpose of reviewStudies show that inhibitors of Bruton’s tyrosine kinase (BTKis), currently FDA-approved for the treatment of B cell malignancies, can prevent IgE-mediated reactions through broad inhibition of the FcεRI signaling pathway in human mast cells and basophils. This review will summarize recent data supporting the use of these drugs as novel therapies in various allergic disorders.Recent findingsRecent studies have shown that BTKis can prevent IgE-mediated degranulation and cytokine production in primary human mast cells and basophils. Two oral doses of the second-generation BTKi acalabrutinib can completely prevent moderate passive systemic anaphylaxis in humanized mice and even protect against death during severe anaphylaxis. Furthermore, two doses of ibrutinib can reduce or eliminate skin prick test responses to foods and aeroallergens in allergic subjects. BTKis in development also show efficacy in clinical trials for chronic urticaria. Unlike other therapies targeting IgE, such as omalizumab, BTKis appear to have rapid onset and transient effects, making them ideal candidates for intermittent use to prevent acute reactions such as IgE-mediated anaphylaxis.SummaryThese studies suggest that BTKis may be capable of preventing IgE-mediated anaphylaxis, paving the way for future trials in food allergy and urticaria.

Revisiting the role of MRGPRX2 on hypersensitivity reactions to neuromuscular blocking drugs.

Anaphylaxis is caused by a variety of triggers including Food and Drug Administration (FDA)-approved antibiotics, contrast media and neuromuscular blocking drugs (NMBDs). Traditionally, drug-induced anaphylaxis was thought to result mainly from IgE-mediated histamine release from mast cells. Recently, a G protein-coupled receptor known as MRGPRX2 has been identified and shown to be highly expressed on human skin but not lung mast cells. The demonstration that many NMBDs induce degranulation in human mast cells via MRGPRX2 led to the idea that this receptor contributes to NMBD-induced hypersensitivity reactions. However, other studies have raised doubts regarding its role in drug-induced hypersensitivity. This review discusses the current status and controversy on MRGPRX2's role on NMBD-induced hypersensitivity.

A group of cationic amphiphilic drugs activates MRGPRX2 and induces scratching behavior in mice

Mas gene-related G protein-coupled receptors (MRGPRs) are a G protein-coupled receptor family responsive to various exogenous and endogenous agonists, playing a fundamental role in pain and itch sensation. The primate-specific family member MRGPRX2 and its murine orthologue MRGPRB2 are expressed by mast cells mediating IgE-independent signaling and pseudoallergic drug reactions. Our aim was to increase knowledge about the function and regulation of MRGPRX2/MRGPRB2, which is of major importance in prevention of drug hypersensitivity reactions and drug-induced pruritus. To identify novel MRGPR (ant)agonists, we screened a library of pharmacologically active compounds by utilizing a high-throughput calcium mobilization assay. The identified hit compounds were analyzed for their pseudoallergic and pruritogenic effects in mice and human. We found a class of commonly used drugs activating MRGPRX2 that, to a large extent, consists of antidepressants, antiallergic drugs, and antipsychotics. Three-dimensional pharmacophore modeling revealed structural similarities of the identified agonists, classifying them as cationic amphiphilic drugs. Mast cell activation was investigated by using the 3 representatively selected antidepressants clomipramine, paroxetine, and desipramine. Indeed, we were able to show a concentration-dependent activation and MRGPRX2-dependent degranulation of the human mast cell line LAD2 (Laboratory of Allergic Diseases-2). Furthermore, clomipramine, paroxetine, and desipramine were able to induce degranulation of human skin and murine peritoneal mast cells. These substances elicited dose-dependent scratching behavior following intradermal injection into C57BL/6 mice but less so in MRGPRB2-mutant mice, as well as wheal-and-flare reactions following intradermal injections in humans. Our results contribute to the characterization of structure-activity relationships and functionality of MRGPRX2 ligands and facilitate prediction of adverse reactions such as drug-induced pruritus to prevent severe drug hypersensitivity reactions.

Mast Cell-Specific MRGPRX2: a Key Modulator of Neuro-Immune Interaction in Allergic Diseases.

Atopic dermatitis (AD) and allergic asthma are complex disorders with significant public health burden. This review provides an overview of the recent developments on Mas-related G protein-coupled receptor-X2 (MRGPRX2; mouse counterpart MrgprB2) as a potential candidate to target neuro-immune interaction in AD and allergic asthma. Domestic allergens directly activate sensory neurons to release substance P (SP), which induces mast cell degranulation via MrgprB2 and drives type 2 skin inflammation in AD. MRGPRX2 expression is upregulated in human lung mast cells and serum of asthmatic patients. Both SP and hemokinin-1 (HK-1 generated from macrophages, bronchial cells, and mast cells) cause degranulation of human mast cells via MRGPRX2. MrgprB2 contributes to mast cell-nerve interaction in the pathogenesis of AD. Furthermore, asthma severity is associated with increased MRGPRX2 expression in mast cells. Thus, MRGPRX2 could serve as a novel target for modulating AD and asthma.

Trichomonas vaginalis-secreted cysteinyl leukotrienes promote migration, degranulation and MCP-1 production in mast cells.

Trichomonas vaginalis, a flagellated extracellular protozoan parasite that infects the human genitourinary tract, is usually transmitted by sexual contact. Our previous study showed that the leukotriene B4 (LTB4 ), a Tvaginalis-secreted lipid mediator, induces interleukin (IL)-8 production and promotes mast cell degranulation and migration via BLT1 in human. In this study, we investigated whether Tvaginalis produces another leukotrienes and whether it causes increased MCP-1 production, mast cell migration and degranulation by activating mast cells. We found that cysteinyl leukotrienes (CysLTs) were contained in Tvaginalis-derived secretory product (TvSP) by ELISA. The TvSP-stimulated human mast cell line (HMC-1) exhibited significantly increased monocyte chemoattractant protein-1 (MCP-1) secretion compared to the unstimulated cells. Inhibition of NOX2 activation of cells by treatment of NOX inhibitor or NOX2 siRNA reduced TvSP-stimulated MCP-1 production in HMC-1 cells. It was also confirmed that the receptor for CysLTs is expressed in mast cells. The CysLT receptor (CysLTR) antagonist inhibited TvSP-stimulated MCP-1 production of mast cells, as well as ROS production, migration and degranulation of mast cells, and reduced phospho-NF-kB expression. These results suggest that Tvaginalis-secreted CysLTs promote migration, degranulation and MCP-1 production in human mast cells through CysLT receptor-mediated NOX2 activation.

Coagulation factors induce human skin mast cell and basophil degranulation via activation of complement 5 and the C5a receptor

Coagulation factors induce human skin mast cell and basophil degranulation via activation of complement 5 and the C5a receptor