Mast Cell-mediated Inflammation Research Articles

Emedastine Inhibits Th1 and Th2 Cell Differentiation Mediated by Mast Cells.

We have previously clarified that emedastine, a second-generation antihistamine drug, inhibits T helper 1(Th1)/Th2 cell differentiation mediated by Langerhans cells (LCs). In addition, although we have recently found that mast cells also function as antigen-presenting cells (APCs) and induce Th1/Th2 cell differentiation, any influence of emedastine on this function remained unclear. Here we investigated the influence of emedastine on Th1/Th2 cell differentiation via mast cells. Mast cells were obtained by long-term culture of murine splenocytes in medium supplemented with tumor necrosis factor (TNF)-α. The mast cells were then incubated in the presence or absence of emedastine, and cultured with naïve CD4+ T cells in the presence of ovalbumin (OVA) peptide. Five days later, Th cells in the culture were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, and Th1/Th2 cytokine production was examined by enzyme-linked immunosorbent assay. When mast cells treated with emedastine were used as APCs, production of interferon (IFN)-γ and interleukin (IL)-4 from activated Th cells was significantly suppressed. This suppression was associated with inhibition of CD86 expression on mast cells, and mast cells treated with emedastine were shown to obstruct the differentiation of both Th1 and Th2 cells by down-regulating their cell surface expression of CD86. The present data provide additional information that topical application of emedastine to the lesional skin of patients with atopic dermatitis (AD) would reduce not only LC- but also mast cell-mediated skin inflammation.

Role of mechanoregulation in mast cell-mediated immune inflammation of the smooth muscle in the pathophysiology of esophageal motility disorders.

Major esophageal disorders involve obstructive transport of bolus to the stomach, causing symptoms of dysphagia and impaired clearing of the refluxed gastric contents. These may occur due to mechanical constriction of the esophageal lumen or loss of relaxation associated with deglutitive inhibition, as in achalasia-like disorders. Recently, immune inflammation has been identified as an important cause of esophageal strictures and the loss of inhibitory neurotransmission. These disorders are also associated with smooth muscle hypertrophy and hypercontractility, whose cause is unknown. This review investigated immune inflammation in the causation of smooth muscle changes in obstructive esophageal bolus transport. Findings suggest that smooth muscle hypertrophy occurs above the obstruction and is due to mechanical stress on the smooth muscles. The mechanostressed smooth muscles release cytokines and other molecules that may recruit and microlocalize mast cells to smooth muscle bundles, so that their products may have a close bidirectional effect on each other. Acting in a paracrine fashion, the inflammatory cytokines induce genetic and epigenetic changes in the smooth muscles, leading to smooth muscle hypercontractility, hypertrophy, and impaired relaxation. These changes may worsen difficulty in the esophageal transport. Immune processes differ in the first phase of obstructive bolus transport, and the second phase of muscle hypertrophy and hypercontractility. Moreover, changes in the type of mechanical stress may change immune response and effect on smooth muscles. Understanding immune signaling in causes of obstructive bolus transport, type of mechanical stress, and associated smooth muscle changes may help pathophysiology-based prevention and targeted treatment of esophageal motility disorders.NEW & NOTEWORTHY Esophageal disorders such as esophageal stricture or achalasia, and diffuse esophageal spasm are associated with smooth muscle hypertrophy and hypercontractility, above the obstruction, yet the cause of such changes is unknown. This review suggests that smooth muscle obstructive disorders may cause mechanical stress on smooth muscle, which then secretes chemicals that recruit, microlocalize, and activate mast cells to initiate immune inflammation, producing functional and structural changes in smooth muscles. Understanding the immune signaling in these changes may help pathophysiology-based prevention and targeted treatment of esophageal motility disorders.

P2X3 Potentiates IgE Mediated Mast Cell Function

Abstract The rising incidence of allergic disease is a public health challenge needing novel interventions. Mast cell activation by IgE is an established feature of allergy, but fundamental aspects of IgE-induced signaling remain unclear. Prior data from our lab testing the effects of fluoxetine on IgE-driven mast cell function indicated a role for the purinergic receptor P2X3. The P2X3 receptor is an ATP-activated cationic channel most often associated with pain that is expressed on the surface of mast cells. IgE cross-linkage elicits a rapid release of ATP from mast cells, suggesting ATP-P2X3 receptor signaling may potentiate IgE-driven responses. We show that diminishing P2X3 function pharmacologically and genetically reduces IgE-mediated cytokine production. P2X3 knockout bone marrow-derived mast cells showed a diminished capability to produce cytokine in vitro when stimulated by IgE cross-linkage. The P2X3 inhibitors BLU-5937 and Gefapixant administered intraperitoneally suppressed cytokine levels measured in the plasma of mice subjected to the mast cell-dependent passive systemic anaphylaxis model (PSA). Furthermore, P2X3 knock-out mice showed reduced IL-6 compared to wild-type mice in the PSA model. Interestingly, inhibition or deletion of the P2X3 receptor showed no effect on mast cell degranulation in vitro and did not reduce hypothermia during PSA. We propose that an ATP-P2X3 autocrine or paracrine loop augments mast cell-mediated inflammation, thus offering a potential new target for the treatment of allergic disease. RO1 AI164710-01, TL1 DK 132771-1

Mast cell–mediated inflammation relies on insulin-regulated aminopeptidase controlling cytokine export from the Golgi

On activation, mast cells rapidly release preformed inflammatory mediators from large cytoplasmic granules via regulated exocytosis. This acute degranulation is followed by a late activation phase involving synthesis and secretion of cytokines, growth factors, and other inflammatory molecules via the constitutive pathway that remains ill defined. We investigated the role for an insulin-responsive vesicle-like endosomal compartment, marked by insulin-regulated aminopeptidase (IRAP), in the secretion of TNF-α and IL-6 in mast cells and macrophages. Murine knockout (KO) mouse models (IRAP-KO and kit-Wsh/sh) were used to study inflammatory disease models and to measure and mechanistically investigate cytokine secretion and degranulation in bone marrow-derived mast cells invitro. IRAP-KO mice are protected from TNF-α-dependent kidney injury and inflammatory arthritis. In the absence of IRAP, TNF-α and IL-6 but not IL-10 fail to be efficiently secreted. Moreover, chemical targeting of IRAP endosomes reduced proinflammatory cytokine secretion. Mechanistically, impaired TNF-α export from the Golgi and reduced colocalization of vesicle-associated membrane protein (VAMP) 3-positive TNF-α transport vesicles with syntaxin 4 (aka Stx4) was observed in IRAP-KO mast cells, while VAMP8-dependent exocytosis of secretory granules was facilitated. IRAP plays a novel role in mast cell-mediated inflammation through the regulation of exocytic trafficking of cytokines.

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

Human Precision-Cut Lung Slices: Generation of and Measurement of Contractility and Relaxation of Small Airways.

Lung slices have been used since the mid-1990's to study various aspects of lung biology that include, but are not limited to, mechanisms of airway contraction and relaxation; the pulmonary immune response in the context of inflammatory diseases of the lung like asthma and chronic obstructive pulmonary disease; mast cell-mediated airway contractility and inflammation; modulation of airway cells following pathogen exposure; and consequences of environmental toxicant exposure. Here we describe the generation of human precision-cut lung slices (hPCLS) and measurement of contraction and relaxation of small airways within the slices.

Lactic acid suppresses MRGPRX2 mediated mast cell responses

MAS related G-protein coupled receptor X2 (MRGPRX2) is a G-protein coupled receptor (GPCR) expressed in human mast cells that has been implicated to play an important role in causing pseudo-allergic reactions as well as exacerbating inflammation during asthma and other allergic diseases. Lactic acid, a byproduct of glucose metabolism, is abundantly present in inflamed tissues and has been shown to regulate functions of several immune cells. Because the endogenous ligands for MRGPRX2 (substance P and LL-37) are elevated during pathologic conditions, such as cancer and asthma, and given that lactic acid levels are also enhanced in these patients, we explored the role of lactic acid in regulating mast cells response via MRGPRX2 and MrgprB2, the mouse orthologue of the human receptor. We found that lactic acid suppressed both the early (Ca2+ mobilization and degranulation) and late (chemokine/cytokine release) phases of mast cell activation; this data was confirmed in LAD2, human skin and mouse peritoneal mast cells. In LAD2 cells, the reduction in degranulation and chemokine/cytokine production mediated by lactic acid was dependent on pH. In agreement with our in vitro studies, lactic acid also reduced passive systemic anaphylaxis to compound 48/80 (a known MRGPRX2/MrgprB2 ligand) and skin inflammation in a mouse model of rosacea that is dependent on MrgprB2 expression on skin mast cells. Our data thus suggest that lactic acid may serve to inhibit mast cell-mediated inflammation during asthma and reduce immune response during cancer by affecting mast cell activation through MRGPRX2.

Mast cell tryptases in allergic inflammation and immediate hypersensitivity.

Dysregulated mast cell-mediated inflammation and/or activation have been linked to a number of human diseases, including asthma, anaphylaxis, chronic spontaneous urticaria, and mast cell activation syndromes. As a major mast cell granule protein, tryptase is a biomarker commonly used in clinical practice to diagnose mast cell-associated disorders and -mediated reactions, but its mechanistic roles in disease pathogenesis remains incompletely understood. Here, we summarize recent advances in the understanding of human tryptase genetics and the effects that different genetic composition may have on the quaternary structure of tetrameric mature tryptases. We also discuss how these differences may impact clinical phenotypes including allergic inflammation, immediate hypersensitivity, and others seen in patients with mast cell-associated disorders. With the increased application of next-generation sequencing, we foresee that human genetic approaches will be a major focus of understanding human tryptase functions in various human mast cell disorders and in new therapeutic development.

Therapeutic hypothermia reduces inflammation and oxidative stress in the liver after asphyxial cardiac arrest in rats.

Background Few studies have evaluated the effects of hypothermia on cardiac arrest (CA)-induced liver damage. This study aimed to investigate the effects of hypothermic therapy on the liver in a rat model of asphyxial cardiac arrest (ACA).Methods Rats were subjected to 5-minute ACA followed by return of spontaneous circulation (RoSC). Body temperature was controlled at 33°C±0.5°C or 37°C±0.5°C for 4 hours after RoSC in the hypothermia group and normothermia group, respectively. Liver tissues in each group were collected at 6 hours, 12 hours, 1 day, and 2 days after RoSC. To examine hepatic inflammation, mast cells were stained with toluidine blue. Superoxide anion radical production was evaluated using dihydroethidium fluorescence straining and expression of endogenous antioxidants (superoxide dismutase 1 [SOD1] and SOD2) was examined using immunohistochemistry.Results There were significantly more mast cells in the livers of the normothermia group with ACA than in the hypothermia group with ACA. Gradual increase in superoxide anion radical production was found with time in the normothermia group with ACA, but production was significantly suppressed in the hypothermia group with ACA relative to the normothermia group with ACA. SOD1 and SOD2 levels were higher in the hypothermia group with ACA than in the normothermia group with ACA.Conclusions Experimental hypothermic treatment after ACA significantly inhibited inflammation and superoxide anion radical production in the rat liver, indicating that this treatment enhanced or maintained expression of antioxidants. Our findings suggest that hypothermic therapy after CA can reduce mast cell-mediated inflammation through regulation of oxidative stress and the expression of antioxidants in the liver.

Extract of Boehmeria nivea Suppresses Mast Cell-Mediated Allergic Inflammation by Inhibiting Mitogen-Activated Protein Kinase and Nuclear Factor-κB.

Mast cells are effector cells that initiate allergic inflammatory immune responses by inducing inflammatory mediators. Boehmeria nivea (Linn.) Gaudich is a natural herb in the nettle family Urticaceae that possesses numerous pharmacological properties. Despite the various pharmacological benefits of Boehmeria nivea, its effects on allergic inflammation have not yet been determined. Here, we investigated the effect of the ethanol extract of Boehmeria nivea (BNE) on degranulation rat basophilic leukemia (RBL)-2H3 mast cells stimulated with anti-dinitrophenyl (anti-DNP) and bovine serum albumin (BSA) during immunoglobulin E (IgE)-mediated allergic immune response. The results showed inhibition of the release of β-hexosaminidase and histamine from the cells. BNE suppressed pro-inflammatory cytokines (Tumor necrosis factor (TNF)-α, Interleukin (IL)-1β, and IL-6) and reduced T helper (Th)2 cytokine IL-4 expression and/or secretion correlated with the downregulation of p38, extracellular signal-regulated kinases (ERK) mitogen-activated protein kinase (MAPK), and nuclear factor-κB (NF-κB) signaling pathways in treated RBL-2H3 mast cells. In passive cutaneous anaphylaxis, treatment with BNE during IgE-mediated local allergic reaction triggered a reduction in mouse ear pigmentation and thickness. Taken together, these results indicated that BNE suppressed mast cell-mediated inflammation, suggesting that BNE might be a candidate for the treatment of various allergic disorders.

Lactic acid suppresses IgE-mediated mast cell function in vitro and in vivo

Mast cells have functional plasticity affected by their tissue microenvironment, which greatly impacts their inflammatory responses. Because lactic acid (LA) is abundant in inflamed tissues and tumors, we investigated how it affects mast cell function. Using IgE-mediated activation as a model system, we found that LA suppressed inflammatory cytokine production and degranulation in mouse peritoneal mast cells, data that were confirmed with human skin mast cells. In mouse peritoneal mast cells, LA-mediated cytokine suppression was dependent on pH- and monocarboxylic transporter-1 expression. Additionally, LA reduced IgE-induced Syk, Btk, and ERK phosphorylation, key signals eliciting inflammation. In vivo, LA injection reduced IgE-mediated hypothermia in mice undergoing passive systemic anaphylaxis. Our data suggest that LA may serve as a feedback inhibitor that limits mast cell-mediated inflammation.

The Role of Histamine and Histamine Receptors in Mast Cell-Mediated Allergy and Inflammation: The Hunt for New Therapeutic Targets.

Histamine and its receptors (H1R–H4R) play a crucial and significant role in the development of various allergic diseases. Mast cells are multifunctional bone marrow-derived tissue-dwelling cells that are the major producer of histamine in the body. H1R are expressed in many cells, including mast cells, and are involved in Type 1 hypersensitivity reactions. H2R are involved in Th1 lymphocyte cytokine production. H3R are mainly involved in blood–brain barrier function. H4R are highly expressed on mast cells where their stimulation exacerbates histamine and cytokine generation. Both H1R and H4R have important roles in the progression and modulation of histamine-mediated allergic diseases. Antihistamines that target H1R alone are not entirely effective in the treatment of acute pruritus, atopic dermatitis, allergic asthma, and other allergic diseases. However, antagonists that target H4R have shown promising effects in preclinical and clinical studies in the treatment of several allergic diseases. In the present review, we examine the accumulating evidence suggesting novel therapeutic approaches that explore both H1R and H4R as therapeutic targets for histamine-mediated allergic diseases.

IL-33 mast cell axis is central in LL-37 induced bladder inflammation and pain in a murine interstitial cystitis model

Interstitial cystitis (IC), also known as painful bladder syndrome (PBS), is a debilitating chronic condition that afflicts over 3 million women above the age of 18 in the U.S., and most patients fail to respond to current treatment options. Mast cells have previously been implicated as both a diagnostic and prognostic marker in IC/PBS. Patients with IC/PBS have been shown to have elevated levels of IL-33, a cytokine released in response to tissue insult, in their urine. We hypothesize that mast cell-mediated inflammation induced from IL-33 may play an important role in initiating pain and inflammation in IC/PBS. A human cathelicidin, LL-37, which is found at elevated levels in IC/PBS patients, was used to induce an IC/PBS-like state of inflammation and bladder pain in mast cell deficient C-kit (−/−) and wild type C57Bl/6 (WT) mice. Inflammation was quantified using myeloperoxidase (MPO) expression in bladder tissues measured via ELISA. Response rate to suprapubic stimulation from von Frey filaments was used to assess the relative pain and discomfort. Both types of mice increased IL-33 expression in response to LL-37 exposure. However, mast cell deficient mice demonstrated significantly lower levels of inflammation (p < 0.001) and reduced pain response (p < 0.001) compared to WT mice. These findings implicate an IL-33-mast cell dependent axis with a potential etiology of pain and inflammation in IC/PBS. Future therapeutics aimed at targeting the IL-33 - mast cell axis could potentially serve as useful targets for treating IC/PBS.

Functional role of kynurenine and aryl hydrocarbon receptor axis in chronic rhinosinusitis with nasal polyps

Chronic rhinosinusitis with nasal polyps (CRSwNP) is associated with mast cell-mediated inflammation and heightened oxidant stress. Kynurenine (KYN), an endogenous tryptophan metabolite, can promote allergen-induced mast cell activation through the aryl hydrocarbon receptor (AhR). We sought to determine the role of the KYN/AhR axis and oxidant stress in mast cell activation and the development of CRSwNP. We measured the expression of indoleamine 2,3-dioxygenase 1, tryptophan 2,3-dioxygenase, KYN, and oxidized calmodulin-dependent protein kinase II (ox-CaMKII) in nasal polyps and controls. KYN-potentiated ovalbumin (OVA)-induced ROS generation, cell activation, and ox-CaMKII expression were investigated in wild-type and AhR-deficient (AhR-/-) mast cells. The role of ox-CaMKII in mast cell activation was further investigated. Nasal polyps in CRSwNP showed an increased expression of indoleamine 2,3-dioxygenase 1, tryptophan2,3-dioxygenase, and KYN compared with controls. AhR was predominantly expressed in mast cells in nasal polyps. Activated mast cells and local IgE levels were substantially increased in eosinophilic polyps compared with noneosinophilic polyps and controls. Furthermore, KYN potentiated OVA-induced ROS generation, intracellular Ca2+ levels, cell activation, and expression of ox-CaMKII in wild-type, but not in AhR-/- mast cells. Compared with noneosinophilic polyps and controls, eosinophilic polyps showed increased expression of ox-CaMKII in mast cells. Mast cells from ROS-resistant CaMKII MMVVδ mice or pretreated with CaMKII inhibitor showed protection against KYN-promoted OVA-induced mast cell activation. These studies support a potentially critical but previously unidentified function of the KYN/AhR axis in regulating IgE-mediated mast cell activation through ROS and ox-CaMKII in CRSwNP.

Carbonic anhydrase enzymes regulate mast cell-mediated inflammation

Abstract Type 2 cytokine responses are necessary for the development of protective immunity to helminth parasites but also cause the inflammation associated with allergies and asthma. Recent studies have found that peripheral hematopoietic progenitor cells contribute to type 2 cytokine-mediated inflammation through their enhanced ability to develop into mast cells. Here we show that carbonic anhydrase (Car) enzymes are upregulated in type 2-associated progenitor cells and demonstrate that Car enzyme inhibition is sufficient to prevent murine mast cell responses and inflammation following Trichinella spiralis infection or the induction of food allergy-like disease. Further, we employed CRISPR/Cas9 technology and illustrate that genetically editing Car1 is sufficient to selectively reduce mast cell development. Finally, we demonstrate that Car enzymes can be targeted to prevent human mast cell development. Collectively these studies identify a previously unrecognized role for Car enzymes in regulating mast cell lineage commitment and suggest that Car enzyme inhibitors may possess therapeutic potential that can be employed to treat mast cell-mediated inflammation.

Abnormal Sensory Protein Expression and Urothelial Dysfunction in Ketamine-Related Cystitis in Humans.

PurposeThe aim of this study was to analyze patterns of sensory protein expression and urothelial dysfunction in ketamine-related cystitis (KC) in humans.MethodsBiopsies of bladder mucosa were performed in 29 KC patients during cystoscopy. Then specimens were analyzed for tryptase, zonula occludens-1 (ZO-1), E-cadherin, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) with immunofluorescence staining and quantification. In addition, 10 healthy control bladder specimens were analyzed and compared with the KC specimens. Another 16 whole bladder specimens obtained from partial cystectomy were also analyzed for the muscarinic receptors M2 and M3, endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), β-3 adrenergic receptors (β3-ARs), and the P2X3 receptor by western blotting. In addition, 3 normal control bladder specimens were analyzed and compared with the KC specimens.ResultsThe KC bladder mucosa revealed significantly less expression of ZO-1 and E-cadherin, and greater expression of TUNEL and tryptase activity than the control samples. The expression of M3 and β3-AR in the KC specimens was significantly greater than in the controls. The expression of iNOS, eNOS, M2, and P2X3 was not significantly different between the KC and control specimens.ConclusionsThe bladder tissue of KC patients revealed significant urothelial dysfunction, which was associated with mast-cell mediated inflammation, increased urothelial cell apoptosis, and increased expression of the M3 and β3-AR.

Carbonic anhydrase enzymes regulate mast cell-mediated inflammation.

Type 2 cytokine responses are necessary for the development of protective immunity to helminth parasites but also cause the inflammation associated with allergies and asthma. Recent studies have found that peripheral hematopoietic progenitor cells contribute to type 2 cytokine-mediated inflammation through their enhanced ability to develop into mast cells. In this study, we show that carbonic anhydrase (Car) enzymes are up-regulated in type 2-associated progenitor cells and demonstrate that Car enzyme inhibition is sufficient to prevent mouse mast cell responses and inflammation after Trichinella spiralis infection or the induction of food allergy-like disease. Further, we used CRISPR/Cas9 technology and illustrate that genetically editing Car1 is sufficient to selectively reduce mast cell development. Finally, we demonstrate that Car enzymes can be targeted to prevent human mast cell development. Collectively, these experiments identify a previously unrecognized role for Car enzymes in regulating mast cell lineage commitment and suggest that Car enzyme inhibitors may possess therapeutic potential that can be used to treat mast cell-mediated inflammation.

Carbonic anhydrase enzymes regulate mast cell-mediated immunity to Trichinella spiralis.

Abstract Type 2 cytokine responses are necessary for the development of protective immunity to helminth parasites but also cause the detrimental inflammation associated with allergies and asthma. Recent studies have found that peripheral hematopoietic progenitor cell populations contribute to type 2 cytokine-mediated inflammation through their enhanced ability to develop into mast cells. Here we identify that carbonic anhydrase (Car) enzymes are upregulated in type 2-asscoaited progenitor cells and demonstrate that Car enzyme inhibition was sufficient to prevent murine mast cell development, type 2 cytokine-mediated inflammation and protective immunity to Trichinella spiralis. Moreover, we show that Car enzyme inhibition was also sufficient to prevent intestinal mast cell responses in a murine model of food allergy-like disease. Finally, we performed translational studies and demonstrated that Car enzymes can be targeted with an FDA-approved inhibitor to prevent human mast cell development. Collectively these studies identify a previously unrecognized role for Car enzymes in regulating mast cell lineage commitment and suggest that FDA-approved Car enzyme inhibitors may possess additional off-label therapeutic potential that can be employed to treat mast cell-mediated inflammation.

Decreased tryptophan and increased kynurenine levels in mastocytosis associated with digestive symptoms.

The main metabolism pathway of tryptophan is protein formation, but it can also be metabolized into serotonin and kynurenine. Indoleamine 2,3-dioxygenase (IDO) is the enzyme that catalyzes the degradation of tryptophan into kynurenine. Mastocytosis is a heterogeneous disease characterized by mast cell accumulation in various tissues with 57% of patients having gastrointestinal involvement. We studied tryptophan metabolism in mastocytosis patients displaying or not gastrointestinal features and healthy subjects (n = 26 in each group). Mastocytosis patients with digestive symptoms displayed significantly increased kynurenine level and IDO activity as compared to healthy controls and mastocytosis patients without digestive symptoms. This could be linked to mast cell-mediated digestive inflammation among patients with mastocytosis. This work is the first focusing on kynurenine pathway in a mast cell disease and could help to understand the pathogenesis of digestive features in mastocytosis as well as in other mast cell-mediated diseases.

Negative regulatory roles of ORMDL3 in the FcεRI-triggered expression of proinflammatory mediators and chemotactic response in murine mast cells.

Single-nucleotide polymorphism studies have linked the chromosome 17q12-q21 region, where the human orosomucoid-like (ORMDL)3 gene is localized, to the risk of asthma and several other inflammatory diseases. Although mast cells are involved in the development of these diseases, the contribution of ORMDL3 to the mast cell physiology is unknown. In this study, we examined the role of ORMDL3 in antigen-induced activation of murine mast cells with reduced or enhanced ORMDL3 expression. Our data show that in antigen-activated mast cells, reduced expression of the ORMDL3 protein had no effect on degranulation and calcium response, but significantly enhanced phosphorylation of AKT kinase at Ser 473 followed by enhanced phosphorylation and degradation of IκBα and translocation of the NF-κB p65 subunit into the nucleus. These events were associated with an increased expression of proinflammatory cytokines (TNF-α, IL-6, and IL-13), chemokines (CCL3 and CCL4), and cyclooxygenase-2 dependent synthesis of prostaglandin D2. Antigen-mediated chemotaxis was also enhanced in ORMDL3-deficient cells, whereas spreading on fibronectin was decreased. On the other hand, increased expression of ORMDL3 had no significant effect on the studied signaling events, except for reduced antigen-mediated chemotaxis. These data were corroborated by increased IgE-antigen-dependent passive cutaneous anaphylaxis in mice with locally silenced ORMDL3 using short interfering RNAs. Our data also show that antigen triggers suppression of ORMDL3 expression in the mast cells. In summary, we provide evidence that downregulation of ORMDL3 expression in mast cells enhances AKT and NF-κB-directed signaling pathways and chemotaxis and contributes to the development of mast cell-mediated local inflammation in vivo.