IgE-mediated Allergic Responses Research Articles

On the role of antibody affinity and avidity in the IgE-mediated allergic response.

Type I hypersensitivity, also known as classical allergy, is mediated via allergen-specific IgE antibodies bound to type I FcR (FcεRI) on the surface of mast cells and basophils upon cross-linking by allergens. This IgE-mediated cellular activation may be blocked by allergen-specific IgG through multiple mechanisms, including direct neutralization of the allergen or engagement of the inhibitory receptor FcγRIIb which blocks IgE signal transduction. In addition, co-engagement of FcεRI and FcγRIIb by IgE-IgG-allergen immune complexes causes down regulation of receptor-bound IgE, resulting in desensitization of the cells. Both, activation of FcεRI by allergen-specific IgE and engagement of FcγRIIb by allergen-specific IgG are driven by allergen-binding. Here we delineate the distinct roles of antibody affinity versus avidity in driving these processes and discuss the role of IgG subclasses in inhibiting basophil and mast cell activation.

Point-of-Care Fluorescence Biosensing System for Rapid Multi-Allergen Screening.

With the steady increase in allergy prevalence worldwide, there is a strong need for novel diagnostic tools for precise, fast, and less invasive testing methods. Herein, a miniatured fluorescence-based biosensing system is developed for the rapid and quantitative detection of allergen-specific immunoglobulin-E. An antibody-based fluorescence assay in a microfluidic-patterned slide, combined with a custom-made portable fluorescence reader for image acquisition and user-friendly software for the data analysis, enables obtaining results for multiple allergens in just ~1 h with only 80 μL of blood serum. The multiplexed detection of common birch, timothy grass, cat epithelia, house dust mite, and dog epithelia shows quantitative IgE-mediated allergic responses to specific allergens in control serum samples with known total IgE concentration. The responses are verified with different control tests and measurements with a commercial fluorescence reader. These results open the door to point-of-care allergy screening for early diagnosis and broader access and for large-scale research in allergies.

Hispidulin Alleviates Mast Cell-Mediated Allergic Airway Inflammation through FcεR1 and Nrf2/HO-1 Signaling Pathway.

Allergic asthma is a type 2 immune-response-mediated chronic respiratory disease. Mast cell activation influences the pathogenesis and exacerbation of allergic asthma. Therefore, the development of mast cell-targeting pharmacotherapy is important for managing allergic airway inflammation. We investigated the efficacy of hispidulin (HPD), natural flavone, in a mast-cell-mediated ovalbumin (OVA)-induced allergic airway inflammation model. HPD alleviated symptoms of allergic asthma and decreased the levels of immunoglobulin (Ig) E, type 2 inflammation, immune cell infiltration, and mast cell activation in the lung. Furthermore, in vivo analysis confirmed the efficacy of HPD through the evaluation of IgE-mediated allergic responses in a mast cell line. HPD treatment inhibited mast cell degranulation through inhibition of the FcεR1 signaling pathway and suppressed the expression of inflammatory cytokines (TNF-α, IL-4, IL-6, and IL-13) through suppression of the NF-κB signaling pathway. The antioxidant effects of HPD in activated mast cells were identified through modulation of antioxidant enzymes and the Nrf2/HO-1 signaling pathway. In conclusion, HPD may be a potential therapeutic candidate for allergic airway inflammation of asthma and acts by suppressing mast cell activation and oxidative stress.

A novel natural Syk inhibitor suppresses IgE-mediated mast cell activation and passive cutaneous anaphylaxis

Spleen tyrosine kinase (Syk) plays a crucial role as a target for allergy treatment due to its involvement in immunoreceptor signaling. The purpose of this study was to identify natural inhibitors of Syk and assess their effects on the IgE-mediated allergic response in mast cells and ICR mice. A list of eight compounds was selected based on pharmacophore and molecular docking, showing potential inhibitory effects through virtual screening. Among these compounds, sophoraflavanone G (SFG) was found to inhibit Syk activity in an enzymatic assay, with an IC50 value of 2.2 μM. To investigate the conformational dynamics of the SYK-SFG system, we performed molecular dynamics simulations. The stability of the binding between SFG and Syk was evaluated using root mean square deviation (RMSD) and root mean square fluctuation (RMSF). In RBL-2H3 cells, SFG demonstrated a dose-dependent suppression of IgE/BSA-induced mast cell degranulation, with no significant cytotoxicity observed at concentrations below 10.0 μM within 24 h. Furthermore, SFG reduced the production of TNF-α and IL-4 in RBL-2H3 cells. Mechanistic investigations revealed that SFG inhibited downstream signaling proteins, including phospholipase Cγ1 (PLCγ1), as well as mitogen-activated protein kinases (AKT, Erk1/2, p38, and JNK), in mast cells in a dose-dependent manner. Passive cutaneous anaphylaxis (PCA) experiments demonstrated that SFG could reduce ear swelling, mast cell degranulation, and the expression of COX-2 and IL-4. Overall, our findings identify naturally occurring SFG as a direct inhibitor of Syk that effectively suppresses mast cell degranulation both in vitro and in vivo.

Potential honey bee (Apis mellifera) allergens associated with IgE-mediated allergy- An In-silico study.

Allergies due to honeybee venom (HBV) are reported to be the second most common form of allergy toHymenopteravenom that occurs after being stung. Indeed, 15-20% of people test IgE positive after being stung. However, accurate data on the incidence of honey bee allergens is missing and estimated to be less than 0.001%. Beekeeping is an ancient and widely practiced activity across the Kingdom of Saudi Arabia. Still, studies on the allergenic effect of the different subspecies of honey bees are very rare in Saudi Arabia. Hence, in this study, using the In-silico approach, we aimed to study and evaluate the effect of allergens from honey bees in Ha'il City, Saudi Arabia on IgE-mediated allergies. A list of potential allergens from Apis mellifera was prepared, and the 3D structure was prepared using the SWISS-MODEL web server and the PDB database was used for retrieving the structure of the immunoglobulin E- fragment antigen-binding (IgE-Fab) region. Molecular docking (clusPro webserver) and molecular dynamics (Schrödinger) results revealed that the B2D0J5 protein from Apis mellifera might be the key protein associated with IgE-mediated allergic response. Overall, the identified knowledge can be used for exploring prophylactic vaccine candidates and improving the diagnosis of allergic reactions to honey bees in the Ha'il region of Saudi Arabia.

AT9283, 1-Cyclopropyl-3-(3-(5-(Morpholinomethyl)-1H-Benzo[d]Imidazole-2-yl)-1H-Pyrazol-4-yl) Urea, Inhibits Syk to Suppress Mast Cell-Mediated Allergic Response.

Mast cells are an effector cell that plays a pivotal role in type I hypersensitive immune responses. Mast cells exist in connective tissues, such as skin and mucosal tissue, and contain granules which contain bioactive substances such as histamine and heparin in cells. The granules of mast cells are secreted by antigen stimulation to cause the type I allergic hypersensitivity. In addition, stimulated by antigen, mast cells synthesize and secrete various eicosanoids and cytokines. While AT9283 is known to have anticancer effects, the therapeutic effect of AT9283 on allergic disorders is completely unknown. In this study, it was found that AT9283 reversibly inhibited antigen-IgE binding-induced degranulation in mast cells (IC50, approx. 0.58 μM) and suppressed the secretion of the inflammatory cytokines IL-4 (IC50, approx. 0.09 μM) and TNF-α (IC50, approx. 0.19 μM). For a mechanism of mast cell inhibition, while not inhibiting Syk phosphorylation, AT9283 suppressed the activation of LAT, a downstream substrate protein of Syk, in a dose-dependent manner. As expected, AT9283 also inhibited the activation of PLCγ1 and Akt, downstream signaling molecules of Syk/LAT, and MAP kinases such as JNK, Erk1/2, and P38. In an in vitro protein tyrosine kinase assay, AT9283 directly inhibited Syk activity. Next, AT9283 dose-dependently inhibited passive cutaneous anaphylaxis (PCA), an IgE-mediated allergic acute response, in mice (ED50, approx. 34 mg/kg, p.o.). These findings suggest that AT9283 has potential to use as a new drug for alleviating the symptoms of IgE-mediated allergic disorders.

IL-27 signaling negatively regulates FcɛRI-mediated mast cell activation and allergic response.

IL-27 is a member of the IL-12 family, exerting both anti- and pro-inflammatory activity in a cell-dependent and disease context-specific manner. Antigen-mediated cross-linking of IgE on mast cells triggers a signaling cascade that results in mast cell degranulation and proinflammatory cytokine production, which are key effectors in allergic reactions. Here, we show that the activation of mast cells is negatively regulated by IL-27 signaling. We found that mice lacking IL-27Rα (WSX-1) displayed increased sensitivity to IgE-mediated skin allergic response and chronic airway inflammation. The bone marrow-derived mast cells (BMMCs) of IL-27Rα-deficient mouse showed greater high-affinity receptor Fc epsilon RI (FcεRI)-mediated activation with significantly enhanced degranulation and cytokine production. Mechanistically, the dysregulated signaling in IL-27Rα-/- mast cells is associated with increased activation of Grb2-PLC-γ1-SLP-76, PI3K/Akt/IκBα signaling and decreased phosphorylation level of SH2 domain-containing protein phosphatase1 (SHP1). Furthermore, IL-27 treatment could inhibit mast cell activation directly, and retrovirus-based IL-27 expression in lung attenuated the airway inflammation in mice. Collectively, our findings reveal that IL-27 signaling negatively regulates mast cell activation and its mediated allergic response.

Fel d1 Blocking Antibodies: A Novel Method to Reduce IgE-Mediated Allergy to Cats.

Fel d1 is an important allergen produced by cats that causes IgE reactions in up to 95% of cat-allergic adults. Immunotherapy to reduce human allergy to cats has demonstrated that people have the capacity to produce allergen-specific neutralizing antibodies that block IgE-mediated allergic responses. We wished to determine if “blocking” antibodies could be used to reduce the IgE binding ability of cat allergens prior to their exposure to humans. Here, we describe the characterization of Fel d1-specific antibodies. We demonstrated the efficacy of a rabbit polyclonal and an allergen-specific chicken IgY to bind to Fel d1 in cat saliva and block Fel d1-IgE binding and IgE-mediated basophil degranulation. Fel d1 blocking antibodies offer a new and exciting approach to the neutralization of cat allergens.

Inhibition of c-Fos expression attenuates IgE-mediated mast cell activation and allergic inflammation by counteracting an inhibitory AP1/Egr1/IL-4 axis

BackgroundActivator protein-1 (AP1), a c-Fos–JUN transcription factor complex, mediates many cytobiological processes. c-Fos has been implicated in immunoglobulin (Ig)E activation of mast cells (MCs) via high-affinity IgE Fc receptor (FcεRI) binding. This study examined c-Fos involvement in MC activation and tested the effects of the c-Fos/AP1 inhibitor T-5224 on MCs activation and allergic responses.MethodsIn vitro studies were conducted with two MC model systems: rat basophilic leukemia cells (RBLs) and mouse bone marrow derived mast cells (BMMCs). MC degranulation and effector functions were examined with β-hexosaminidase release and cytokine secretion assays. c-Fos/AP1 was inhibited with T-5224. c-Fos activity was suppressed with short hairpin RNA targeting c-Fos (shFos). In vivo immune responses were evaluated in passive cutaneous anaphylaxis (PCA) and ovalbumin-induced active systemic anaphylaxis (ASA) models, as well as in an oxazolone (OXA)-induced model of atopic dermatitis, a common allergic disease.Resultsc-Fos expression was elevated transcriptionally and translationally in IgE-stimulated MCs. c-Fos binding of the Egr1 (early growth response 1) promoter upregulated Egr1 transcription, leading to production of interleukin (IL)4. T-5224 reduced FcεRI-mediated MC degranulation (evidenced by β-hexosaminidase activity and histamine levels) and diminished EGR1 and IL4 expression. T-5224 attenuated IgE-mediated allergic responses in PCA and ASA models, and it suppressed MC-mediated atopic dermatitis in mice.ConclusionIgE binding can activate MCs via a c-Fos/Egr1/IL-4 axis. T-5224 suppresses MC activation in vitro and in vivo and thus represents a promising potential strategy for targeting MC activation to treat allergic diseases.

Intranasal delivery of allergen in a nanoemulsion adjuvant inhibits allergen-specific reactions in mouse models of allergic airway disease.

Atopic diseases are an increasing problem that involve both immediate hypersensitivity reactions mediated by IgE and unique cellular inflammation. Many forms of specific immunotherapy involve the administration of allergen to suppress allergic immune responses but are focused on IgE-mediated reactions. In contrast, the effect of allergen-specific immunotherapy on allergic inflammation is complex, not entirely consistent and not well understood. We have previously demonstrated the ability of allergen administered in a nanoemulsion (NE) mucosal adjuvant to suppress IgE-mediated allergic responses and protect from allergen challenge in murine food allergy models. This activity was associated with decreases in allergen-specific IL-10 and reductions in allergic cytokines and increases in regulatory T cells. Here, we extend these studies to using 2 distinct models, the ovalbumin (OVA) and cockroach (CRA) models of allergic airway disease, which are based predominantly on allergic inflammation. Acute or chronic allergic airway disease was induced in mice using ovalbumin and cockroach allergen models. Mice received three therapeutic immunizations with allergen in NE, and reactivity to airway challenge was determined. Therapeutic immunization with cockroach or OVA allergen in NE markedly reduced pathology after airway challenge. The 2 models demonstrated protection from allergen challenge-induced pathology that was associated with suppression of Th2-polarized immune responses in the lung. In addition, the reduction in ILC2 numbers in the lungs of allergic mice along with reduction in epithelial cell alarmins, IL-25 and IL-33, suggests an overall change in the lung immune environment induced by the NE immunization protocol. These results demonstrate that suppression of allergic airway inflammation and bronchial hyper-reactivity can be achieved using allergen-specific immunotherapy without significant reductions in allergen-specific IgE and suggest that ILC2 cells may be critical targets for this activity.

IL-33 sensitizes mast cells to respond to a second alarmin, ATP, but not PAMPs.

Abstract Barrier defenses must respond rapidly to pathogen infection but avoid responses to innocuous antigens or microbes. Aberrant recognition can lead to hypersensitivity and/or chronic inflammatory conditions. Barrier tissues actively release alarmins such as IL-33 in response to cell stress helping to identify pathogenic stimuli. Additional alarmin molecules such as ATP, uric acid crystals, and HMBG protein have also been identified but it remains unclear how the innate immune system integrates these multiple signals. We have used mast cells to address how IL-33 and ATP signals interact. While best known for their role in IgE-mediated allergic responses, mast cells readily produce inflammatory cytokines in response to either IL-33 or ATP. Measuring secreted IL-6, TNF alpha and IL-13 we found that bone marrow-derived and peritoneal mast cells had a 3–6-fold increased response to ATP when sensitized by pretreatment with IL-33. This synergistic effect was observed at relatively low ATP concentrations (<300μM) and required <1hr exposure to IL-33. Stimulating with PAMPs (LPS, LTA, poly I:C, ß-glucan) failed to show any enhancement with IL-33 pretreatment. Inhibitor studies indicated that the P2X3 receptor and the NFAT transcription factor mediated the ATP response. To test IL-33 and ATP synergy in vivo, we assessed eosinophil mobilization to the peritoneal cavity in mice. Consistent with the in vitro studies, co-administration of IL-33 and ATP increased eosinophil numbers compared to either alarmin alone. These results support the hypothesis that alarmins collaborate to surpass a threshold level of barrier stress necessary to initiate an inflammatory immune response.

Protein and Antibody Engineering: Suppressing Degranulation of the Mast Cells and Type I Hypersensitivity Reaction.

With an increase in atopic cases and owing to a significant role of mast cells in type I hypersensitivity, a therapeutic need to inhibit degranulation of mast cells has risen. Mast cells are notorious for IgE-mediated allergic response. Advancements have allowed researchers to improve clinical outcomes of already available therapies. Engineered peptides and antibodies can be easily manipulated to attain desired characteristics as per the biological environment. A number of these molecules are designed to target mast cells in order to regulate the release of histamine and other mediators, thereby controlling type I hypersensitivity response. The aim of this review paper is to highlight some of the significant molecules designed for the purpose.

Physiopathologie de la dermatite atopique et des autres maladies atopiques : une approche globale est-elle possible ?: Pathophysiology of atopic dermatitis and other atopic diseases: is a global approach possible?

Atopy is defined by the propensity to develop an exaggerated type-2 inflammatory response to environmental molecules. Clinically, atopy is diagnosed when atopic disease occurs: atopic dermatitis, food allergy, atopic asthma and allergic rhinitis and conjunctivitis. Whereas the classical "atopic march" is increasingly challenged through epidemiological studies, type-2 cellular inflammation is a characteristic shared by the atopic diseases. This inflammation can be innate (non-specific: eosinophils, mast cells, dendritic cells, innate lymphoid cells [ILC]), or adaptive (antigen-specific, involving T cells). Interleukins (IL-)4, 5 and 13 are major actors of type-2 inflammation and are mainly produced by ILC and T cells. The efficacy of treatments targeting these type-2 cytokines highlight the importance of type-2 inflammation in atopic diseases. However, several patients do not respond to type-2 targeting treatments, highlighting the presence of other actors in pathophysiology of atopic diseases: alteration of epithelial barrier, IgE-mediated allergic responses, type-17 inflammation. Thus, the term "endotype" can illustrate this diversity in pathophysiology. Finally, a global approach of atopic diseases, as type-2 inflammatory diseases, is fundamental, but not sufficient. An approach by endotype is advisable, in a personalized medicine perspective. © 2020 Elsevier Masson SAS. All rights reserved.

Two Japanese pepper (Zanthoxylum piperitum) fruit-derived compounds attenuate IgE-mediated allergic response in vitro and in vivo via inhibition of mast cell degranulation

Zanthoxylum piperitum (ZP, ‘Japanese pepper’) is a traditional medicine and pepper used in Asian countries such as Japan. Hydroxy-α-sanshool, a pungent-tasting substance contained within ZP, has been reported to slightly suppress immunoglobulin E (IgE)-mediated mast cell degranulation. The current study aims to newly identify anti-allergic compounds derived from ZP. We examine the inhibitory mechanisms behind IgE-mediated mast cell degranulation. By inhibitory effect-guided isolation, we identified degranulation inhibitory compounds derived from ZP fruit: 1-acetoxy-7-hydroxy-3, 7-dimethylocta-2E, 5E-diene (ZP1) and 8-hydroxygeranyl acetate (ZP2). ZP1 and ZP2 inhibited IgE-mediated degranulation and A23187-mediated degranulation in RBL-2H3 mast cells. Our findings suggest the inhibition of degranulation by ZP1 and ZP2 was by inhibition of Lyn phosphorylation, followed by inhibition of intracellular Ca2+ mobilization, protein kinase C alpha phosphorylation, membrane ruffling, and granule-to-plasma membrane fusion. Oral administration of ZP1 or ZP2 attenuated an IgE-mediated passive cutaneous anaphylactic reaction in mice. Histological observation suggests that this effect occurred via inhibition of mast cell degranulation. These findings indicate that ZP1 and ZP2 attenuate allergic reaction via inhibition of IgE-mediated mast cell degranulation.

Direct Inhibition of the Allergic Effector Response by Raw Cow's Milk-An Extensive In Vitro Assessment.

The mechanisms underlying the allergy-protective effects of raw cow’s milk are poorly understood. The current focus is mainly on the modulation of T cell responses. In the present study, we investigated whether raw cow’s milk can also directly inhibit mast cells, the key effector cells in IgE-mediated allergic responses. Primary murine bone marrow-derived mast cells (BMMC) and peritoneal mast cells (PMC), were incubated with raw milk, heated raw milk, or shop milk, prior to IgE-mediated activation. The effects on mast cell activation and underlying signaling events were assessed. Raw milk was furthermore fractionated based on molecular size and obtained fractions were tested for their capacity to reduce IgE-mediated mast cell activation. Coincubation of BMMC and PMC with raw milk prior to activation reduced β-hexosaminidase release and IL-6 and IL-13 production, while heated raw milk or shop milk had no effect. The reduced mast cell activation coincided with a reduced intracellular calcium influx. In addition, SYK and ERK phosphorylation levels, both downstream signaling events of the FcεRI, were lower in raw milk-treated BMMC compared to control BMMC, although differences did not reach full significance. Raw milk-treated BMMC furthermore retained membrane-bound IgE expression after allergen stimulation. Raw milk fractionation showed that the heat-sensitive raw milk components responsible for the reduced mast cell activation are likely to have a molecular weight of > 37 kDa. The present study demonstrates that raw cow’s milk can also directly affect mast cell activation. These results extend the current knowledge on mechanisms via which raw cow’s milk prevents allergic diseases, which is crucial for the development of new, microbiologically safe, nutritional strategies to reduce allergic diseases.

Thymoquinone attenuates IgE-mediated allergic response via pi3k-Akt-NFκB pathway and upregulation of the Nrf2-HO1 axis.

IgE-dependent reactions mediate the majority of allergic diseases. This study explores the effects of thymoquinone (Tq) on IgE-mediated allergic response in activated mast cells, basophils, and neutrophils. Tq treatment resulted in a dose-dependent decrease in levels of TNF-α and IL-4 in activated RBL-2H3 cells. Tq inhibited the degranulation of these cells with an IC50 value of 56.37µM. Moreover, the compound suppressed basophil activation induced through FcεRI receptors with an IC50 value of 45.76µM in heparinized human whole blood. Likewise, neutrophil migration and elastase activity were dose-dependently reduced. While Tq decreased the phosphorylation of Akt and NFκB in activated RBL-2H3 cells, it increased nuclear Nrf2 and HO-1 antioxidant proteins. Our results indicate that Tq possesses demonstrable activity in cellular models of IgE-mediated allergic reactions. PRACTICAL APPLICATIONS: The current study sheds light on the mechanistic pathways of Tq on IgE-based response in activated mast cells, basophils, and neutrophils. The output of this preclinical in vitro study may be translated into better chemotherapeutic applications of Tq and its analogs in the treatment of allergic inflammation. However, a detailed investigation of in vivo models is recommended.

In vitro anti-inflammatory effects of curcumin on mast cell-mediated allergic responses via inhibiting FcεRI protein expression and protein kinase C delta translocation.

Allergy is a hypersensitivity reaction when exposed to certain environmental substances. It shows high relation between immunoglobulin E (IgE) binding to a specific receptor (FcεRI), pro-inflammatory cytokines, and mediators with allergic inflammation responses. Curcumin is a yellow pigment isolated from the turmeric. Curcumin possesses antioxidant and anti-inflammatory properties as well as exhibits significant chemopreventive activity. This study was aimed to investigate the in vitro assessment of the regulation of curcumin on allergic inflammatory responses on rat basophil leukemia (RBL)-2H3 and human pre-basophils (KU812) cell lines. Curcumin showed the activity against histamine and β-hexosaminidase releases from both IgE-mediated and A23187-induced cells degranulation. The morphological observation also confirmed that curcumin inhibits cells degranulation. IgE-mediated allergic responses and significantly induced mast cells intracellular reactive oxygen species (ROS) production. Curcumin reduced ROS production from IgE-mediated or A23187-induced cells degranulation. Curcumin also successfully reduced FcεRI expressions and some pro-inflammatory cytokines, such as interleukin (IL)-4 and IL-13. Furthermore, curcumin inhibited protein kinase C (PKC)-δ translocation from cytosolic to particulate. These results suggested that curcumin can alleviate both the IgE-mediated and calcium ionosphere A23187-stimulated allergic responses through reducing the release of the allergic mediators.

WZ3146 inhibits mast cell Lyn and Fyn to reduce IgE-mediated allergic responses in vitro and in vivo

Mast cells (MCs) play an important role as effector cells that cause allergic responses in allergic diseases. For these reasons, MC is considered an attractive therapeutic target for allergic disease treatment. In this study, we investigated the inhibitory effect of WZ3146, N-[3-[5-chloro-2-[4-(4-methylpiperazin-1-yl)anilino]pyrimidin-4-yl]oxyphenyl]prop-2-enamide, and the mechanisms of its actions on the MC activation and IgE-mediated allergic response by using three types of MCs such as rat basophilic leukemia (RBL)-2H3 cells, mouse bone marrow mast cells (BMMCs), and human Laboratory of Allergic Diseases 2 (LAD2) cells. WZ3146 inhibited antigen-stimulated degranulation in a dose-dependent manner (IC50, ~ 0.35 μM for RBL-2H3 cells; ~ 0.39 μM for BMMCs; ~ 0.41 for LAD2 cells). WZ3146 also suppressed the production of histamine, tumor necrosis factor (TNF)-α and interleukin (IL)-6, which mediate various allergic responses, in a dose-dependent manner. As the mechanism of WZ3146 to inhibit MCs, it inhibited the activation of spleen tyrosine kinase (Syk) and the downstream signaling proteins of Syk such as linker for activation of T cell (LAT) and phospholipase (PL) Cγ1 in the signaling pathway of FcεRI. In addition, WZ3146 inhibited the activation of Akt, extracellular signal-regulated kinase (ERK)1/2, p38, and c-Jun N-terminal kinase (JNK). However, WZ3146 did not inhibit degranulation of MCs by thapsigargin or ionomycin, which increase calcium concentration in cytosol. Notably, WZ3146 inhibited the activity of Lyn and Fyn, but not Syk. In an following animal experiment, WZ3146 inhibited IgE-dependent passive cutaneous anaphylaxis (PCA) in a dose-dependent manner (ED50, ~ 20 mg/kg). Taken together, in this study we show that the pyrimidine derivative, WZ3146, inhibits the IgE-mediated allergic response by inhibiting Lyn and Fyn Src-family kinases, which are initially activated by antigen stimulation in MCs. Therefore, we propose that WZ3146 could be used as a new therapeutic agent for the treatment of allergic diseases.

Therapeutic Potential of Morin in Ovalbumin-induced Allergic Asthma Via Modulation of SUMF2/IL-13 and BLT2/NF-kB Signaling Pathway.

Allergic asthma is a chronic immune-inflammatory disorder, characterized by airway inflammation and airway hyperresponsiveness (AHR). Morin is a natural flavonoid reported to exhibit inhibitory action against IgE-mediated allergic response. To determine the efficacy of murine model of ovalbumin (OVA)-induced AHR inhibition by morin and decipher the molecular mechanism involved. Sprague-Dawley rats were sensitized and challenged with OVA to induce AHR. Rats received treatment with morin (10, 30 and 100 mg/kg, p.o.) for the next 28 days. Morin (30 and 100 mg/kg) significantly and dose-dependently attenuated (p < 0.01 and p < 0.001) OVA-induced alterations in pulse oxy and lung function test, increased bronchoalveolar lavage fluid cell counts, elevated total protein and albumin levels in serum, BALF, and lungs, increased serum total and OVA-specific IgE levels and, elevated oxidative stress levels in the lung. RT-PCR analysis revealed that morin treatment (30 and 100 mg/kg) significantly (p < 0.001) up-regulated SUMF2 mRNA expression in lungs whereas mRNA expressions of BLT2, NF-κB, and Th2-cytokine (TNF-α, IL-1β, IL-4, IL-6, and IL-13) were down-regulated significantly and dose-dependently (p < 0.01 and p < 0.001). Also, histologic and ultrastructural studies showed that morin significantly inhibited (p < 0.001) OVAinduced perivascular and peribranchial inflammatory infiltration and interstitial fibrosis. Morin exhibited inhibitory effect against OVA-induced allergic asthma by activation of SUMF2 which impeded IL-13 expression and in turn attenuated Th2-cytokines, BLT2, NF-κB, and IgE levels to ameliorate AHR. Thus, our findings suggested that morin could be considered as a potential alternative therapeutic agent for the management of allergic asthma.

Characterization of thymoquinone/hydroxypropyl-β-cyclodextrin inclusion complex: Application to anti-allergy properties

Thymoquinone is an effective phytochemical compound in the treatment of various diseases. However, its practical administration has been limited due to poor aqueous solubility and bioavailability. In this work, we developed a novel inclusion complex of thymoquinone and hydroxypropyl-β-cyclodextrin that features improved solubility and bioactivity. The drug solubility was markedly accelerated in the increasing ratio of hydroxypropyl-β-cyclodextrin to thymoquinone amount. The formation of the thymoquinone/hydroxypropyl-β-cyclodextrin inclusion complex was evidenced using X-ray diffraction, differential scanning calorimetry, thermal gravimetric analysis, Fourier transform infrared, scanning electron microscopy and nuclear magnetic resonance. The release behavior of the complex, as well as of their mixtures, was examined in artificial gastric (pH 1.2) and intestinal (pH 6.8) dissolution media. The formulated complex released the drug rapidly at the initial stage, followed by a slow release. Thermodynamic parameters ΔH, ΔS and ΔG were calculated with temperatures ranging from 20 to 45 °C to evaluate the complexation process. The activity of the inclusion complex was evaluated on IgE-mediated allergic response in rat basophilic leukemia (RBL-2H3) cells by monitoring key allergic mediators. The results revealed that compared with free thymoquinone, the inclusion complex more strongly inhibited the release of histamine, tumor necrosis factor-α, and interleukin-4, and was not cytotoxic at the tested thymoquinone concentrations (0.125–4 μg/mL) indicating the inclusion complex possibly had better antiallergic effects. Our finding suggested that the inclusion complex achieved prolonged action and reduced side-effect of thymoquinone.