Allergic Inflammation In Mice Research Articles

Investigation of gut microbiota changes and allergic inflammation of mice with milk protein-induced allergic enteritis.

This study aimed to investigate the changes of gut microbiota and allergic inflammation in mice with allergic enteritis caused by milk protein. In this study, female BALB\C mice in the whey protein (WP-sensitized) group were gavaged with WP and normal saline, the sham-sensitized group was given normal saline once a week for 5 weeks. One week later, the WP-sensitized mice were administered 60mg β-lactoglobulin (BLG). The results showed that mice's body weight decreased, feces with loose and bloody, and systemic allergic reactions and ear swelling increased in the WP-sensitized group. The levels of WP-specific Ig, mMCP-1, calprotectin of feces, and inflammation-related factors in the WP-sensitized group were increased. WP-sensitized group intestine tissues were damaged severely and the expressions of ZO-1, Claudin-1, and Occludin reduced. The results of 16S rRNA sequencing showed that there were differences in operational taxonomic units (OUT) levels of gut microbes between the two groups, o_Clostridiales, c_Clostridia, and f_Lachnospiraceae were more abundant in the WP-sensitized group. In conclusion, the WP sensitization can induce the allergic inflammation, intestinal injury and intestinal barrier dysfunction in mice, and the gut microbes were also changed, which provided a reference for the treatment of WP-sensitized mice.

Dietary digestible carbohydrates are associated with higher prevalence of asthma in humans and with aggravated lung allergic inflammation in mice.

Dietary carbohydrates and fats are intrinsically correlated within the habitual diet. We aimed to disentangle the associations of starch and sucrose from those of fat, in relation to allergic sensitization, asthma and rhinoconjuctivitis prevalence in humans, and to investigate underlying mechanisms using murine models. Epidemiological data from participants of two German birth cohorts (age 15) were used in logistic regression analyses testing cross-sectional associations of starch and sucrose (and their main dietary sources) with aeroallergen sensitization, asthma and rhinoconjunctivitis, adjusting for correlated fats (saturated, monounsaturated, omega-6 and omega-3 polyunsaturated) and other covariates. For mechanistic insights, murine models of aeroallergen-induced allergic airway inflammation (AAI) fed with a low-fat-high-sucrose or -high-starch versus a high-fat diet were used to characterize and quantify disease development. Metabolic and physiologic parameters were used to track outcomes of dietary interventions and cellular and molecular responses to monitor the development of AAI. Oxidative stress biomarkers were measured in murine sera or lung homogenates. We demonstrate a direct association of dietary sucrose with asthma prevalence in males, while starch was associated with higher asthma prevalence in females. In mice, high-carbohydrate feeding, despite scant metabolic effects, aggravated AAI compared to high-fat in both sexes, as displayed by humoral response, mucus hypersecretion, lung inflammatory cell infiltration and TH 2-TH 17 profiles. Compared to high-fat, high-carbohydrate intake was associated with increased pulmonary oxidative stress, signals of metabolic switch to glycolysis and decreased systemic anti-oxidative capacity. High consumption of digestible carbohydrates is associated with an increased prevalence of asthma in humans and aggravated lung allergic inflammation in mice, involving oxidative stress-related mechanisms.

AMPK suppresses Th2 cell responses by repressing mTORC2

Allergic inflammation is a T helper 2 (Th2) cell-driven pathophysiological phenomenon, but the mechanism by which the metabolic cascade affects Th2 cell differentiation remains unclear. In this study, we investigated the roles of AMP-activated protein kinase (AMPK) and intracellular energy sensors in Th2 cell differentiation and the pathogenesis of allergic inflammation. Accordingly, T-cell-specific AMPK or Sirtuin 1 (Sirt1)-knockout mice were subjected to allergic inflammation, and their Th2 cell responses were investigated. The results demonstrated that inducing allergic inflammation in AMPK- and Sirt1-knockout mice increased Th2 cell responses and exacerbated allergic phenotypes. Furthermore, treatment with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), an activator of AMPK, ameliorated allergic inflammation in mice. Mechanistically, our findings revealed that AMPK repressed mechanistic target of rapamycin complex 2 (mTORC2), which downregulated the expression of suppressor of cytokine signaling 5 (SOCS5) in CD4+ T cells. In addition, the loss of AMPK signaling reduced SOCS5 expression and increased interleukin-4-STAT6–GATA3 axis-mediated Th2 cell differentiation. Finally, the T-cell-specific deletion of Rictor, a member of mTORC2, in Sirt1T-KO mice led to the reversal of allergic exacerbation to the level in control mice. Overall, our findings suggest that AMPK in CD4+ T cells inhibits the differentiation of Th2 cells by repressing mTORC2 and thus serves as a potential target for Th2 cell-associated diseases.

Nano-silica particles synergistically IgE-mediated mast cell activation exacerbating allergic inflammation in mice.

BackgroundAllergic respiratory diseases have increased dramatically due to air pollution over the past few decades. However, studies are limited on the effects of inorganic components and particulate matter with different particle sizes in smog on allergic diseases, and the possible molecular mechanism of inducing allergies has not been thoroughly studied.MethodsFour common mineral elements with different particle sizes in smog particles were selected, including Al2O3, TiO2, Fe2O3, and SiO2. We studied the relationship and molecular mechanism of smog particle composition, particle size, and allergic reactions using mast cells, immunoglobulin E (IgE)-mediated passive cutaneous anaphylaxis (PCA) model, and an ovalbumin (OVA)-induced asthmatic mouse model in vitro and in vivo, combined with transmission electron microscopy, scanning transmission X-ray microscopy analysis, and transcriptome sequencing.ResultsOnly 20 nm SiO2 particles significantly increased β-hexosaminidase release, based on dinitrophenol (DNP)-human serum albumin (HSA) stimulation, from IgE-sensitized mast cells, while other particles did not. Meanwhile, the PCA model showed that Evan’s blue extravasation in mice was increased after treatment with nano-SiO2 particles. Nano-SiO2 particles exposure in the asthmatic mouse model caused an enhancement of allergic airway inflammation as manifested by OVA-specific serum IgE, airway hyperresponsiveness, lung inflammation injury, mucous cell metaplasia, cytokine expression, mast cell activation, and histamine secretion, which were significantly increased. Nano-SiO2 particles exposure did not affect the expression of FcϵRI or the ability of mast cells to bind IgE but synergistically activated mast cells by enhancing the mitogen-activated protein kinase (MAPK) signaling pathway, especially the phosphorylation levels of the extracellular signal-regulated kinase (ERK)1/2. The ERK inhibitors showed a significant inhibitory effect in reducing β-hexosaminidase release.ConclusionOur results indicated that nano-SiO2 particles stimulation might synergistically activate IgE-sensitized mast cells by enhancing the MAPK signaling pathway and that nano-SiO2 particles exposure could exacerbate allergic inflammation. Our experimental results provide useful information for preventing and treating allergic diseases.

Highly efferocytic M2 macrophages are generated at the termination stage of basophil-dependent skin allergy and dampen the inflammation

Basophils play critical roles in the development of delayed-onset skin allergic inflammation in mice (IgE-CAI model). Importantly, they also contribute to the resolution of allergic inflammation by promoting the generation of M2 macrophages. However, it remains unclear how M2 macrophages suppress excess inflammation. To address this, we conducted single-cell RNA-seq (scRNA-seq) analysis of the IgE-CAI skin lesion. scRNA-seq analysis identified two distinct M2 macrophage populations, namely early and late M2 macrophages, in IgE-CAI skin lesion. The former population was preferentially observed at the peak of inflammation (3 days after allergen challenge), whereas the latter one at the termination phase of inflammation (5 days). Gene ontology analysis revealed that genes associated with phagocytosis were enriched in late M2 macrophages. In particular, late M2 macrophages displayed upregulated expression of Gas6 and Mertk, key genes responsible for phagocytic clearance of apoptotic cells. Of note, MERTK inhibitor significantly aggravated ear swelling and accumulation of inflammatory cells in the skin lesion. Taken together, scRNA-seq identified M2 macrophages that display high capacity of dead cell clearance and contribute to the resolution of IgE-CAI.

FAK inhibitor PF-431396 suppresses IgE-mediated mast cell activation and allergic inflammation in mice

Mast cells (MCs) initiate and maintain allergic inflammation. Upon being stimulated with immunoglobulin (Ig)E and antigen (Ag), MCs exhibit FcεRI (high-affinity IgE) receptor-mediated degranulation, cytokine secretion, and increased focal adhesion kinase (FAK) activity. The aims of this study were to examine mechanisms of FAK regulation in IgE-mediated MC activation and the effects of FAK inhibition on MC-mediated allergic responses. FAK activity was manipulated with short hairpin RNA (shRNA) knockdown, FAK overexpression, and the FAK inhibitor PF-431396 (PF). Gene expression and kinase activation were analyzed with quantitative molecular biology assays. PF effects were tested in the passive cutaneous anaphylaxis (PCA), active systemic anaphylaxis (ASA), and allergic conjunctivitis (AC) mouse models. Our results showed that FAK overexpression increased IgE-mediated degranulation and reduced the dexamethasone inhibitory effect on MCs activation. The FAK inhibitor PF diminished MC release of β-hexosaminidase (β-hex), histamine, and inflammatory cytokines, via a mechanism that involves MAPK and NF-κB signaling pathways. CaMKII was identified as a robust FAK-associating protein. Inhibition of CaMKII activation by KN-93 suppressed FAK activity and its downstream pathway. PF attenuated inflammatory responses in our PCA and ASA models, and relieved signs of allergic disease in AC model mice. In conclusions, MC degranulation and production of inflammatory mediators in allergic disease may be consequent to FcεRI crosslinking inducing CaMKII-mediated activation of FAK activity. FAK inhibition may represent a new MC-suppressing treatment strategy for the treatment of allergic diseases.

Correlation between the seasonal variations in phlorotannin content and the antiallergic effects of the brown alga Ecklonia cava subsp. stolonifera

Here, we investigated the antiallergic effects of brown algae (Ecklonia cava subsp. stolonifera) harvested on the coast of Nishinoshima Town, Oki Islands, Shimane Prefecture, Japan. Specifically, we investigated whether the active components of E. cava ssp. stolonifera vary seasonally and whether such variation correlates with seasonal variation in the antiallergic effects of the algae. High-performance liquid chromatography, liquid chromatography–mass spectrometry, and nuclear magnetic resonance spectra were used to analyze 80% methanol (80M) extract from E. cava ssp. stolonifera and to isolate eckol, 6,6′-bieckol, 8,8′-bieckol, dieckol, and phlorofucofuroeckol-A as the major active compounds in the species' phlorotannins. Monthly measurements of phlorotannin content and the isolated phlorotannins in the 80M extract showed that the crude phlorotannin and isolated phlorotannin content was higher in the summer–autumn period than in the winter–spring period. The antiallergic effects of the monthly 80M extracts and the isolated phlorotannins were confirmed by measuring inhibition of inflammation-related enzymatic activity, suppression of degranulation in rat basophile leukemia-2H3 cells, and effects on allergic inflammation in Institute of Cancer Research strain mice. There was partial seasonal variation in the effect of 80M extract in combination with the amount of isolated phlorotannin and the antiallergic effects. When phlorotannin was removed from the extracts using polyvinylpolypyrrolidone, the antiallergic effects were no longer observed; thus, phlorotannins are the active components. Overall, these results show that correlations exist between the seasonal variations in the suppressive effect of 80M extract on enzymatic activities, allergic inflammation in mice, and the amount of isolated phlorotannins.

MelLec Exacerbates the Pathogenesis of Aspergillus fumigatus-Induced Allergic Inflammation in Mice.

Environmental factors, particularly fungi, influence the pathogenesis of allergic airway inflammation, but the mechanisms underlying these effects are still unclear. Melanin is one fungal component which is thought to modulate pulmonary inflammation. We recently identified a novel C-type lectin receptor, MelLec (Clec1a), which recognizes fungal 1,8-dihydroxynaphthalene (DHN)-melanin and is able to regulate inflammatory responses. Here we show that MelLec promotes pulmonary allergic inflammation and drives the development of Th17 T-cells in response to spores of Aspergillus fumigatus. Unexpectedly, we found that MelLec deficiency was protective, with MelLec-/- animals showing normal weight gain and significantly reduced pulmonary inflammation in our allergic model. The lungs of treated MelLec-/- mice displayed significantly reduced inflammatory foci and reduced bronchial wall thickening, which correlated with a reduced cellular influx (particularly neutrophils and inflammatory monocytes) and levels of inflammatory cytokines and chemokines. Notably, fungal burdens were increased in MelLec-/- animals, without apparent adverse effects, and there were no alterations in the survival of these mice. Characterization of the pulmonary T-cell populations, revealed a significant reduction in Th17 cells, and no alterations in Th2, Th1 or Treg cells. Thus, our data reveal that while MelLec is required to control pulmonary fungal burden, the inflammatory responses mediated by this receptor negatively impact the animal welfare in this allergic model.

Correction.

British Journal of PharmacologyVolume 178, Issue 10 p. 2161-2161 CORRECTIONFree Access Correction First published: 01 May 2021 https://doi.org/10.1111/bph.15449AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat In the paper “The anti-parasitic drug miltefosine suppresses human eosinophil activation and ameliorates murine allergic inflammation in vivo” by Knuplez et al, the title has been amended to “The anti-parasitic drug miltefosine suppresses activation of human eosinophils and ameliorates allergic inflammation in mice.” REFERENCE Knuplez, E., Kienzl, M., Trakaki, A., Schicho, R., Heinemann, A., Sturm, E. M., & Marsche, G. (2021). The anti-parasitic drug miltefosine suppresses activation of human eosinophils and ameliorates allergic inflammation in mice. British Journal of Pharmacology, 178, 1234– 1248. Wiley Online LibraryCASPubMedWeb of Science®Google Scholar Volume178, Issue10Themed Issue: Cellular metabolism and diseasesMay 2021Pages 2161-2161 ReferencesRelatedInformation

Selective deletion of SHIP-1 in hematopoietic cells in mice leads to severe lung inflammation involving ILC2 cells

Src homology 2 domain–containing inositol 5-phosphatase 1 (SHIP-1) regulates the intracellular levels of phosphotidylinositol-3, 4, 5-trisphosphate, a phosphoinositide 3–kinase (PI3K) product. Emerging evidence suggests that the PI3K pathway is involved in allergic inflammation in the lung. Germline or induced whole-body deletion of SHIP-1 in mice led to spontaneous type 2-dominated pulmonary inflammation, demonstrating that SHIP-1 is essential for lung homeostasis. However, the mechanisms by which SHIP-1 regulates lung inflammation and the responsible cell types are still unclear. Deletion of SHIP-1 selectively in B cells, T cells, dendritic cells (DC) or macrophages did not lead to spontaneous allergic inflammation in mice, suggesting that innate immune cells, particularly group 2 innate lymphoid cells (ILC2 cells) may play an important role in this process. We tested this idea using mice with deletion of SHIP-1 in the hematopoietic cell lineage and examined the changes in ILC2 cells. Conditional deletion of SHIP-1 in hematopoietic cells in Tek-Cre/SHIP-1 mice resulted in spontaneous pulmonary inflammation with features of type 2 immune responses and airway remodeling like those seen in mice with global deletion of SHIP-1. Furthermore, when compared to wild-type control mice, Tek-Cre/SHIP-1 mice displayed a significant increase in the number of IL-5/IL-13 producing ILC2 cells in the lung at baseline and after stimulation by allergen Papain. These findings provide some hints that PI3K signaling may play a role in ILC2 cell development at baseline and in response to allergen stimulation. SHIP-1 is required for maintaining lung homeostasis potentially by restraining ILC2 cells and type 2 inflammation.

Sapropterin (BH4) Aggravates Autoimmune Encephalomyelitis in Mice

Depletion of the enzyme cofactor, tetrahydrobiopterin (BH4), in T-cells was shown to prevent their proliferation upon receptor stimulation in models of allergic inflammation in mice, suggesting that BH4 drives autoimmunity. Hence, the clinically available BH4 drug (sapropterin) might increase the risk of autoimmune diseases. The present study assessed the implications for multiple sclerosis (MS) as an exemplary CNS autoimmune disease. Plasma levels of biopterin were persistently low in MS patients and tended to be lower with high Expanded Disability Status Scale (EDSS). Instead, the bypass product, neopterin, was increased. The deregulation suggested that BH4 replenishment might further drive the immune response or beneficially restore the BH4 balances. To answer this question, mice were treated with sapropterin in immunization-evoked autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. Sapropterin-treated mice had higher EAE disease scores associated with higher numbers of T-cells infiltrating the spinal cord, but normal T-cell subpopulations in spleen and blood. Mechanistically, sapropterin treatment was associated with increased plasma levels of long-chain ceramides and low levels of the poly-unsaturated fatty acid, linolenic acid (FA18:3). These lipid changes are known to contribute to disruptions of the blood–brain barrier in EAE mice. Indeed, RNA data analyses revealed upregulations of genes involved in ceramide synthesis in brain endothelial cells of EAE mice (LASS6/CERS6, LASS3/CERS3, UGCG, ELOVL6, and ELOVL4). The results support the view that BH4 fortifies autoimmune CNS disease, mechanistically involving lipid deregulations that are known to contribute to the EAE pathology.

The anti-parasitic drug miltefosine suppresses activation of human eosinophils and ameliorates allergic inflammation in mice.

Background and PurposeMiltefosine is an alkylphosphocholine drug with proven effectiveness against various types of parasites and cancer cells. Miltefosine is not only able to induce direct parasite killing but also modulates host immunity, for example by reducing the severity of allergies in patients. To date, there are no reports on the effect of miltefosine on eosinophils, central effector cells involved in allergic inflammation.Experimental ApproachWe tested the effect of miltefosine on the activation of human eosinophils and their effector responses in vitro and in mouse models of eosinophilic migration and ovalbumin‐induced allergic lung inflammation.Key ResultsThe addition of miltefosine suppressed several eosinophilic effector reactions such as CD11b up‐regulation, degranulation, chemotaxis and downstream signalling. Miltefosine significantly reduced the infiltration of immune cells into the respiratory tract of mice in an allergic cell recruitment model. Finally, in a model of allergic inflammation, treatment with miltefosine resulted in an improvement of lung function parameters.Conclusion and ImplicationsOur observations suggest a strong modulatory activity of miltefosine in the regulation of eosinophilic inflammation in vitro and in vivo. Our data underline the potential efficacy of miltefosine in the treatment of allergic diseases and other eosinophil‐associated disorders and may raise important questions regarding the immunomodulatory effect of miltefosine in patients treated for leishmania infections.

Propolis suppresses cytokine production in activated basophils and basophil-mediated skin and intestinal allergic inflammation in mice

BackgroundPropolis is a resinous mixture produced by honey bees that contains cinnamic acid derivatives and flavonoids. Although propolis has been reported to inhibit mast cell functions and mast cell-dependent allergic responses, the effect of propolis on basophil biology remains unknown. This study aimed to investigate the inhibitory effect of propolis on FcεRI-mediated basophil activation. MethodsTo determine the inhibitory effect of propolis on basophil activation in vitro, cytokine production and FcεRI signal transduction were analyzed by ELISA and western blotting, respectively. To investigate the inhibitory effect of propolis in vivo, IgE-CAI and a food allergy mouse model were employed. ResultsPropolis treatment resulted in the suppression of IgE/antigen-induced production of IL-4, IL-6 and IL-13 in basophils. Phosphorylation of FcεRI signaling molecules Lyn, Akt and ERK was inhibited in basophils treated with propolis. While propolis did not affect the basophil population in the treated mice, propolis did inhibit IgE-CAI. Finally, ovalbumin-induced intestinal anaphylaxis, which involves basophils and basophil-derived IL-4, was attenuated in mice prophylactically treated with propolis. ConclusionsTaken together, these results demonstrate the ability of propolis to suppress IgE-dependent basophil activation and basophil-dependent allergic inflammation. Therefore, prophylactic treatment with propolis may be useful for protection against food allergic reactions in sensitive individuals.

Lysophosphatidic Acid Regulates the Differentiation of Th2 Cells and Its Antagonist Suppresses Allergic Airway Inflammation

Background: Lysophosphatidic acid (LPA), a prototypic member of a large family of lysophospholipids, has been recently shown to play a role in immune responses to respiratory diseases. The involvement of LPA in allergic airway inflammation has been reported, but the mechanism remains unclear. Object: We analyzed the biological activity of LPA in vitro and in vivo and investigated its role in allergic inflammation in mice using an LPA receptor 2 (LPA2) antagonist. Methods: We used a murine model with acute allergic inflammation, in which mice are sensitized and challenged with house dust mite, and analyzed airway hyperresponsiveness (AHR), pathological findings, Th2 cytokines, and IL-33 in bronchoalveolar lavage fluid (BALF) and lung homogenates. The effect of LPA on Th2 differentiation and cytokine production was examined in vitro using naive CD4⁺ T cells isolated from splenocytes. We also investigated in vivo the effects of LPA on intranasal administration in mice. Results: The LPA2 antagonist suppressed the increase of AHR, the number of total cells, and eosinophils in BALF and lung tissue. It also decreased the production of IL-13 in BALF and IL-33 and CCL2 in the lung. LPA promoted Th2 cell differentiation and IL-13 production by Th2 cells in vitro. Nasal administration of LPA significantly increased the number of total cells and IL-13 in BALF via regulating the production of IL-33 and CCL-2-derived infiltrating macrophages. Conclusion: These findings suggest that LPA plays an important role in allergic airway inflammation and that the blockade of LPA2 might have therapeutic potential for bronchial asthma.

Microneedle-Mediated Allergen-Specific Immunotherapy for the Treatment of Airway Allergy in Mice.

Subcutaneous allergen-specific immunotherapy (SCIT) qualifies as a promising approach for the permanent cure of IgE-mediated airway allergies, which can often manifest into allergic rhinitis and other allergic respiratory diseases. SCIT entails repeated administration of a high allergen dose into the subcutaneous (sc) region using a hypodermic needle for many (3-5) years, which is inconvenient and painful and reduces patient compliance. To overcome these limitations, we hypothesized that microneedles (MNs), which are minimally invasive and painless, could provide a novel approach for allergen desensitization by depositing the allergen into the superficial layers of the skin. To test this hypothesis, we compared MNs and SCIT for allergen desensitization in a mouse model of ovalbumin (Ova)-induced airway allergy. Mice were first made allergic to Ova and then treated with MNs coated with Ova (with or without CpG as an adjuvant) or via SCIT-Ova + alum (subcutaneous Ova + alum injections) for comparison. Treatment with coated MNs significantly induced Ova-specific serum IgG antibodies in a manner comparable to SCIT-Ova + alum-treated group. To test the efficacy against allergen challenge, treated mice were challenged with Ova via the nasal route. Coated MNs with Ova and CpG (MN-Ova + CpG) considerably suppressed the airway inflammation in allergic mice, evidenced by downregulation of proinflammatory cytokines (IL-5 and IL-13), upregulation of anti-inflammatory cytokine IL-10, and activation of Ova-specific immune response in bronchoalveolar (BAL) fluid. The therapeutic capacity of MN-based allergy treatment was further validated by the reduction in eosinophil and mast cell infiltration in the lung tissues of mice treated with MN-Ova + CpG, and low deposition of mucus inside their lung bronchioles. Overall, coated MNs ameliorated the symptoms of airway allergy in mice similar to SCIT and could provide a novel means of painless allergen-specific immunotherapy.

Defining mechanisms of semaphorin 4A stimulatory action on human Treg cells

Abstract We had shown previously that semaphorin 4A (Sema4A) suppresses allergic inflammation in mice (Mucosal Immunol., 2012, 5:409-19), regulates human Treg cell stability, and induces the generation of Treg cells in PBMC cultures acting through its Plexin B1 receptor (Immunohorizons, 2019, 3:78–87). Plexin B1 forms functional homodimers or heterodimers with other Plexin B molecules and co-precipitates with Neuropilins (NRPs) in transfected 293T cells. Furthermore, NRPs were shown to stabilize Sema/Plexin interactions. In this study, we sought to identify potential partner(s) for Plexin B1 in mediating Sema4A action. We examined surface expression of Plexin B2, B3, and NRPs 1, 2 in human Treg cells by flow cytometry, and observed that they express NRP-2 but not NRP-1, Plexin B2, or B3. Moreover, culture of PBMC with recombinant human Sema4A-Fc (rhSema4A) in vitro resulted in the downregulation of NRP-2 expression on human Treg cells suggesting a specific ligand-receptor interaction. We propose that Sema4A action on human Treg cells may be mediated by a Plexin B1-NRP-2 complex. Our ongoing research is focused on cryoEM studies of ligand-receptor and receptor-receptor interactions in conjunction with sequence/structure statistical analyses using the DARC (Deep Analysis of Residue Constraints) program (Scientific Reports, in press) with the goal of developing Sema4A-based therapies for allergic disease.

Aerosolized Toll-like Receptor Agonists Suppress Allergic Inflammation

Abstract Recent studies have linked exposure to microbes and a reduction of allergic diseases. We sought to determine whether activation of innate immunity by aerosolized Toll-like receptor (TLR) agonists could attenuate the development of allergic inflammation in mice. BALB/cJ mice were sensitized and challenged with House Dust Mite (HDM). Some mice were treated by aerosol with ligands for TLR9 (ODNm362) and TLR2/6 (Pam2CSK4) in combination (Pam2-ODN). Allergic inflammation was assessed by quantification of leukocytes in bronchoalveolar lavage fluid (BALF) and epithelial mucin content with periodic acid fluorescent Schiff stain (PAFS). Pam2-ODN-treated mice exhibited a 94% reduction in eosinophils, 90% reduction in lymphocytes, and a 44% reduction in epithelial mucin content. Lymphocyte subset analysis showed a 50% reduction in TH2 cells without a detectable change in any other TH subset. Using an alternate allergic inflammation model with Ovalbumin (OVA) Pam2-ODN-treated mice showed greater than 50% reductions in total and OVA-specific IgE serum concentrations. IgG2a concentrations, which are reduced in OVA-sensitized mice, returned to baseline with Pam2-ODN treatment. There is a stark contrast of the efficacy in mice treated before and after sensitization that suggests Pam2-ODN may modulate the tendency of sensitization when exposed to aeroallergens. Previous studies have shown that lung epithelial cells are required for Pam2-ODN lung responses, and taken together with these new data, Pam2-ODN may reprogram airway epithelial cells to be tolerogenic to aeroallergens, but additional investigation is required to understand the precise molecular mechanism of Pam2-ODN in this context.

Therapeutic DNA vaccine attenuates itching and allergic inflammation in mice with established biting midge allergy.

Forcipomyia taiwana is a tiny hematophagous midge that attacks en masse. It is responsible for the most prevalent biting insect allergy in Taiwan. For t 2 is its major allergen. The intense itchy reactions can prevent allergic individuals from performing their regular daily outdoor activities. This study aimed to investigate whether the For t 2 DNA vaccine was effective in treating mice with established biting midge allergy. Mice were sensitized with recombinant For t 2 proteins or whole midge extracts. Two to four consecutive shots of various concentrations of For t 2 DNA vaccine, with or without CpG adjuvants, were then administered to midge-sensitized mice. Mice that received two shots of 50-100 μg For t 2 DNA vaccine showed a significant reduction in allergen-induced bouts of scratching, For t 2-specific IgE, specific IgG1/IgG2a ratio in sera, skin eosinophil infiltration, and IL-31 production, as well as IL-4 and IL-13 production by splenocytes. Two doses of For t 2 DNA vaccine one week apart was sufficient to treat mice with established biting midge allergy. The treatment resulted in clinical, immunological, and histopathological improvements. We recommend that this low-cost, convenient treatment strategy be developed for use in humans who are allergic to biting midges.

CARsomes inhibit airway allergic inflammation in mice by inducing antigen-specific Th2 cell apoptosis.

Skewed T helper (Th)2 response plays a crucial role in the pathogenesis of allergic diseases. The therapeutic efficacy for allergic diseases is unsatisfactory currently. This study aims to regulate the skewed Th2 response with CARsomes. The CARsome consisted of an epitope of Dermatophagoides farina-1 (Derf1), a segment of the anti-DEC205 antibody, the scFv, and an open reading frame of perforin. This fusion protein binds to DEC205 molecule on the surface of exosomes derived from dendritic cells (DC). The effects of CARsome on inducing antigen (Ag)-specific Th2 cell apoptosis were assessed both in vivo and in vitro. Exposure to CARsomes in the culture induced Ag-specific Th2 cell apoptosis. Injection of CARsomes through the vein puncture also induced Ag-specific Th2 cell apoptosis in the lungs of sensitized mice. CARsomes could induce Ag-specific regulatory T cells. Administration of CARsomes efficiently inhibited experimental allergic airway inflammation. The CARsomes can inhibit allergic airway inflammation by inducing Ag-specific Th2 cell apoptosis and induce Ag-specific regulatory T cells. The data suggest that CARsomes have the translational potential to be used to treat allergic airway inflammation.

Phosphatidylinositol 3-kinase-δ controls endoplasmic reticulum membrane fluidity and permeability in fungus-induced allergic inflammation in mice.

Background and PurposePhosphatidylinositol 3‐kinase (PI3K), especially PI3K‐δ, and endoplasmic reticulum (ER) stress play important roles in refractory asthma induced by the fungus Aspergillus fumigatus through mechanisms that are not well understood. Here we have investigated these mechanisms, using BEAS‐2B human bronchial epithelial cells and a mouse model of A. fumigatus‐induced allergic lung inflammation.Experimental ApproachA selective PI3K‐δ inhibitor, IC87114, and an ER folding chaperone, 4‐phenylbutyric acid (4‐PBA), were applied to a model of A. fumigatus‐induced asthma in female C57BL/6 mice. The therapeutic potential of IC87114 and 4‐PBA was assessed in relevant primary cell, tissue, and disease models, using immunohistochemistry, western blotting and assessment of ER redox state and membrane fluidity.Key ResultsTreatment with IC87114 or 4‐PBA alleviated pulmonary inflammation and airway remodelling and reduced ER stress and inflammation‐associated intra‐ER hyperoxidation, disrupting protein disulfide isomerase (PDI) chaperone activity. IC87114 and 4‐PBA also reversed changes in ER membrane fluidity and permeability and the resultant mitochondrial hyperactivation (i.e., Ca2+ accumulation) under hyperoxidation, thereby restoring the physiological state of the ER and mitochondria. These compounds also abolished mitochondria‐associated ER membrane (MAM) formation caused by the physical contact between these subcellular organelles.Conclusion and ImplicationsPI3K‐δ and ER stress mediate A. fumigatus‐induced allergic lung inflammation by altering the ER redox state, PDI chaperone function, and ER membrane fluidity and permeability and by amplifying ER signalling to mitochondria through MAM formation. Thus, therapeutic strategies that target the PI3K‐δ–ER stress axis could be an effective treatment for allergic asthma caused by fungi.