Ovalbumin-induced Airway Inflammation Research Articles

IgE cross-linking induces activation of human and mouse mast cell progenitors

The concept of innate and adaptive effector cells that are repleted by maturing inert progenitor cell populations is changing. Mast cells develop from rare mast cell progenitors populating peripheral tissues at homeostatic conditions, or as a result of induced recruitment during inflammatory conditions. Because FcεRI-expressing mast cell progenitors are the dominating mast cell type during acute allergic lung inflammation invivo, we hypothesized that they are activated by IgE cross-linking. Mouse peritoneal and human peripheral blood cells were sensitized and stimulated with antigen, or stimulated with anti-IgE, and the mast cell progenitor population analyzed for signs of activation by flow cytometry. Isolated peritoneal mast cell progenitors were studied before and after anti-IgE stimulation at single-cell level by time-lapse fluorescence microscopy. Lung mast cell progenitors were analyzed for their ability to produce IL-13 by intracellular flow cytometry in a mouse model of ovalbumin-induced allergic airway inflammation. Sensitized mouse peritoneal mast cell progenitors demonstrate increased levels of phosphorylation of tyrosines on intracellular proteins (total tyrosine phosphorylation), and spleen tyrosine kinase (Syk) phosphorylation after antigen exposure. Anti-IgE induced cell surface-associated lysomal-associated membrane protein-1 (LAMP-1) in naive mast cell progenitors, and prompted loss of fluorescence signal and altered morphology of isolated cells loaded with lysotracker. In human mast cell progenitors, anti-IgE increased total tyrosine phosphorylation, cell surface-associated LAMP-1, and CD63. Lung mast cell progenitors from mice with ovalbumin-induced allergic airway inflammation produce IL-13. Mast cell progenitors become activated by IgE cross-linking and may contribute to the pathology associated with acute allergic airway inflammation.

Vitamin D Supplementation: Oxidative Stress Modulation in a Mouse Model of Ovalbumin-Induced Acute Asthmatic Airway Inflammation.

Asthma oxidative stress disturbances seem to enable supplementary proinflammatory pathways, thus contributing to disease development and severity. The current study analyzed the impact of two types of oral vitamin D (VD) supplementation regimens on the redox balance using a murine model of acute ovalbumin-induced (OVA-induced) asthmatic inflammation. The experimental prevention group received a long-term daily dose of 50 µg/kg (total dose of 1300 µg/kg), whereas the rescue group underwent a short-term daily dose of 100 µg/kg (total dose of 400 µg/kg). The following oxidative stress parameters were analyzed in serum, bronchoalveolar lavage fluid (BALF) and lung tissue homogenate (LTH): total oxidative status, total antioxidant response, oxidative stress index, malondialdehyde and total thiols. Results showed that VD significantly reduced oxidative forces and increased the antioxidant capacity in the serum and LTH of treated mice. There was no statistically significant difference between the two types of VD supplementation. VD also exhibited an anti-inflammatory effect in all treated mice, reducing nitric oxide formation in serum and the expression of nuclear factor kappa B p65 in the lung. In conclusion, VD supplementation seems to exhibit a protective role in oxidative stress processes related to OVA-induced acute airway inflammation.

Atractylodin attenuates the expression of MUC5AC and extracellular matrix in lipopolysaccharide-induced airway inflammation by inhibiting the NF-κB pathway.

This study aimed to explore the effects of atractylodin (ATR) on lipopolysaccharide (LPS)-induced inflammatory response in human airway epithelial cells. The cytotoxicity was assessed by CCK-8 assay. The mRNA expression and concentration of interleukin (IL)-6, IL-8, and mucin 5AC (MUC5AC) were measured by qRT-PCR and ELISA, respectively. Western blotting was performed to determine protein expression. We found that LPS stimulation increased the mRNA expression and concentrations of IL-6, IL-8, and MUC5AC, as well as the expression of Col-I and FN in 16HBE cells, but this effect of LPS was attenuated by ATR treatment. Mechanistically, ATR suppressed LPS-induced activation of the NF-κB pathway in 16HBE cells. Moreover, ATR repressed ovalbumin-induced airway inflammation and NF-kB pathway in mice. In conclusion, ATR attenuated the expression of MUC5AC and ECM in LPS-induced airway inflammation by inhibiting the NF-κB pathway.

Ovalbumin-Induced Airway Inflammation Is Ameliorated in Dectin-1-Deficient Mice, in Which Pulmonary Regulatory T Cells Are Expanded through Modification of Intestinal Commensal Bacteria.

Asthma is an allergic chronic respiratory disease that affects more than 300 million people around the world. Dysbiosis of intestinal commensal microbiota influences the development of asthma. Dectin-1 (gene symbol: Clec7a), a C-type lectin receptor, plays an important role in the intestinal immune homeostasis by controlling regulatory T (Treg) cell differentiation through regulation of intestinal microbiota. However, it is not clear whether intestinal immune conditions affect immune responses in other organs. In this study, we examined the effects of Dectin-1 deficiency on allergic airway inflammation (AAI). OVA-induced AAI was attenuated in Clec7a -/- mice. Treg cells were more abundant in colonic lamina propria, mesenteric lymph nodes, and bronchoalveolar lavage fluid of Clec7a -/- mice after AAI induction. Treatment with antibiotics, but not an antifungal agent, decreased the abundance of intestinal Treg cells and aggravated the symptoms of AAI in Clec7a -/- mice. Transplantation of gut microbiota from Clec7a -/- mice into antibiotic-treated hosts increased the abundance of intestinal Treg cells and ameliorated AAI. Overcolonization by Lactobacillus murinus, a Dectin-1 signaling-regulated commensal bacterium, also promoted expansion of Treg cells in the colon and suppressed lung inflammation. Depletion of Treg cells with anti-CD25 Ab eliminated the phenotypic differences between wild-type and Clec7a -/- mice in OVA-induced AAI. These observations suggest that inhibition of Dectin-1 signaling ameliorates AAI by increasing the abundance of Treg cells in lungs through modification of intestinal commensal bacteria, suggesting a role for commensal microbiota in regulating inflammation in organs other than the intestine.

Cryptotympana pustulata Extract and Its Main Active Component, Oleic Acid, Inhibit Ovalbumin-Induced Allergic Airway Inflammation through Inhibition of Th2/GATA-3 and Interleukin-17/RORγt Signaling Pathways in Asthmatic Mice.

Cicadae Periostracum (CP), derived from the slough of Cryptotympana pustulata, has been used as traditional medicine in Korea and China because of its diaphoretic, antipyretic, anti-inflammatory, antioxidant, and antianaphylactic activities. The major bioactive compounds include oleic acid (OA), palmitic acid, and linoleic acid. However, the precise therapeutic mechanisms underlying its action in asthma remain unclear. The objective of this study was to determine the antiasthmatic effects of CP in an ovalbumin (OVA)-induced asthmatic mouse model. CP and OA inhibited the inflammatory cell infiltration, airway hyperresponsiveness (AHR), and production of interleukin (IL)7 and Th2 cytokines (IL-5) in the bronchoalveolar lavage fluid and OVA-specific imunoglobin E (IgE) in the serum. The gene expression of IL-5, IL-13, CCR3, MUC5AC, and COX-2 was attenuated in lung tissues. CP and OA might inhibit the nuclear translocation of GATA-binding protein 3 (GATA-3) and retinoic acid receptor-related orphan receptor γt (RORγt) via the upregulation of forkhead box p3 (Foxp3), thereby preventing the activation of GATA-3 and RORγt. In the in vitro experiment, a similar result was observed for Th2 and GATA-3. These results suggest that CP has the potential for the treatment of asthma via the inhibition of the GATA-3/Th2 and IL-17/RORγt signaling pathways.

Requirement of Gαi1 and Gαi3 in interleukin-4-induced signaling, macrophage M2 polarization and allergic asthma response.

IL-4 induces Akt activation in macrophages, required for full M2 (alternative) polarization. We examined the roles of Gαi1 and Gαi3 in M2 polarization using multiple genetic methods.Methods and Results: In MEFs and primary murine BMDMs, Gαi1/3 shRNA, knockout or dominant negative mutations attenuated IL-4-induced IL4Rα endocytosis, Gab1 recruitment as well as Akt activation, leaving STAT6 signaling unaffected. Following IL-4 stimulation, Gαi1/3 proteins associated with the intracellular domain of IL-4Rα and the APPL1 adaptor, to mediate IL-4Rα endosomal traffic and Gab1-Akt activation in BMDMs. In contrast, gene silencing of Gαi1/3 with shRNA or knockout resulted in BMDMs that were refractory to IL-4-induced M2 polarization. Conversely, Gαi1/3-overexpressed BMDMs displayed preferred M2 response with IL-4 stimulation. In primary human macrophages IL-4-induced Akt activation and Th2 genes expression were inhibited with Gαi1/3 silencing, but augmented with Gαi1/3 overexpression. In Gαi1/3 double knockout (DKO) mice, M2 polarization, by injection of IL-4 complex or chitin, was potently inhibited. Moreover, in a murine model of asthma, ovalbumin-induced airway inflammation and hyperresponsiveness were largely impaired in Gαi1/3 DKO mice.Conclusion: These findings highlight novel and essential roles for Gαi1/3 in regulating IL-4-induced signaling, macrophage M2 polarization and allergic asthma response.

Solid lipid nanoparticle delivery of rhynchophylline enhanced the efficiency of allergic asthma treatment via the upregulation of suppressor of cytokine signaling 1 by repressing the p38 signaling pathway

ABSTRACT Allergic asthma is one of the most common chronic airway diseases, and there is still a lack of effective drugs for the treatment of allergic asthma. The purpose of this work is to formulate rhynchophylline (Rhy)-solid lipid nanoparticles (SLNs) to improve their therapeutic efficacy in a mice allergic model of asthma. A solvent injection method was employed to prepare the Rhy-SLNs. Physicochemical characterization of Rhy-SLNs was measured, and the release assessment was investigated, followed by the release kinetics. Next, a model of murine experimental asthma was established. Mice were subcutaneously injected with 20 μg ovalbumin mixed with 1 mg aluminum hydroxide on days 0, 14, 28, and 42 and administrated aerosolized 1% ovalbumin (w/v) by inhalation from day 21 to day 42. Mice were intraperitoneally injected with 20 mg/kg Rhy-SLNs or Rhy at one hour before the airway challenge with ovalbumin. The results showed that Rhy-SLNs revealed a mean particle size of 62.06 ± 1.62 nm with a zeta potential value of −6.53 ± 0.04 mV and 82.6 ± 1.8% drug entrapment efficiency. The release curve of Rhy-SLNs was much higher than the drug released in phosphate buffer saline at 0, 1, 1.5, 2, 4, or 6 h. Moreover, Rhy-SLNs exerted better effects on inhibiting ovalbumin-induced airway inflammation, oxidative stress, airway remodeling (including collagen deposition and mucus gland hyperplasia) than Rhy in murine experimental asthma. Subsequently, we found that Rhy-SLNs relieved allergic asthma via the upregulation of the suppressor of cytokine signaling 1 by repressing the p38 signaling pathway.

Critical role of IL-33, but not IL-25 or TSLP, in silica crystal-mediated exacerbation of allergic airway eosinophilia

Silica crystals (silica), which are a major mineral component of volcanic ash and desert dust, contribute to the pathogenesis of pulmonary disorders such as asthma and fibrosis. Although administration of silica or sand dust to rodents exacerbates development of ovalbumin-induced or house dust mite-induced asthma-like airway inflammation, the detailed mechanisms remain unclear. Here, using murine models, we found that silica can induce IL-33 expression in pulmonary epithelial cells. IL-33, but not IL-25 or TSLP, and type 2 cytokines such as IL-5 and IL-13 were critically involved in silica’s exacerbation of OVA-induced airway eosinophilia in mice. Innate lymphoid cells (ILCs), but not T, B or NKT cells, were also involved in the setting. Moreover, a scavenger receptor that recognized silica was important for silica’s exacerbating effect. These observations suggest that IL-33 induced in epithelial cells by silica activates ILCs to produce IL-5 and/or IL-13, contributing to silica’s exacerbation of OVA-induced airway eosinophilia in mice. Our findings provide new insight into the underlying mechanisms of exacerbation of pulmonary disorders such as asthma following inhalation of silica-containing materials such as volcanic ash and desert dust.

TRICHINELLA SPIRALIS INFECTION PROTECTS AGAINST OVALBUMININDUCED ALLERGIC BRONCHIAL ASTHMA IN A MURINE MODEL

Allergic bronchial asthma is a long-lasting illness of the respiratory system characterized bychronic inflammation of the respiratory airways with abnormal Th2-type immune responses tospecific allergens. The accumulation of scientific evidence supports the beneficial effects ofsome parasitic infections in animal models of allergic diseases. Thus, the aim of this work wasto discover the influence of infection with T. spiralis on ovalbumin-induced acute allergic airwayinflammation in mice and to investigate whether this effect is correlated to the infectiondose or not. Therefore, the severity of respiratory airways inflammation, the leukocyte counts inthe blood and BALF, immunohistochemistry of FOXP3+CD4+T cells and the levels of IL-6, IL-13, and IL-10 were assessed. In this experiment, 120 male laboratory bred mice were randomlydivided into six groups: induced asthma, saline control, low dose preventive, high dose preventive,low dose therapeutic and high dose therapeutic groups. T. spiralis infection attenuated theintensity of pulmonary inflammation, decreased numbers of eosinophils in BALF and blood,reduced levels of IL-6 and IL-13 and elevated levels of IL-10 with a significant upregulation ofFOXP3+CD4+T cells expression compared to the uninfected induced asthma group. The preventivegroups showed the best results with no significant difference between effects of the low andthe high doses. In conclusion, T. spiralis infection reduced Th2 type infammation and augmentedregulatory immune response in ovalbumin-induced acute allergic airway inflammation withbetter preventive than therapeutic effects. Further investigations are needed to determine theminimal dose of infection that gives the best influence.

TRICHINELLA SPIRALIS INFECTION PROTECTS AGAINST OVALBUMININDUCED ALLERGIC BRONCHIAL ASTHMA IN A MURINE MODEL

Allergic bronchial asthma is a long-lasting illness of the respiratory system characterized bychronic inflammation of the respiratory airways with abnormal Th2-type immune responses tospecific allergens. The accumulation of scientific evidence supports the beneficial effects ofsome parasitic infections in animal models of allergic diseases. Thus, the aim of this work wasto discover the influence of infection with T. spiralis on ovalbumin-induced acute allergic airwayinflammation in mice and to investigate whether this effect is correlated to the infectiondose or not. Therefore, the severity of respiratory airways inflammation, the leukocyte counts inthe blood and BALF, immunohistochemistry of FOXP3+CD4+T cells and the levels of IL-6, IL-13, and IL-10 were assessed. In this experiment, 120 male laboratory bred mice were randomlydivided into six groups: induced asthma, saline control, low dose preventive, high dose preventive,low dose therapeutic and high dose therapeutic groups. T. spiralis infection attenuated theintensity of pulmonary inflammation, decreased numbers of eosinophils in BALF and blood,reduced levels of IL-6 and IL-13 and elevated levels of IL-10 with a significant upregulation ofFOXP3+CD4+T cells expression compared to the uninfected induced asthma group. The preventivegroups showed the best results with no significant difference between effects of the low andthe high doses. In conclusion, T. spiralis infection reduced Th2 type infammation and augmentedregulatory immune response in ovalbumin-induced acute allergic airway inflammation withbetter preventive than therapeutic effects. Further investigations are needed to determine theminimal dose of infection that gives the best influence.

Lin28B Regulates Angiotensin II-Mediated Let-7c/miR-99a MicroRNA Formation Consequently Affecting Macrophage Polarization and Allergic Inflammation.

Angiotensin-II (Ang-II) receptor plays a role in allergic airway inflammation; however, the underlying mechanism and role of macrophages need better understanding. In the present study, angiotensin-II infusion (1μg/kg/min) in ovalbumin-induced airway inflammation mice model significantly decreased immune cell infiltration, goblet cell hyperplasia, and eosinophil numbers in lungs. Ang-II infusion increased M1 and decreased M2 macrophage population in bronchoalveolar lavage fluid and respective macrophage markers in lung macrophages. Similarly, in vitro Ang-II treatment in murine bone marrow-derived macrophages (BMDMs) induced M1 and reduced M2 macrophage phenotype with enhanced bactericidal activity. Mechanistically, Ang-II inhibits Let-7c and miR-99a expression in BMDMs and in vivo as well. Lentiviral overexpression of Let-7c and miR-99a miRNAs in BMDMs abrogated Ang-II-induced M1 phenotype activation and promoted M2 phenotype, which is governed by targeting TNFα by miR-99a. In lung macrophages, ovalbumin-induced TNFα inhibition was rescued after Ang-II treatment. In BMDMs, knockdown of TNFα abrogated Ang-II-induced M2 to M1 macrophage phenotype switch and associated bactericidal activity. Ang-II affects mature miRNA formation by enhancing Lin28B levels in macrophages in vivo and in vitro. Furthermore, Lin28B knockdown prevented Ang-II-mediated inhibition of mature Let-7c/miR-99a miRNA formation, M2 to M1 macrophage phenotype switch, and increased bactericidal activity. Therefore, present study suggests a role of Lin28B in Ang-II-induced Let-7c/miR-99a miRNA formation that consequently affects TNFα production, M1 phenotype activation, and allergic airway inflammation. Graphical Abstract Ovalbumin inhibits LIN28B expression thereby fails to inhibit premature to mature Let-7c/miR-99a miRNA formation. Mature miR-99a miRNA that inhibits TNFα consequently promotes M2 polarization and allergic airway inflammation. While Ang-II induces Lin28B, which inhibits Let-7c/miR-99a miRNA processing and mature miRNA formation, this results in increased TNFα levels that lead to M1 polarization and allergic airway inflammation inhibition.

Intestinal fungi are causally implicated in microbiome assembly and immune development in mice

The gut microbiome consists of a multi-kingdom microbial community. Whilst the role of bacteria as causal contributors governing host physiological development is well established, the role of fungi remains to be determined. Here, we use germ-free mice colonized with defined species of bacteria, fungi, or both to differentiate the causal role of fungi on microbiome assembly, immune development, susceptibility to colitis, and airway inflammation. Fungal colonization promotes major shifts in bacterial microbiome ecology, and has an independent effect on innate and adaptive immune development in young mice. While exclusive fungal colonization is insufficient to elicit overt dextran sulfate sodium-induced colitis, bacterial and fungal co-colonization increase colonic inflammation. Ovalbumin-induced airway inflammation reveals that bacterial, but not fungal colonization is necessary to decrease airway inflammation, yet fungi selectively promotes macrophage infiltration in the airway. Together, our findings demonstrate a causal role for fungi in microbial ecology and host immune functionality, and therefore prompt the inclusion of fungi in therapeutic approaches aimed at modulating early life microbiomes.

Mammalian Target of Rapamycin Signaling Enhances Ovalbumin-Induced Neutrophilic Airway Inflammation by Promoting Th17 Cell Polarization in Murine Noneosinophilic Asthma Model.

Background: T helper 17 (Th17) is regarded as key immune cell in the pathogenesis of noneosinophilic asthma (NEA) due to the recruitment of neutrophils into the airways. The mammalian target of rapamycin (mTOR) is an important signaling molecule that plays a critical role in immune regulation. This study focused on mTOR signaling pathway in the regulation of Th17-mediated neutrophilic airway inflammation. Methods: Ovalbumin (OVA) T cell receptor transgenic DO11.10 mice (DO11.10 mice) were used to establish NEA model, and few mice received specific mTORC1 inhibitor rapamycin (RAPA) before intranasal administration of OVA. The severity of airway inflammation was determined by differential cell counts in bronchoalveolar lavage (BAL) fluids and histopathologic lung analysis. The levels of various cytokines in BAL fluids and lung tissues were measured. To determine the role of mTORC1 signaling in Th17 differentiation, naive T cells from wild-type (WT) and TSC1 knockout (KO) mice were cultured in Th17 skewing condition with or without RAPA in vitro and the production of IL-17A was compared. Results: Treatment with RAPA markedly attenuated OVA-induced neutrophilic airway inflammation in DO11.10 mice. Also the production of IL-17A was inhibited without affecting the production of interferon-γ (IFN-γ) and IL-4 in lungs. Furthermore, RAPA suppressed differentiation of Th17 cells in vitro, whereas enhanced activity of mTORC1 promoted Th17 cell differentiation and increased the expression of Th17-related transcription factors RORγt and RORα. Conclusion: These results suggested that mTOR promoted Th17 cell polarization and enhanced OVA-induced neutrophilic airway inflammation in experimental NEA.

Combination of Sarsasapogenin and Fluticasone attenuates ovalbumin-induced airway inflammation in a mouse asthma model

Objective: Asthma is a very common airway inflammatory disease for which the existing drug therapy options are insufficient. In this study, we explored the mechanisms underlying the anti-inflammatory potential of Sarsapogenin (SG) and its combination with Fluticasone (FC) in ovalbumin (OVA)-induced allergic asthma in mice.Methods: In a standard experimental model, asthma in mice was sensitized and challenged by OVA. The mice were treated with SG and SG + FC during OVA challenge. At the completion, lung weight, inflammatory cell count in bronchoalveolar lavage fluid (BALF), serum cytokines levels, immunoglobulin E (IgE) levels, lung nitrate/nitrite (NO) levels, and lung tissue oxidative stress biomarkers were determined. Histopathological evaluation of the lung tissue was also performed.Key findings: Treatment of mice with SG and SG + FC combination intensely diminished the trafficking of total and differential inflammatory cells count into BALF. SG and SG + FC administration significantly reduced the production of inflammatory cytokines, serum IgE levels and restoration of antioxidant stress markers. Histopathological analysis of lung samples effectually weakened bronchial inflammation and mucus production in the lung with a significant reduction in inflammation and mucus score.Conclusion: Our study results suggested that SG and SG + FC effectively reduced allergic airway inflammation via inhibiting pro-inflammatory cytokines, NO expressions and oxidative stress parameters. So, it could be used as a therapeutic potential agent for the treatment of asthma by decreasing its dose in combination with FC to avoid the chronic adverse effects of FC.

Preventive effects of "ovalbumin-conjugated celastrol-loaded nanomicelles'' in a mouse model of ovalbumin-induced allergic airway inflammation

Allergies affect a significant proportion of the world's population, and existing vaccination strategies to restrict their adverse pathologies often render side-effects. The aim of this study was to design a new vaccine for allergen-specific immunotherapy (SIT), and to investigate its preventive effects during allergic inflammation. We constructed ovalbumin (OVA)-conjugated celastrol-loaded nanomicelles (OVA-NMs-celastrol), wherein celastrol (a bioactive anti-inflammatory compound) was loaded into carboxyl-functioned polymeric nanomicelles using a thin-film hydration method. OVA was used as a model allergen and conjugated on nanomicelles. The OVA-NMs-celastrol obtained were characterized based on particle size, morphology, drug encapsulation efficiency, and drug loading percentage. Further, the preventive effect of OVA-NMs-celastrol was evaluated in a mouse model of allergic asthma. Our results showed that OVA-NMs-celastrol possessed valuable characteristics such as small particle size (50.72 ± 0.98 nm) and spherical-like shape, with celastrol encapsulation efficiency of 99.89 ± 0.85% and a drug loading percentage of 4.76 ± 0.03%. Further, in vivo results showed that treatment with OVA-NMs-celastrol could decrease OVA specific IgE and histamine levels, Th2 cytokine (IL-4, IL-5) levels, and inflammatory cell infiltration in the lung tissues. Moreover, it could enhance the OVA specific IgG1 and IgG2a levels and decrease the IgE / IgG2a ratio. These results demonstrate the successful construction of OVA-NMs-celastrol as a potential vaccine candidate for use in SIT for allergic inflammation.

Adoptive transfer of type 1 regulatory T cells suppressed the development of airway hyperresponsiveness in ovalbumin-induced airway inflammation model mice

Type 1 regulatory T (Tr1) cells are CD4+ T cells that produce a large amount of IL-10, an anti-inflammatory cytokine. However, it has not been fully elucidated whether Tr1 cells suppress allergic asthma. In this study, the effects of adoptive transfer of in vitro-induced Tr1 cells on allergic asthma were evaluated. Splenocytes from ovalbumin (OVA)-sensitized BALB/c mice were cultured with OVA, IL-21, IL-27, and TGF-β. After culture, IL-10-producing CD4+ T cells were isolated by Dynabeads mouse CD4 and IL-10 secretion assay, and analyzed by flow cytometry. Purified Tr1 cells (IL-10+ CD4+ T cells) were intravenously injected into OVA-sensitized BALB/c mice. The recipient mice were intratracheally challenged with OVA. Airway hyperresponsiveness to methacholine was assessed by the forced oscillation technique, followed by bronchoalveolar lavage (BAL). Almost all of the induced IL-10-producing CD4+ T cells were negative for interferon-γ, IL-4, IL-17A, and forkhead box P3, suggesting that the cells were Tr1 cells. The adoptive transfer of Tr1 cells significantly suppressed the development of airway hyperresponsiveness, and increases in IL-5, eosinophils, and neutrophils in BAL fluid. In conclusion, we demonstrated that Tr1 cells suppressed allergic asthma in mice.

The amino acid sensor general control nonderepressible 2 (GCN2) controls TH9 cells and allergic airway inflammation

TH9 cells have emerged as important mediators of allergic airway inflammation. There is evidence that general control nonderepressible 2 (GCN2) affects the immune response under some stress conditions. However, whether GCN2 regulates CD4+ T-cell differentiation during allergic inflammation remains unknown. We sought to clarify the regulatory roles of GCN2 in CD4+ T-cell subset differentiation and its significance in patients with allergic airway inflammation. The effects of GCN2 in differentiation of TH cell subsets were detected by using the invitro induction system. GCN2 knockout mice, ovalbumin-induced allergic airway inflammation, and adoptive transfer mouse models were used to determine the significance of GCN2 in TH9 differentiation and allergic airway inflammation invivo. RNA sequencing, real-time PCR, Western blotting, and other molecular approaches were used to identify the molecular mechanisms relevant to regulation of GCN2 in TH9 cell differentiation. GCN2 deficiency significantly inhibited differentiation of TH9 cells but not TH1, TH2, and regulatory T cells. GCN2 knockout mice and recombination-activating gene 2 knockout (Rag2KO) mice that received adoptively transferred GCN2-deficient CD4+ T cells exhibited reduced TH9 differentiation and less severe allergic airway inflammation. Furthermore, the isolated GCN2-deficient TH9 cells also mediated less severe allergic airway inflammation on adoptive transfer. Mechanistically, GCN2 deficiency inhibits TH9 cell differentiation through a hypoxia-inducible factor 1α-dependent glycolytic pathway. Our results reveal a novel role of GCN2 in TH9 cell differentiation. Our findings indicate that new strategies to inhibit GCN2 activity might provide novel approaches to attenuate allergic airway inflammation.

KGC3P attenuates ovalbumin-induced airway inflammation through downregulation of p-PTEN in asthmatic mice

BackgroundThe roots of Korean red ginseng (Panax ginseng C.A.Mey.; KGC) have been used as an herbal supplement to enhance vital energy and immune capacity. Salvia plebeia R.Br. has been used to treat inflammatory diseases. PurposeThe aim of this study was to examine the anti-asthmatic effects of a mixture of Korean red ginseng and Salvia plebeia R.Br. (KGC3P), its component nepetin, and their modes of action in alleviating ovalbumin (OVA)-induced asthma in mice. MethodBALB/c mice were sensitized with OVA then subjected to intratracheal, intraperitoneal, and aerosol challenges. KGC3P and nepetin were administered orally for four weeks. Airway hyperresponsiveness (AHR), OVA-specific IgE levels, and Th2 cytokine- and gene expression levels in bronchoalveolar lavage fluid (BALF) and splenocytes were measured. Histological and immune cell subtype analyses were performed. PTEN and Akt phosphorylation levels were also evaluated. ResultsKGC3P reduced OVA-induced AHR, serum IgE levels, histological changes, and eosinophils infiltration but also the absolute number of immune cell subtypes including CD3+/CD4+, CD3+/CD8+, CD4+/CD69+, and Gr-1+/CD11b+ in the lungs, BALF, and mesenteric lymph nodes (MLN). KGC3P also lowered the Th2 cytokines IL-4, IL-5, and IL-13 in the BALF and splenocytes and downregulated the IL-4, IL-13, IL-17, TNF-α, and MUC5AC genes in the lung. KGC3P upregulated the peroxisome proliferator-activated receptor (PPAR)γ gene but downregulated the p-Akt and p-PTEN phosphorylation. Similar results were obtained with nepetin treatment. ConclusionKGC3P and nepetin are anti-asthmatic because they reduce various immune cells such as eosinophils and Th2 cell as well as Th2 cytokines. These mechanisms may be accompanied by the regulation of PPARγ expression and the PTEN pathway. Taken together, our results indicate that KGC3P and nepetin may potentially prevent and treat asthma.

Indoor PM2.5 from coal combustion aggravates ovalbumin-induced asthma-like airway inflammation in BALB/c mice.

We hypothesized that indoor PM2.5 exposure from coal combustion exaggerates airway inflammation in the lung tissue of asthmatic mice induced with ovalbumin (OVA). Forty BALB/c mice, randomly divided into four groups (n = 10 per group), were intratracheally instilled with normal saline alone, PM2.5 (2.5 mg/ml PM2.5 alone), OVA (15 μg/ml OVA alone), and PM2.5+OVA (2.5 mg/ml PM2.5 and 15 μg/ml OVA), respectively, four times at 2-wk intervals. Daily mean concentration of PM2.5 from indoor coal combustion was 156.95 μg/m3. The highest metal composition in PM2.5 was Zn (34.81 ± 1.8 μg/m3). Exposure to PM2.5+OVA significantly elevated IL-4 and decreased IFN-γ production in mice compared with the control (P < 0.05). Exposure to PM2.5+OVA showed a significant increase in the protein levels of granulocyte-macrophage colony-stimulating factor and IL-8 and a decrease in the protein level of transforming growth factor-β1 in bronchoalveolar lavage fluid of mice compared with the control (P < 0.05). The expression of IL-4 mRNA was significantly increased, whereas the expression of IFN-γ mRNA was decreased in lung tissue of the PM2.5+OVA group (P < 0.05). The expression level of Foxp3 mRNA in the PM2.5+OVA group was significantly lower than that in the control group in lung tissue (P < 0.05). Treatment with PM2.5+OVA promoted a prominent neutrophil sequestration into the lung parenchyma, goblet cell proliferation, and severe inflammatory cell infiltration in the airways. Exposure to PM2.5 from indoor coal combustion might induce airway inflammatory immune responses and exacerbate peribronchiolar inflammation due to infiltration of inflammatory cells into the airway submucosa and airway structural pathological changes.

NLRC5 negatively regulates LTA-induced inflammation via TLR2/NF-κB and participates in TLR2-mediated allergic airway inflammation.

NLRC5, the largest member of the Nod-like receptor (NLR) family, has been reported to play a pivotal role in regulating inflammatory responses. Recent evidence suggests that NLRC5 participates in Toll-like receptor (TLR) signaling pathways and negatively modulates nuclear factor-κB (NF-κB) activation. In this study, we investigated the interaction between NLRC5 and TLR2 in the NF-κB inflammatory signaling pathway and the involvement of NLRC5 in TLR2-mediated allergic airway inflammation. We knocked down TLR2 and NLRC5, respectively in the RAW264.7 macrophage cell line by small interfering RNA (siRNA) and then stimulated the knockdown cells with lipoteichoic acid (LTA). In comparison with the negative siRNA group, the level of NLRC5 expression was lower in the TLR2 siRNA group, with a reduction in the NF-κB-related inflammatory response. Conversely, in the NLRC5 knockdown cells, after LTA-treated the level of TLR2 expression did not change but the expression levels of both NF-κB pp65 and NLRP3 increased remarkably. Thus, we hypothesize that NLRC5 participates in the LTA-induced inflammatory signaling pathway and regulates the inflammation via TLR2/NF-κB. Similarly, in subsequent in vivo experiments, we demonstrated that the expression level of NLRC5 was significantly increased in the ovalbumin-induced allergic airway inflammation. However, this effect disappeared in TLR2-deficient (TLR2 -/- ) mice and was accompanied by reduced levels of NF-κB expression and airway inflammation. In conclusion, NLRC5 negatively regulates LTA-induced inflammatory response via a TLR2/NF-κB pathway in macrophages and also participates in TLR2-mediated allergic airway inflammation.