OVA-induced Mouse Model Research Articles

The consistently up-regulated expression of NLRP3 in severe asthma patients from mRNA microarray and ovalbumin-induced mouse model of asthma.

Severe asthma (SA) is a chronic lung disease, resistant to current treatments, symbolized by repeated symptoms of reversible airflow obstruction, airway hyper-responsiveness, and inflammation. The aim of this study was to identify genes exhibiting differential expression in individuals without asthma and SA patients. We aimed to pinpoint hub differentially expressed genes (DEGs) by utilizing a mouse model of asthma sensitized to ovalbumin (OVA). Microarray data for SA were acquired from the Gene Expression Omnibus (GEO) databases. DEGs were identified, and functional enrichment analyses were carried out. STRING and Cytoscape were utilized to design a protein-protein interaction (PPI) network and conduct module analysis. An OVA-induced asthma mice model was established. Lung tissue from the mice was collected for quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blot, and immunohistochemistry (IHC) to assess the expression of DEGs. A total of 545 DEGs were identified, among which 172 genes were upregulated in SA patients compared to healthy controls. The nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) was significantly up-regulated in SA patients [adjusted P value (Padj) =0.001]. Analysis of lung tissue using qRT-PCR, western blot, and IHC revealed higher expression of NLRP3 in OVA-induced asthma mice compared to the control group. Enrichment analysis suggests the involvement of NLRP3 in pathways related to pyroptosis, c-type lectin receptor signaling, and NOD-like receptor signaling. Through bioinformatics analysis, we identified a multitude of DEGs that could potentially contribute significantly to the development of SA. Notably, our findings highlight NLRP3 as a potential pivotal player in asthma pathogenesis, underscoring its prospective utility as a biomarker for SA.

Malvidin From Malva sylvestris L. Ameliorates Allergic Responses in Ovalbumin-Induced Allergic Rhinitis Mouse Model via the STAT6/GATA3 Pathway.

Malva sylvestris L. (commonly known as mallow) has been widely used in traditional Tibetan formulations to treat allergic rhinitis (AR), and malvidin is a key anti-inflammation constituent of this plant. The present study aimed to evaluate the potential therapeutic effect and mechanism of malvidin in an AR mouse model. Malvidin's efficacy was evaluated in an AR mouse model induced by ovalbumin (OVA) sensitization and challenge. The factors, such as nasal symptoms, serum OVA-specific immunoglobulin E (IgE) levels, histological changes in the nasal mucosa, and expressions of Th1, Th2, Th17, and Tregs and their cytokines, were assessed. Western blotting was used to analyze the effect of malvidin on signal transducer and activator of transcription 6 (STAT6) and GATA3 expression levels. Malvidin reduced the allergic symptoms and serum levels of OVA-specific IgE in the AR model. Histological analysis indicated that malvidin alleviates nasal mucosal edema, eosinophil infiltration, and goblet cell proliferation. In addition, it altered the expression of Th1/Th2/Th17-related cytokines, enhanced the Treg population, and reduced Th2-mediated immunity by suppressing the phosphorylation of STAT6 and expression of the GATA3 protein. Malvidin significantly improved allergic symptoms in an OVA-induced AR mouse model by modulating Th1/Th2 immune responses and suppressing the STAT6/GATA3 pathway, indicating its potential as a naturally sourced agent for AR management.

Herbacetin Inhibits Asthma Development by Blocking the SGK1/NF-κB Signaling Pathway.

Asthma severely interferes with people's lives through coughing, wheezing and inflammation of the lungs. Herbacetin is a class of natural compounds that inhibit the development of inflammation. However, whether Herbacetin inhibits asthma has not been definitively studied. Lipopolysaccharides (LPS)-induced lung epithelial (BASE-2B) cells injury model was established, and then the relief of damaged BASE-2B cells with different concentrations of Herbacetin was examined. The cell counting kit (CCK8) was used to detect the effect of Herbacetin on the proliferation ability in ovalbumin (OVA)-induced asthma mice model, and Western Blot and flow cytometry were used to detect the effect of Herbacetin on the apoptosis in OVA-induced asthma mice model. Additionally, pulmonary pathology was detected by HE and Masson staining, and serum inflammatory factors were detected by alveolar lavage fluid. Herbacetin reduces BESA-2B cells induced by LPS level of inflammation, and reactive oxygen species (ROS) generation, inhibits cell apoptosis, promotes cell proliferation, OVA-induced mice lung histopathology test HE staining, serum inflammatory factors show the same results. Western Blot shows that Herbacetin regulates the expression of Caspase-3, Bax, and Bcl-2. SGK1 overexpression increased the rate of apoptosis, and Herbacetin reversed this phenomenon. By silencing the expression of SGK1, it was found that Herbacetin was an inhibitor of SGK1, which could inhibit the NF-κB/p-P65 pathway in asthmatic airway inflammation. Herbacetin reduces pro-inflammatory cytokine levels by inhibiting the SGK1/NF-κB pathway. Our data suggest that Herbacetin has a significant anti-inflammatory effect on asthma and can be used as a potential therapeutic agent.

Dimethyl fumarate alleviates allergic asthma by strengthening the Nrf2 signaling pathway in regulatory T cells.

Allergic asthma is a widely prevalent inflammatory condition affecting people across the globe. T cells and their secretory cytokines are central to the pathogenesis of allergic asthma. Here, we have evaluated the anti-inflammatory impact of dimethyl fumarate (DMF) in allergic asthma with more focus on determining its effect on T cell responses in allergic asthma. By utilizing the ovalbumin (OVA)-induced allergic asthma model, we observed that DMF administration reduced the allergic asthma symptoms and IgE levels in the OVA-induced mice model. Histopathological analysis showed that DMF treatment in an OVA-induced animal model eased the inflammation in the nasal and bronchial tissues, with a particular decrease in the infiltration of immune cells. Additionally, RT-qPCR analysis exhibited that treatment of DMF in an OVA-induced model reduced the expression of inflammatory cytokine (IL4, IL13, and IL17) while augmenting anti-inflammatory IL10 and Foxp3 (forkhead box protein 3). Mechanistically, we found that DMF increased the expression of Foxp3 by exacerbating the expression of nuclear factor E2-related factor 2 (Nrf2), and the in-vitro activation of Foxp3+ Tregs leads to an escalated expression of Nrf2. Notably, CD4-specific Nrf2 deletion intensified the allergic asthma symptoms and reduced the in-vitro iTreg differentiation. Meanwhile, DMF failed to exert protective effects on OVA-induced allergic asthma in CD4-specific Nrf2 knock-out mice. Overall, our study illustrates that DMF enhances Nrf2 signaling in T cells to assist the differentiation of Tregs, which could improve the anti-inflammatory immune response in allergic asthma.

YiQi GuBen capsule alleviates OVA-induced asthma through improving mitochondrial dysfunction

Objective: This study aims to explore the effect of YiQi GuBen capsule on improving mitochondrial dysfunction in an animal model of asthma.Methods: The mice (n = 8) were divided into four groups including control (NC), ovalbumin (OVA), dexamethasone (OVA + DEX), and YiQi GuBen (OVA + YQGB) groups. Firstly, we established an OVA-induced mouse asthma model except for the NC group, which then were treated with dexamethasone and YiQi GuBen capsule. Subsequently, HE staining and Masson staining were used for pathological analysis of mice lung tissues. Next, we used transmission electron microscopy (TEM) to observe the effect of the Yiqi Guben capsule on the ultrastructure of mitochondria. Flow cytometry was used to analyze the ROS level, membrane potential, and the number of mitochondria in lung tissue. Moreover, we analyzed the copy number of mitochondrial DNA (mtDNA) and the expression levels of activator peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) and mitochondrial transcription factor A (TFAM).Results: The results of the pathological analysis showed that after treatment with the YiQi GuBen capsule, the lung tissue damage was significantly reduced. In addition, we observed that the ultrastructural damage of mitochondria was improved. Flow cytometry proved that after treatment with the YiQi GuBen capsule, the level of ROS in the mitochondria was effectively reduced, while the mitochondrial membrane potential decreased and the number increased significantly. Moreover, we found that the copy number of mtDNA was significantly increased and the expression levels of PGC-1α and TFAM were significantly upgraded.Conclusion: This study suggests YiQi GuBen capsule can effectively improve mitochondrial dysfunction in the OVA-induced mouse model.

MiR-410 Regulates Helper T Cell Differentiation in Ovalbumin-Induced Asthma through the PI3K-AKT-VEGF Signaling Pathway

Introduction: Asthma has been attributed to Th1/Th2 imbalance and inappropriate Th2 responses to environmental allergens. MicroRNAs (miRNAs), 21 to 23 RNA molecules, are first found in mammals and have been implicated in various biological activities. Our previous study found that miR-410 effectively ameliorates airway inflammation in the ovalbumin (OVA)-induced asthma murine model. However, the role of miR-410 in regulating helper T (Th) cell differentiation is not clear. In the present study, we aimed to explore the regulatory effects of miR-410 on the differentiation of Th cells through both in vivo and in vitro studies. Methods: Dual-luciferase reporter assay was used to find if miR-410 has any direct binding position with VEGF mRNAs. PBMC and CD4+ T cells were isolated and stimulated with OVA. The miR-410 mimics and inhibitors were transfected into CD4+ T cells. The differentiation of Th cells was evaluated by enzyme-linked immunosorbent assay (ELISA) for the concentration of IL-4, IFN-γ, and TGF-β levels in supernatants. Western Blot was used to detect protein expression and phosphorylation of PI3K and AKT. BALB/c mice were kept in a specific pathogen-free condition and received sterile OVA-free food and water. OVA-induced asthmatic mice model was established. ELISA was used to measure the bronchoalveolar lavage fluid (BALF) concentrations of IL-4, IFN-γ, TGF-β, and VEGF. Hematoxylin and eosin staining and immunohistochemical staining were conducted to analyze inflammatory cell infiltration, pathological changes, and the expression of VEGF. Results: Dual-luciferase reporter assay showed that miR-410 has no direct binding position with VEGF mRNAs. In the OVA-primed mononuclear cells compared to normal cells, IFN-γ and TGF-β were decreased while IL-4 and VEGF were increased. This change was reversed while miRNA-410 mimics were transfected into CD4+ T cells. Besides, the OVA-primed CD4+ T cells treated with miR-410 decrease the proliferation of cytokine of Th2 cells as well as phosphorylation of PI3K, and AKT. In OVA-induced asthma mice, IFN-γ and TGF-β were decreased in BALF while the IL-4 and VEGF were increased. OVA-induced mice with asthma treated with miR-410 mimics showed marked reductions in the infiltration of inflammatory cells as well as IL-4 and VEGF in BALF. The immunohistochemical staining of the expression of VEGF also decreased in OVA-induced asthma mice with the instillation of miR-410. Conclusions: In this study, we revealed that miR-410 could regulate the differentiation of Th cells via the PI3K-AKT-VEGF signaling pathway in asthma.

Leuconostoc mesenteroides WHH1141 ameliorates ovalbumin-induced food allergy in mice.

Food allergy (FA) is acknowledged as a significant public health and food safety issue, due to its manifestation as an amplified immune reaction to food antigens. Recently, probiotics within Lactobacillus and Bifidobacterium have been highlighted as a promising strategy against allergic disease by modulating the balance of Th1/Th2 responses. However, the allergy-alleviating effects of probiotic Leuconostoc mesenteroides strains are unknown. Therefore, this study investigated the potentials of eleven L. mesenteroides strains on the Th1/Th2 balance in vitro by evaluating the expression patterns of interferon-gamma (IFN-γ) (Th1 cytokine) and interleukin-4 (IL-4) (Th2 cytokine) in mesenteric lymph node-derived lymphocytes from ovalbumin (OVA)-sensitized mice. Among strains, WHH1141 incubation caused the highest IFN-γ/IL-4 ratio. Oral administration of WHH1141 (1×109 CFU/mL) in the OVA-induced FA mouse model for 40 days improved the weight loss and FA pathological symptoms and normalized the serum immunoglobulin E levels. Meanwhile, the OVA-induced elevated gene expressions of cytokines (IL-4, IL-5, and IL-13) and tight-junction proteins (ZO-1 and Occludin) and levels of cytokines (IL-4, IL-5, and IL-13) and histamine in the jejunum were restored by WHH1141. Furthermore, WHH1141 reversed the reduced gut microbial diversity and short-chain fatty acid (SCFA) levels, specifically increased Bacteroidota abundance, and decreased Firmicutes abundance in OVA-induced mice. Overall, these findings suggest that WHH1141 exerts FA-alleviating effects on OVA-induced mice, which is involved with the inhibition of the jejunal Th2 immune responses and the modulation of gut microbiome composition and SCFA productions. PRACTICAL APPLICATION: Leuconostoc mesenteroides WHH1141 with FA-alleviating potentials may be considered a promising approach in the mitigation of FA symptoms.

Xiao-qing-long-tang ameliorates OVA-induced allergic rhinitis by inhibiting ILC2s through the IL-33/ST2 and JAK/STAT pathways

BackgroundAllergic rhinitis (AR) is a chronic inflammatory disease of the nasal mucosa that is mediated by immunoglobulin E (IgE). Xiao-qing-long-tang (XQLT) is a traditional Chinese medicine compound that is widely used to treat respiratory diseases such as AR. However, the underlying mechanism of the effect of XQLT on AR remains unclear. PurposeTo elucidate the effect of XQLT on ovalbumin (OVA)-induced AR and the mechanisms of action. MethodsThe therapeutic efficacy of XQLT was evaluated in a well-established OVA-induced AR mouse model. Nasal symptoms were analyzed, type 2 cytokines and OVA-sIgE levels were measured, nasal mucosa tissues were collected for histological analysis, and the changes of Group 2 innate lymphoid cells (ILC2s) and the IL-33/ST2 and JAK/STAT signaling pathways in the nasal mucosa were observed. ResultsXQLT significantly alleviated the nasal symptoms and histological damage to the nasal mucosa in AR mice, and reduced the levels of type 2 cytokines and OVA-sIgE. In addition, after XQLT treatment, the numbers of ILC2s in the nasal mucosa of AR mice were reduced, and the mRNA levels of the transcription factors GATA3 and ROR-α were decreased. Moreover, IL-33/ST2 signaling pathway was inhibited. The costimulatory cytokine associated JAK/STAT signaling pathway was also inhibited in ILC2s. ConclusionOur study demonstrated that XQLT regulated ILC2s through the IL-33/ST2 and JAK/STAT pathways to ameliorate type 2 inflammation in OVA-induced AR. These findings suggest that XQLT might be used to treat AR.

Echinacoside alleviates airway remodeling and inflammation in an ovalbumin-induced neonatal mouse model of asthma by modulating the SIRT1-NF-κB pathway.

Echinacoside (ECH) has been reported to have anti-inflammatory and anti-immune effects, and may be effective for treating asthma. This study aimed to investigate the effect of ECH on asthma. A mouse model of asthma was established by ovalbumin (OVA) induction, and the effect of ECH on airway remodeling in mice was evaluated using the Periodic Acid-Schiff stain and enzyme-linked immunosorbent serologic assay (ELISA). Additionally, the effect of ECH on collagen deposition in asthmatic mice was assessed using Western blotting (WB) analysis, and response to airway inflammation was evaluated by ELISA. The signaling pathway regulated by ECH was also investigated using WB. Our findings demonstrated that ECH restored OVA-induced increase in mucin, -immunoglobulin E, and respiratory resistance. ECH also alleviated OVA-induced collagen -deposition, including collagen I, collagen III, alpha smooth muscle actin, and epithelial (E)-cadherin. Moreover, ECH restored the elevated levels of interleukin (IL)-13, IL-17, and the increased -number of macrophages, eosinophils, lymphocytes, and neutrophills induced by OVA. ECH mainly exerted its regulatory effects by modulating the silent mating type information regulation 2 homolog 1 (Sirtuin 1/SIRT1)-nuclear factor kappa B (NF-κB) signaling pathway in the mouse models of asthma. This study highlights the therapeutic potential of ECH for attenuating airway remodeling and inflammation in an OVA-induced neonatal mouse model of asthma through the modulation of SIRT1/NF-κB pathway.

Schisandrin A ameliorates airway inflammation in model of asthma by attenuating Th2 response

Asthma is a persistent respiratory ailment that displays periodicity and is linked to the equilibrium of T cells. Several compounds obtained from Chinese herbal medicines display beneficial impacts on T cell regulation and the attenuation of inflammatory mediator synthesis. Schisandrin A, an active lignan derived from the Schisandra fruit, exhibits anti-inflammatory characteristics. In the present study, the network analysis conducted revealed that the nuclear factor-kappaB (NF-κB) signaling pathway is likely a prominent contributor to the anti-asthmatic effects of schisandrin A. In addition, it has been established that the inhibition of cyclooxygenase 2 (COX-2/PTGS2) is likely a significant factor in this process. The results of in vitro experiments have substantiated that schisandrin A can effectively lower the expression of COX-2 and inducible nitric oxide synthase (iNOS) in 16 HBE cells and RAW264.7 cells in a manner that is dependent on the dosage administered. It was able to effectively reduce the activation of the NF-κB signaling pathway while simultaneously improving the injury to the epithelial barrier function. Furthermore, an investigation utilizing immune infiltration as a metric revealed an inequity in Th1/Th2 cells and a surge in Th2 cytokines in asthma patients. In the OVA-induced asthma mice model, it was observed that schisandrin A treatment effectively suppressed inflammatory cell infiltration, reduced the Th2 cell ratio, inhibited mucus secretion, and prevented airway remodeling. To summarize, the administration of schisandrin A has been found to effectively alleviate the symptoms of asthma by impeding the production of inflammation, which includes reducing the Th2 cell ratio and improving the integrity of the epithelial barrier function. These findings offer valuable insights into the potential therapeutic applications of schisandrin A for the treatment of asthma.

PM2.5 exposure regulates Th1/Th2/Th17 cytokine production through NF-κB signaling in combined allergic rhinitis and asthma syndrome

Particulate matter (PM) is a major component of air pollution from emissions from anthropogenic and natural sources and is a serious problem worldwide due to its adverse effects on human health. Increased particulate air pollution increases respiratory disease-related mortality and morbidity. However, the impact of PM with an aerodynamic diameter of≤2.5μm (PM2.5) on combined allergic rhinitis and asthma syndrome (CARAS) remains to be elucidated. Accordingly, in the present study, we investigated the effect of PM2.5 in an ovalbumin (OVA)-induced CARAS mouse model with a focus on NF-κB signaling. We established an OVA-induced mouse model of CARAS to determine the effects of exposure to PM2.5. BALB/c mice were randomly divided into four groups: (1) naive, (2) PM2.5, (3) CARAS, and (4) CARAS/PM2.5. Mice were systemically sensitized with OVA and challenged with inhalation of ultrasonically nebulized 5% OVA three times by intranasal instillation of OVA in each nostril for 7 consecutive days. Mice in the PM2.5 and CARAS/PM2.5 groups were then exposed to PM2.5 by intranasal instillation of PM2.5 for several days. We then examined the impacts of PM2.5 exposure on histopathology and NF-κB signaling in our OVA-induced CARAS mouse model. PM2.5 increased infiltration of eosinophils in bronchoalveolar lavage fluid (BALF) samples and inflammatory cells in lung tissue. It also increased production of GATA3, RORγ, IL-4, IL-5, IL-13, and IL-17 in nasal lavage fluid (NALF) and BALF samples in the CARAS mouse model, but secretion of IL-12 and IFN-γ was suppressed. Exposure to PM2.5 increased OVA-specific IgE and IgG1 levels in serum, inflammatory cell infiltration in the airways, and fibrosis in lung tissue. It also activated the NF-κB signaling pathway, increasing Th2/Th17 cytokine levels while decreasing Th1 cytokine expression, thereby inducing an inflammatory response and promoting inflammatory cell infiltration in nasal and lung tissue. Our results demonstrate that PM2.5 can aggravate OVA-induced CARAS.

MiR-29a-3p promotes nasal epithelial barrier dysfunction via direct targeting of CTNNB1-VCL module in allergic rhinitis

Allergic rhinitis (AR) is resulted from immunoglobulin E (IgE)-mediated reactions to inhaled allergens which elicit mucosal inflammation and impair epithelial barrier integrity. However, whether miR-29a-3p as an epigenetic regulator that can contribute to epithelial barrier dysfunction in the pathogenesis of AR, and its underlying mechanism remians unclear. In this study, we discovered that miR-29a-3p was upregulated in AR patients and preferentially expressed in epithelial and glandular cells of nasal mucosa. VCL and CTNNB1, candidate target genes of miR-29a-3p, were predicted with several databases, including miRDB, miRanda, microT-CDS and TargetScan, and were validated through dual-luciferase reporter assay system. These two proteins were strongly associated with adherens junction (AJ) and tight junction (TJ) of nasal mucosa epithelial cells, in which played vital roles in mucosal integrity and nasal epithelial barrier function stability. Results for HNEpC culture and in vitro treatment experiments showed that expression of VCL and CTNNB1 were inhibited by miR-29a-3p mimic and were enhanced by miR-29a-3p inhibitor. In OVA-induced AR mice model, the expression pattern of miR-29a-3p and its target genes (Vcl and Ctnnb1) were consistent with the aforementioned quantitative results in AR patients, and miR-29a-3p antagomir could partially alleviate the symptom of OVA-induced AR in mice, restoring mucosal integrity and paracellular barrier function. In conclusion, our findings indicate that miR-29a-3p targets CTNNB1 and VCL to regulate nasal epithelial permeability and barrier function integrity, which may serve as a potential novel therapeutic target for the treatment of AR.

Baicalin Inhibits Airway Smooth Muscle Cells Proliferation through the RAS Signaling Pathway in Murine Asthmatic Airway Remodeling Model.

Studies that looked at asthma airway remodeling pathogenesis and prevention have led to the discovery of the rat sarcoma viral oncogene (RAS) signaling pathway as a key mechanism that controls airway smooth muscle cell (ASMC) proliferation. Baicalin has great anti-inflammatory, proliferation-inhibited, and respiratory disease-relieving properties. However, the inhibitory effects and mechanisms of baicalin on ASMC-mediated airway remodeling in mice are still poorly understood. After establishing the asthmatic mice model by ovalbumin (OVA) and interfering with baicalin, airway remodeling characteristics such as airway resistance, mRNA, and protein expression levels of remodeling-related cytokines were measured by histopathological assessment, quantitative real-time polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA), and western blot. Further efforts on detailed mechanisms were used antibody arrays to compare the expression and activation of proteins involved in the RAS signaling pathway. In addition, validation experiments were performed in ASMC proliferation model and low-expression cells of the target gene by using shRNA. In OVA-induced asthmatic mice model, baicalin significantly reduced the infiltration of inflammatory cells in lung tissue, attenuated airway resistance, and decreased mRNA and protein expression levels of remodeling-related cytokines such as interleukin-13 (IL-13), vascular endothelial growth factor (VEGF), transforming growth factor-beta 1 (TGF-β1), matrix metallopeptidase 9 (MMP9), and tissue inhibitor of metalloproteinase 1 (TIMP1). The results of antibody arrays involved in RAS signaling pathway revealed that OVA and baicalin administration altered the activation of protein kinase C alpha type (PKC-α), A-rapidly accelerated fibrosarcoma (A-RAF), mitogen-activated protein kinase 2 (MEK2), extracellular regulated MAP kinase (ERK), MAPK interacting serine/threonine kinase 1 (MNK1), and ETS transcription factor 1 (ELK1). The above results were further verified in the ASMC proliferation model. A-RAF silencing (shA-RAF) could promote ASMC proliferation and downregulate p-MEK2, p-ERK, p-MNK1, and p-ELK1 expression. The effects of baicalin against airway remodeling and ASMC proliferation might partially be achieved by suppressing the RAS signaling pathway. Baicalin may be a new therapeutic option for managing airway remodeling in asthma patients.

CCL4 Regulates Eosinophil Activation in Eosinophilic Airway Inflammation.

Eosinophilic chronic rhinosinusitis (ECRS) is a refractory airway disease accompanied by eosinophilic inflammation, the mechanisms of which are unknown. We recently found that CCL4/MIP-1β-a specific ligand for CCR5 receptors-was implicated in eosinophil recruitment into the inflammatory site and was substantially released from activated eosinophils. Moreover, it was found in nasal polyps from patients with ECRS, primarily in epithelial cells. In the present study, the role of epithelial cell-derived CCL4 in eosinophil activation was investigated. First, CCL4 expression in nasal polyps from patients with ECRS as well as its role of CCL4 in eosinophilic airway inflammation were investigated in an in vivo model. Furthermore, the role of CCL4 in CD69 expression-a marker of activated eosinophils-as well as the signaling pathways involved in CCL4-mediated eosinophil activation were investigated. Notably, CCL4 expression, but not CCL5, CCL11, or CCL26, was found to be significantly increased in nasal polyps from patients with ECRS associated with eosinophil infiltration as well as in BEAS-2B cells co-incubated with eosinophils. In an OVA-induced allergic mouse model, CCL4 increased eosinophil accumulation in the nasal mucosa and the bronchoalveolar lavage (BALF). Moreover, we found that CD69 expression was upregulated in CCL4-stimulated eosinophils; similarly, phosphorylation of several kinases, including platelet-derived growth factor receptor (PDGFR)β, SRC kinase family (Lck, Src, and Yes), and extracellular signal-regulated kinase (ERK), was upregulated. Further, CCR5, PDGFRβ, and/or Src kinase inhibition partially restored CCL4-induced CD69 upregulation. Thus, CCL4, which is derived from airway epithelial cells, plays a role in the accumulation and activation of eosinophils at inflammatory sites. These findings may provide a novel therapeutic target for eosinophilic airway inflammation, such as ECRS.

Intranasal administration of nucleus-deliverable GATA3-TMD alleviates the symptoms of allergic asthma

Although the T helper 2 (Th2) subset is a critical player in the humoral immune response to extracellular parasites and suppression of Th1-mediated inflammation, Th2 cells have been implicated in allergic inflammatory diseases such as asthma, allergic rhinitis, and atopic dermatitis. GATA binding protein 3 (GATA3) is a primary transcription factor that mediates Th2 differentiation and secretion of Th2 cytokines, including IL-4, IL-5, and IL-13. Here, a nucleus-deliverable form of GATA3-transcription modulation domain (TMD) (ndG3-TMD) was generated using Hph-1 human protein transduction domain (PTD) to modulate the transcriptional function of endogenous GATA3 without genetic manipulation. ndG3-TMD was shown to be efficiently delivered into the cell nucleus quickly without affecting cell viability or intracellular signaling events for T cell activation. ndG3-TMD exhibited a specific inhibitory function for the endogenous GATA3-mediated transcription, such as Th2 cell differentiation and Th2-type cytokine production. Intranasal administration of ndG3-TMD significantly alleviated airway hyperresponsiveness, infiltration of immune cells, and serum IgE level in an OVA-induced mouse model of asthma. Also, Th2 cytokine secretion by the splenocytes isolated from the ndG3-TMD-treated mice substantially decreased. Our results suggest that ndG3-TMD can be a new therapeutic reagent to suppress Th2-mediated allergic diseases through intranasal delivery.

Pyrroloquinoline Quinone Administration Alleviates Allergic Airway Inflammation in Mice by Regulating the JAK-STAT Signaling Pathway.

The current asthma therapies are inadequate for many patients with severe asthma. Pyrroloquinoline quinone (PQQ) is a naturally-occurring redox cofactor and nutrient that can exert a multitude of physiological effects, including anti-inflammatory and antioxidative effects. We sought to explore the effects of PQQ on allergic airway inflammation and reveal the underlying mechanisms. In vitro, the effects of PQQ on the secretion of epithelial-derived cytokines by house dust mite- (HDM-) incubated 16-HBE cells and on the differentiation potential of CD4+ T cells were investigated. In vivo, PQQ was administered to mice with ovalbumin- (OVA-) induced asthma, and lung pathology and inflammatory cell infiltration were assessed. The changes in T cell subsets and signal transducers and activators of transcription (STATs) were evaluated by flow cytometry. Pretreatment with PQQ significantly decreased HDM-stimulated thymic stromal lymphopoietin (TSLP) production in a dose-dependent manner in 16-HBE cells and inhibited Th2 cell differentiation in vitro. Treatment with PQQ significantly reduced bronchoalveolar lavage fluid (BALF) inflammatory cell counts in the OVA-induced mouse model. PQQ administration also changed the secretion of IFN-γ and IL-4 as well as the percentages of Th1, Th2, Th17, and Treg cells in the peripheral blood and lung tissues, along with inhibition the phosphorylation of STAT1, STAT3, and STAT6 while promoting that of STAT4 in allergic airway inflammation model mice. PQQ can alleviate allergic airway inflammation in mice by improving the immune microenvironment and regulating the JAK-STAT signaling pathway. Our findings suggest that PQQ has great potential as a novel therapeutic agent for inflammatory diseases, including asthma.

Zeaxanthin attenuates OVA-induced allergic asthma in mice by regulating the p38 MAPK/β-catenin signaling pathway

Background: Asthma is a heterogeneous and complex chronic airway disease with a high incidence rate, characterized by chronic airway inflammation. Although the anti-inflammatory effect of zeaxanthin has been demonstrated in various disease models, its explicit role in allergic asthma remains elusive. Methods: An allergic asthma model was established by ovalbumin (OVA) stimulation in BALB/c nude mice. The pathological examination, collagen deposition and expression of α-smooth muscle actin (α-SMA) in lung tissues were determined by hematoxylin and eosin (H&E), MASSON and immunofluorescence staining, respectively. Besides, the effect of zeaxanthin on inflammation and oxidative stress was assessed by the enzyme-linked immunosorbent assay (ELISA) and spectrophotometry measure. Moreover, the underlying mechanism was analyzed by detecting the expression of phosphorylated p38 (p-p38), p38, β-catenin, p-c-Jun N-terminal kinase (p-JNK) and JNK with western blot assays. Results: The distinct infiltration of inflammatory cells was observed in the OVA-induced asthma mice model with significantly increased concentrations of immunoglobulin E (IgE), interleukin-4 (IL-4), IL-5, IL-13 and eotaxin (p˂0.001), which were prominently reversed by zeaxanthin treatment (p˂0.001). In addition, zeaxanthin treatment decreased the OVA-induced collagen deposition and α-SMA expression. A similar inhibitory effect of zeaxanthin on the oxidative stress was also observed in the OVA-induced asthma mice model, as evidenced by the prominent decrease of malondialdehyde (MDA) concentration and the remarkable increase of superoxide dismutase (SOD), glutathione S transferase (GST) and Glutathione (GSH) concentrations (p˂0.001). Moreover, zeaxanthin introduction markedly reduced the relative expressions of p-p38/p38, β-catenin and p-JNK/JNK in the OVA-induced asthma mice model (p˂0.001), indicating that zeaxanthin suppressed the p38 mitogen-activated protein kinase (p38 MAPK)/β-catenin signaling pathway in the OVA-induced asthma mice model. Conclusions: Zeaxanthin attenuated OVA-induced allergic asthma in mice via modulating the p38 MAPK/β-catenin signaling pathway.

Therapeutic Effect of Renifolin F on Airway Allergy in an Ovalbumin-Induced Asthma Mouse Model In Vivo.

Renifolin F is a prenylated chalcone isolated from Shuteria involucrata, a traditional minority ethnic medicine used to treat the respiratory diseases and asthma. Based on the effects of the original medicine plant, we established an in vivo mouse model of allergic asthma using ovalbumin (OVA) as an inducer to evaluate the therapeutic effects of Renifolin F. In the research, mice were sensitized and challenged with OVA to establish an allergic asthma model to evaluate the effects of Renifolin F on allergic asthma. The airway hyper-reactivity (AHR) to methacholine, cytokine levels, ILC2s quantity and mircoRNA-155 expression were assessed. We discovered that Renifolin F attenuated AHR and airway inflammation in the OVA-induced asthmatic mouse model by inhibiting the regulation of ILC2s in the lung, thereby, reducing the upstream inflammatory cytokines IL-25, IL-33 and TSLP; the downstream inflammatory cytokines IL-4, IL-5, IL-9 and IL-13 of ILC2s; and the co-stimulatory factors IL-2 and IL-7; as well as the expression of microRNA-155 in the lung. The findings suggest a therapeutic potential of Renifolin F on OVA-induced airway inflammation.

Phosphatidylethanolamine-binding protein 1 (PEBP1) mediates the regulatory role of microRNAs (miRNAs)-205-5p in degranulation and histamine release

ABSTRACT miR-205-5p plays a vital role in the inflammation of allergic rhinitis (AR). The study is designed to investigate the effects and mechanism of miR-205-5p in AR in vivo and in vitro. An OVA-induced mice model and anti-DNP IgE-induced RBL-2H3 cell model were established. The pathological alterations in the nasal mucosa were evaluated by hematoxylin-eosin (HE) staining. IgE and histamine levels were detected by corresponding kits and the expressions of PEBP1, High mobility group box-1 (HMGB1) and Toll-like receptor 4 (TLR4) were detected by western blot. The association of miR-205-5p and PEBP1 was determined by dual-luciferase reported assay. β-hexosaminidase activity was to evaluate the degranulation of RBL-2H3 cell. The pathological injury of nasal mucosa was significantly improved by miR-205-5p inhibition compared to AR mice. Following the treatment of miR-205-5p inhibitor, the levels of helper T cell (Th1) cytokines, interleukin (IL)-2 and interferon-γ (IFN-γ) were increased, while the levels of Th2 cytokines, IL-4 and IL-13, as well as the levels of IgE and histamine were markedly decreased in AR mice. We further found that miR-205-5P inhibition induced increased expression of PEBP1 and decreased expressions of HMGB1and TLR4. In vitro, miR-205-5P was verified to bind to PEBP1. PEBP1 silencing led to the reverse of miR-205-5p effects on decreasing the levels of β-hexosaminidase activity and histamine, as well as the expressions of HMGB1 and TLR4 on anti-DNP IgE-induced RBL-2H3 cells. Our results indicate that miR-205-5P inhibition may ameliorate pathological injury via PEBP1. MiR-205-5P/ PEBP1 could be potential drug targets in AR

Follistatin-related protein 1 in asthma: miR-200b-3p interactions affect airway remodeling and inflammation phenotype

Follistatin-related protein 1 (FSTL1) is significantly associated with the asthma severity and outcome in humans and diverse mouse models of asthma. Previous studies have also suggested that FSTL1 could activate autophagy and NLRP3, thus playing as a causative agent in the asthma progression. However, mechanisms that regulate airway epithelial cell-specific FSTL1 expression and function in asthma are unknown. Here, we further evaluated the spatiotemporal relationships between the FSTL1 and asthma development through ovalbumin (OVA) -induced asthma models. Integrative analysis in asthmatics airway epithelium identifies microRNA (miR)-200b-3p as a novel upstream of FSTL1. Next, we collected airway biopsies, induced sputum, and blood samples isolated from asthmatics patients and the OVA-induced mouse model. We revealed that miR-200b-3p expression is downregulated in asthmatics airway epithelium, while its expression was negatively correlated with FSTL1. On this basis, the function and expression pattern analysis of miR-200b-3p were performed using miRNA-target prediction databases and long non-coding RNA (lncRNA) microarray assay. It is illustrated that miR-200b-3p, which is downregulated with pro-fibrotic stimulation of TGF-β1, could also be sponged by lncRNA PCAT19 and regulate FSTL1 expression in asthma progression. In vivo, miR-200b-3p overexpression in mice prevents OVA-induced airway remodeling and inflammation. Lastly, protective roles of miR-200b-3p are partly attributed to the direct and functional repression of FSTL1. Our findings suggest a crucial role for the miR-200b-3p/FSTL1 axis in regulating asthmatic’s airway remodeling and inflammation phenotype.