Studying of anti-inflammatory and antioxidant effects of tectorigenin in ovalbumin-induced asthma mice models.

BACKGROUND: Allergic asthma is an important respiratory system problem characterized by airway inflammation, breathlessness, and bronchoconstriction. Allergic asthma and its outcomes are triggered by type 2 allergic immune responses. Tectorigenin is a methoxy-isoflavone with anti-inflammatory effects. In this study, we investigated the effects of tectorigenin on the pathophysiology of allergic asthma in an animal model. METHODS: Asthmatic mice were treated with tectorigenin. Then airway hyperresponsiveness (AHR), eosinophil percentage, levels of interleukin (IL)-33, IL-25, IL-13, IL-5, IL-4, total and ovalbumin (OVA)-specific immunoglobulin (Ig)E, and lung histopathology were evaluated. RESULT: Tectorigenin significantly (P 〈 0.05) reduced eosinophil infiltration (41 ± 7%) in the broncho-alveolar lavage fluid (BALF), serum IL-5 level (41 ± 5, pg/mL), and bronchial and vascular inflammation (scores of 1.3 ± 0.2 and 1.1 ± 0.3, respectively) but had no significant effects on AHR, serum levels of IL-33, -25, -13, and -4 (403 ± 24, 56 ± 7, 154 ± 11, and 89 ± 6 pg/mL, respectively), total and OVA-specific IgE (2684 ± 265 and 264 ± 19 ng/mL, respectively), goblet cell hyperplasia, and mucus production. CONCLUSION: Tectorigenin could control inflammation and the secretion of inflammatory mediators of asthma, so it can be regarded as a potential antiasthma treatment with the ability to control eosinophilia-related problems.

PI3Kγδ inhibition suppresses key disease features in a rat model of asthma.

BACKGROUND: Two isoforms of Phosphoinositide 3-kinase (PI3K), p110γ and p110δ, are predominantly expressed in leukocytes and represent attractive therapeutic targets for the treatment of allergic asthma. The study aim was to assess the impact of administration of an inhaled PI3Kγδ inhibitor (AZD8154) in a rat model of asthma. METHODS: Firstly, we checked that the tool compound, AZD8154, inhibited rat PI3K γ & δ kinases using rat cell-based assays. Subsequently, a time-course study was conducted in a rat model of asthma to assess PI3K activity in the lung and how it is temporally associated with other key transcription pathways and asthma like features of the model. Finally, the impact on lung dosed AZD8154 on target engagement, pathway specificity, airway inflammation and lung function changes was assessed. RESULTS: Data showed that AZD8154 could inhibit rat PI3K γ & δ isoforms and, in a rat model of allergic asthma the PI3K pathway was activated in the lung. Intratracheal administration of AZD8154 caused a dose related suppression PI3K pathway activation (reduction in pAkt) and unlike after budesonide treatment, STAT and NF-κB pathways were not affected by AZD8154. The suppression of the PI3K pathway led to a marked inhibition of airway inflammation and reduction in changes in lung function. CONCLUSION: These data show that a dual PI3Kγδ inhibitor suppress key features of disease in a rat model of asthma to a similar degree as budesonide and indicate that dual PI3Kγδ inhibition may be an effective treatment for people suffering from allergic asthma.

Endothelin-1-mediated Brainstem Glial Activation Produces Asthmatic Airway Vagal Hypertonia Via Enhanced ATP-P2X4 Receptor Signaling in Sprague-Dawley Rats.

The occurrence of major asthma symptoms is largely attributed to airway vagal hypertonia, of which the central mechanisms remain unclear. This study tests the hypotheses that endothelin-1-mediated brainstem glial activation produces asthmatic airway vagal hypertonia via enhanced action of adenosine 5'-triphosphate on neuronal purinergic P2X4 receptors. A rat model of asthma was prepared using ovalbumin. Airway vagal tone was evaluated by the recurrent laryngeal discharge and plethysmographic measurement of pulmonary function. The changes in the brainstem were examined using ELISA, Western blot, luciferin-luciferase, quantitative reverse transcription-polymerase chain reaction, enzyme activity assay and immunofluorescent staining, respectively. The results showed that in the medulla of rats, endothelin receptor type B and P2X4 receptors were primarily expressed in astrocytes and neurons, respectively, and both of which, along with endothelin-1 content, were significantly increased after ovalbumin sensitization. Ovalbumin sensitization significantly increased recurrent laryngeal discharge, which was blocked by acute intracisternal injection of P2X4 receptor antagonist 5-BDBD, knockdown of brainstem P2X4 receptors, and chronic intraperitoneal injection of endothelin receptor type B antagonist BQ788, respectively. Ovalbumin sensitization activated microglia and astrocytes and significantly decreased ecto-5'-nucleotidase activity in the medulla, and all of which, together with the increase of medullary P2X4 receptor expression and decrease of pulmonary function, were reversed by chronic BQ788 treatment. These results demonstrated that in rats, allergic airway challenge activates both microglia and astrocytes in the medulla via enhanced endothelin-1/endothelin receptor type B signaling, which subsequently causes airway vagal hypertonia via augmented adenosine 5'-triphosphate/P2X4 receptor signaling in central neurons of airway vagal reflex.

A novel quinoline with airway relaxant effects and anti-inflammatory properties.

BACKGROUND: In chronic pulmonary diseases characterized by inflammation and airway obstruction, such as asthma and COPD, there are unmet needs for improved treatment. Quinolines is a group of small heterocyclic compounds that have a broad range of pharmacological properties. Here, we investigated the airway relaxant and anti-inflammatory properties of a novel quinoline (RCD405). METHODS: The airway relaxant effect of RCD405 was examined in isolated airways from humans, dogs, rats and mice. Murine models of ovalbumin (OVA)-induced allergic asthma and LPS-induced airway inflammation were used to study the effects in vivo. RCD405 (10 mg/kg) or, for comparisons in selected studies, budesonide (3 mg/kg), were administered intratracheally 1 h prior to each challenge. Airway responsiveness was determined using methacholine provocation. Immune cell recruitment to bronchi was measured using flow cytometry and histological analyses were applied to investigate cell influx and goblet cell hyperplasia of the airways. Furthermore, production of cytokines and chemokines was measured using a multiplex immunoassay. The expression levels of asthma-related genes in murine lung tissue were determined by PCR. The involvement of NF-κB and metabolic activity was measured in the human monocytic cell line THP-1. RESULTS: RCD405 demonstrated a relaxant effect on carbachol precontracted airways in all four species investigated (potency ranking: human = rat > dog = mouse). The OVA-specific IgE and airway hyperresponsiveness (AHR) were significantly reduced by intratracheal treatment with RCD405, while no significant changes were observed for budesonide. In addition, administration of RCD405 to mice significantly decreased the expression of proinflammatory cytokines and chemokines as well as recruitment of immune cells to the lungs in both OVA- and LPS-induced airway inflammation, with a similar effect as for budesonide (in the OVA-model). However, the effect on gene expression of Il-4, IL-5 and Il-13 was more pronounced for RCD405 as compared to budesonide. Finally, in vitro, RCD405 reduced the LPS-induced NF-κB activation and by itself reduced cellular metabolism. CONCLUSIONS: RCD405 has airway relaxant effects, and it reduces AHR as well as airway inflammation in the models used, suggesting that it could be a clinically relevant compound to treat inflammatory airway diseases. Possible targets of this compound are complexes of mitochondrial oxidative phosphorylation, resulting in decreased metabolic activity of targeted cells as well as through pathways associated to NF-κB. However, further studies are needed to elucidate the mode of action.

Protective effects of myricetin on airway inflammation and oxidative stress in ovalbumin-induced asthma mice.

Myricetin, a flavonoid isolated from many edible vegetables and fruits, has multiple biological effects, including anti-inflammatory and anti-tumor effects. Myricetin could inhibit mast cell degranulation in vitro, and it reduced the eosinophil content in bronchoalveolar lavage fluid (BALF) of ovalbumin (OVA)-sensitized mice. However, it remains unclear whether myricetin alleviates airway hyperresponsiveness (AHR), airway inflammation, and oxidative stress in asthma. Here, we investigated whether myricetin attenuated AHR, airway inflammation, and eosinophil infiltration in lungs of asthmatic mice. Mice were sensitized with OVA, then injected intraperitoneally with myricetin to investigate anti-inflammatory and antioxidant effects of myricetin. Moreover, we examined its effects on human bronchial epithelial BEAS-2B cells stimulated with TNF-α and IL-4, in vitro. Myricetin effectively mitigated eosinophil infiltration, AHR, and goblet cell hyperplasia in lung, and it reduced Th2 cytokine expression in BALF from asthmatic mice. Myricetin effectively promoted glutathione and superoxide dismutase productions and mitigated malondialdehyde expressions in mice by promoting Nrf2/HO-1 expression. Myricetin also reduced the production of proinflammatory cytokines, eotaxins, and reactive oxygen species in BEAS-2B cells. Myricetin effectively suppressed ICAM-1 expression in inflammatory BEAS-2B cells, which suppressed monocyte cell adherence. These results suggested that myricetin could effectively improve asthma symptoms, mainly through blocking Th2-cell activation, which reduced oxidative stress, AHR, and airway inflammation.

The IL-31/TRPV1 pathway mediates allergic asthma exacerbated by DINP dermal exposure in OVA-sensitized Balb/c mice.

BACKGROUND: The potential role of dermal exposure diisononyl phthalate (DINP) as an adjuvant in allergic inflammation and asthma has been suggested. However, the current findings do not provide enough evidence to support this claim. OBJECTIVES: The purpose of this investigation was to examine the impact and mechanisms of allergic asthma exacerbation through the dermal exposure to DINP. METHODS: The study was undertaken using OVA-sensitized mice. Lung histopathology and airway hyperreactivity (AHR) were assessed. Expression levels of immunoglobulins (t-IgE, OVA-IgE and OVA-IgG1), cytokines (IL-31, IL-4, IL-5, IL-6, IL-13 and INF-γ), and TRPV1 were measured. To investigate the mechanism by which allergic asthma worsens due to dermal exposure to DINP, the blockade analysis using the IL-31 antagonist SB-431542 and the TRPV1 antagonist capsazepine (CZP) were performed. RESULTS: The findings of the study revealed that the simultaneous exposure to DINP and OVA resulted in an increase in inspiratory resistance (Ri) and expiratory resistance (Re), a decrease in the minimum value of lung dynamic compliance (Cldyn), and worsened airway remodeling. Additionally, it was found that this exposure led to an increase in the levels of IL-31 and TRPV1, which are biomarkers of Th2 cytokines (IL-4, IL-5, IL-6, and IL-13), as well as immunoglobulins (Total IgE, OVA-lgE, and OVA-IgG1), while decreasing the biomarker of Th1 cytokines (IFN-γ). However, these impairments showed improvement after the administration of SB-431542 or CZP. CONCLUSION: The findings of this research indicate that the IL-31/TRPV1 pathway plays a moderating function in OVA-induced allergic asthma worsened by dermal exposure to DINP.

ABHD2 deficiency aggravates ovalbumin-induced airway remodeling through the PI3K/Akt pathway in an animal model of chronic asthma.

Airway remodeling is a major pathological characteristic of chronic obstructive pulmonary disease (COPD). This study aimed to investigate the effect of Abhd2 deficiency on ovalbumin (OVA)-induced airway remodeling and inflammation in vivo. Abhd2-deficient mice were used to establish an OVA-induced asthma model. Lung tissues were analyzed using hematoxylin and eosin (HE) staining, Masson staining, immunohistochemistry, quantitative reverse transcription- polymerase chain reaction (qRT-PCR), and western blotting were used to determine the role of Abhd2 in the regulation of OVA-induced airway remodeling and inflammation. Our findings revealed that the RNA expression of inflammatory factors, including IL-1ß, IL-6, IL-4, and IL-13, was significantly increased in OVA-induced Abhd2 Gt/Gt asthmatic mice. The expression of IFN-γ was decreased significantly in OVA-induced Abhd2 Gt/Gt asthmatic mice. The protein expression of airway remodeling factors, including α-SMA, type I collagen, and Ki67, was also increased in OVA-induced Abhd2 Gt/Gt asthmatic mice compared to that in OVA-induced wild-type (WT) mice. Additionally, Abhd2 deficiency promoted the expression of p-Akt in tissues of the asthma model. These results suggest that Abhd2 deficiency exacerbates airway remodeling and inflammation through the PI3K/Akt pathway in chronic asthma.

MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

BACKGROUND: Neutrophilic airway inflammation is a challenge in asthma management and is associated with poor patient prognosis. Mucin 1 (MUC1), which contains a cytoplasmic tail (MUC1-CT), has been found to mediate glucocorticoid sensitivity in asthma; however, its role in modulating neutrophilic airway inflammation in asthma remains unknown. METHODS: Human-induced sputum cells were collected from healthy participants (n = 12), patients with mild-to-moderate asthma (n = 34), and those with severe asthma (n = 18). In vitro human lung bronchial 1 epithelial cell line (BEAS-2B) was transfected with small interfering RNA against MUC1 (MUC1-siRNA) and then stimulated by lipopolysaccharide (LPS), where some cells were pretreated with a TLR4 inhibitor (TAK-242). In vivo mouse model of asthmatic neutrophil airway inflammation was induced by ovalbumin (OVA)/LPS. Some groups were intraperitoneally injected with MUC1-CT inhibitor (GO-203) and/or TAK-242 . RESULTS: The mRNA expression of MUC1 was downregulated in the induced sputum of patients with asthma and correlated with asthmatic neutrophilic airway inflammation. The mRNA expressions of TLR4, MyD88, nucleotide-binding oligomerization domain-like pyrin domain-containing protein 3 (NLRP3), caspase-1, interleukin (IL)-18, and IL-1ß in induced sputum cells of patients with asthma were upregulated and related to the mRNA expression of MUC1. LPS activated the TLR4 pathway and NLRP3-mediated pyroptosis in BEAS-2B cells in vitro, which were significantly aggravated after MUC1-siRNA transfection. Furthermore, MUCl-CT interacted with TLR4, and the interaction between TLR4 and MyD88 was significantly increased after MUCl-siRNA transfection. Moreover, TAK-242 ameliorated TLR4/MyD88/nuclear factor kappa B (NF-κB) pathway activation, NLRP3 inflammasome-mediated pyroptosis, and neutrophilic inflammation exacerbated by MUC1 downregulation. GO-203 exacerbated TLR4/MyD88/NF-κB pathway activation in vivo, and NLRP3 inflammasome-mediated pyroptosis reduced in a mouse model of asthmatic neutrophil airway inflammation induced by OVA/LPS; these pathological changes were partially alleviated after TAK-242 application. CONCLUSION: This study revealed that MUC1 downregulation plays an important role in asthmatic neutrophilic airway inflammation. MUC1-CT reduces NLRP3 inflammasome-mediated pyroptosis by inhibiting the activation of the TLR4/MyD88/NF-κB pathway, thereby attenuating neutrophil airway inflammation in patients with asthma.

Adeno-Associated Viral Vector-Delivered Pannexin-1 Mimetic Peptide Alleviates Airway Inflammation in an Allergen-Sensitized Mouse Model.

Asthma is a chronic inflammatory disease around the world. Extracellular adenosine triphosphate works as a dangerous signal in responding to cellular stress, irritation, or inflammation. It has also been reported its association with the pathogenicity in asthma, with increased level in lungs of asthmatics. Pannexin-1 is one of the routes that contributes to the release of adenosine triphosphate form intracellular to extracellular. The aim of this study was to apply pannexin-1 peptide antagonist 10Panx1 into adeno-associated viral (AAV) vectors on ovalbumin (OVA)-induced asthmatic mouse model. The results demonstrated that this treatment was able to reduce the adenosine triphosphate level in bronchoalveolar lavage fluid and downregulate the major relevant to the symptom of asthma attack, airway hyperresponsiveness to methacholine. The histological data also gave a positive support with decreased tissue remodeling and mucus deposition. Other asthmatic related features, including eosinophilic inflammation and OVA-specific T helper type 2 responses, were also decreased by the treatment. Beyond the index of inflammation, the proportion of effector and regulatory T cells was examined to survey the potential mechanism behind. The data provided a slightly downregulated pattern in lung GATA3+ CD4 T cells. However, an upregulated population of CD25+FoxP3+ CD4 T cells was seen in spleens. These data suggested that exogeneous expression of 10Panx1 peptide was potential to alleviated asthmatic airway inflammation, and this therapeutic effect might be from 10Panx1-mediated disruption of T cell activation or differentiation. Collectively, AAV vector-mediated 10Panx1 expression could be a naval therapy option to develop.

Beneficial Effects of Astragalus membranaceus (Fisch.) Bunge Extract in Controlling Inflammatory Response and Preventing Asthma Features.

Astragalus membranaceus (Fisch.) Bunge root is used as herbal medicine for its immunomodulating activities in Chinese medicine. Recently, beneficial properties of A. membranaceus on allergic diseases have been proposed. Here we investigated the role of a commercial extract of A. membranaceus, standardized to 16% polysaccharides, in regulating the immune-inflammatory response in vitro and in vivo and its therapeutic application in asthma. A. membranaceus extract inhibited prostaglandin E2 and leukotriene C4 production in stimulated J774 and peritoneal macrophages, respectively. The extract also reduced interlukin-1ß, tumor necrosis factor-α, and nitrite production, affecting inducible nitric oxide synthase expression. In vivo experiments confirmed the anti-inflammatory properties of A. membranaceus, as evident by a reduction in zymosan-induced peritoneal cellular infiltration and pro-inflammatory mediator production. The efficacy of A. membranaceus extract in modulating the immune response was confirmed in a model of allergic airway inflammation. Extracts improve lung function by inhibiting airway hyperresponsiveness, airway remodeling, and fibrosis. Its anti-asthmatic effects were further sustained by inhibition of the sensitization process, as indicated by a reduction of ovalbumin-induced IgE levels and the mounting of a Th2 immune response. In conclusion, our data demonstrate the anti-inflammatory properties of the commercial extract of A. membranaceus and its beneficial effects on asthma feature development.

Pi-Pa-Run-Fei-Tang alleviates lung injury by modulating IL-6/JAK2/STAT3/IL-17 and PI3K/AKT/NF-κB signaling pathway and balancing Th17 and Treg in murine model of OVA-induced asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Pi-Pa-Run-Fei-Tang (PPRFT) is an empirical TCM prescription for treating asthma. However, the underlying mechanisms of PPRFT in asthma treatment have yet to be elucidated. Recent advances have revealed that some natural components could ameliorate asthma injury by affecting host metabolism. Untargeted metabolomics can be used to better understand the biological mechanisms underlying asthma development and identify early biomarkers that can help advance treatment. AIM OF THE STUDY: The aim of this study was to verification the efficacy of PPRFT in the treatment of asthma and to preliminarily explore its mechanism. MATERIALS AND METHODS: A mouse asthma model was built by OVA induction. Inflammatory cell in BALF was counted. The level of IL-6, IL-1ß, and TNF-α in BALF were measured. The levels of IgE in the serum and EPO, NO, SOD, GSH-Px, and MDA in the lung tissue were measured. Furthermore, pathological damage to the lung tissues was detected to evaluate the protective effects of PPRFT. The serum metabolomic profiles of PPRFT in asthmatic mice were determined by GC-MS. The regulatory effects on mechanism pathways of PPRFT in asthmatic mice were explored via immunohistochemical staining and western blotting analysis. RESULTS: PPRFT displayed lung-protective effects through decreasing oxidative stress, airway inflammation, and lung tissue damage in OVA-induced mice, which was demonstrated by decreasing inflammatory cell levels, IL-6, IL-1ß, and TNF-α levels in BALF, and IgE levels in serum, decreasing EPO, NO, and MDA levels in lung tissue, elevating SOD and GSH-Px levels in lung tissue and lung histopathological changes. In addition, PPRFT could regulate the imbalance in Th17/Treg cell ratios, suppress RORγt, and increase the expression of IL-10 and Foxp3 in the lung. Moreover, PPRFT treatment led to decreased expression of IL-6, p-JAK2/Jak2, p-STAT3/STAT3, IL-17, NF-κB, p-AKT/AKT, and p-PI3K/PI3K. Serum metabolomics analysis revealed that 35 metabolites were significantly different among different groups. Pathway enrichment analysis indicated that 31 pathways were involved. Moreover, correlation analysis and metabolic pathway analysis identified three key metabolic pathways: galactose metabolism; tricarboxylic acid cycle; and glycine, serine, and threonine metabolism. CONCLUSION: This research indicated that PPRFT treatment not only attenuates the clinical symptoms of asthma but is also involved in regulating serum metabolism. The anti-asthmatic activity of PPRFT may be associated with the regulatory effects of IL-6/JAK2/STAT3/IL-17 and PI3K/AKT/NF-κB mechanistic pathways.

Effect of NO2 exposure on airway inflammation and oxidative stress in asthmatic mice.

Nitrogen dioxide (NO2) is a widespread air pollutant. Epidemiological evidence indicates that NO2 is associated with an increase of incidence rate and mortality of asthma, but its mechanism is still unclear. In this study, we exposed mice to NO2 (5 ppm, 4 h per day for 30 days) intermittently to investigate the development and potential toxicological mechanisms of allergic asthma. We randomly assigned 60 male Balb/c mice to four groups: saline control, ovalbumin (OVA) sensitization, NO2 alone, and OVA+NO2 groups. The involved mechanisms were found from the perspective of airway inflammation and oxidative stress. The results showed that NO2 exposure could aggravate lung inflammation in asthmatic mice, and airway remodeling was characterized by significant thickening of the airway wall and infiltration of inflammatory cells. Moreover, NO2 would aggravate the airway hyperresponsiveness (AHR), which is characterized by significantly elevated inspiratory resistance (Ri) and expiratory resistance (Re), as well as decreased dynamic lung compliance (Cldyn). In addition, NO2 exposure promoted pro-inflammatory cytokines (IL-6 and TNF-α) and serum immunoglobulin (IgE) production. The imbalance of Th1/Th2 cell differentiation (IL-4 increased, IFN-γ reduced, IL-4/IFN-γ significantly increased) played a key role in the inflammatory response of asthma under NO2 exposure. In a nutshell, NO2 exposure could promote allergic airway inflammation and increase asthma susceptibility. The levels of ROS and MDA among asthmatic mice exposed to NO2 increased significantly, while GSH levels sharply decreased. These findings may provide better toxicological evidence for the mechanisms of allergic asthma risk due to NO2 exposure.

Effects of mesenchymal stem cell-derived nanovesicles in experimental allergic airway inflammation.

BACKGROUND: Allergic asthma is associated with airflow obstruction and hyper-responsiveness that arises from airway inflammation and remodeling. Cell therapy with mesenchymal stem cells (MSC) has been shown to attenuate inflammation in asthma models, and similar effects have recently been observed using extracellular vesicles (EV) obtained from these cells. Biologically functional vesicles can also be artificially generated from MSC by extruding cells through membranes to produce EV-mimetic nanovesicles (NV). In this study, we aimed to determine the effects of different MSC-derived vesicles in a murine model of allergic airway inflammation. METHODS: EV were obtained through sequential centrifugation of serum-free media conditioned by human bone marrow MSC for 24 h. NV were produced through serial extrusion of the whole cells through filters. Both types of vesicles underwent density gradient purification and were quantified through nanoparticle tracking analysis. C57BL/6 mice were sensitized to ovalbumin (OVA, 8 µg), and then randomly divided into the OVA group (intranasally exposed to 100 µg OVA for 5 days) and control group (exposed to PBS). The mice were then further divided into groups that received 2 × 109 EV or NV (intranasally or intraperitoneally) or PBS immediately following the first OVA exposure. RESULTS: Administration of EV and NV reduced cellularity and eosinophilia in bronchoalveolar lavage (BAL) fluid in OVA-sensitized and OVA-exposed mice. In addition, NV treatment resulted in decreased numbers of inflammatory cells within the lung tissue, and this was associated with lower levels of Eotaxin-2 in both BAL fluid and lung tissue. Furthermore, both intranasal and systemic administration of NV were effective in reducing inflammatory cells; however, systemic delivery resulted in a greater reduction of eosinophilia in the lung tissue. CONCLUSIONS: Taken together, our results indicate that MSC-derived NV significantly reduce OVA-induced allergic airway inflammation to a level comparable to EV. Thus, cell-derived NV may be a novel EV-mimetic therapeutic candidate for treating allergic diseases such as asthma.

Interaction of high temperature and NO2 exposure on asthma risk: In vivo experimental evidence of inflammation and oxidative stress.

Allergic asthma is a complicated respiratory disease with many concerns. Mounting epidemiological evidence linked temperature (T) and NO2 with allergic asthma, yet toxicological studies remain scarce. We conducted an in vivo study to explore toxicological evidence in T-NO2 interaction on allergic asthma, to investigate underlying toxicological mechanisms. 90 male Balb/c mice were randomly and equally divided into 6 groups including saline control, ovalbumin (OVA)-sensitized, OVA + 35 °C, OVA + NO2, OVA + 35 °C + NO2, and OVA + 35 °C + NO2 + capsazepine (CZP), adopting treatment for 38 days. We measured pulmonary functions of inspiratory resistance (Ri), expiratory resistance (Re) and airway compliance (Cldyn), serum protein biomarkers, indexes of pulmonary inflammation, histopathological changes and protective effects of CZP. Airway hyperresponsiveness (AHR) was aggravated by high T (35 °C) and NO2 (5 ppm) co-exposure with a series of aggravating asthmatic symptoms including airway wall thickening, lumen stenosis, goblet cell proliferation, mucus hypersecretion, and subepithelial fibrotic hyperplasia, providing evidence in the toxicological impact of high T-NO2 interaction. The biomarkers of serum immune functions (Total-IgE, OVA-sIgE and IL-4), pro-inflammation (IL-6 and TNF-α), oxidative stress cytokines (8-OHdG, ROS and MDA), airway resistance (Ri and Re), and TRPV1 expression significantly increased, while IFN-γ, GSH and airway compliance (Cldyn) significantly decreased with co-exposure to high T and NO2. We observed that CZP addition significantly ameliorated these toxicological effects and biomarker levels induced by heat-NO2 interaction. Our results suggest a toxicity of heat-NO2 interaction on asthma with clear mechanisms, which can be ameliorated by CZP, indicating that both oxidative stress and TRPV1 expression may be primarily responsible for asthma of heat-NO2-induced toxicity.

Amphiregulin induces CCN2 and fibronectin expression by TGF-ß through EGFR-dependent pathway in lung epithelial cells.

BACKGROUND: Airway fibrosis is one of the pathological characteristics of severe asthma. Transforming growth factor (TGF)-ß has been known to promote epithelial-mesenchymal transition formation and to play a role in the progression of tissue fibrosis. Cellular communication network factor 2 (CCN2) and fibronectin (FN) are well-known markers of EMT and fibrosis. However, whether AREG is involved in TGF-ß-induced CCN2 and FN expression in human lung epithelial cells is unknown. METHODS: AREG and FN were analyzed by immunofluorescence staining on ovalbumin-challenged mice. CCN2 and FN expression were evaluated in human lung epithelial (A459) cells following TGF or AREG treatment for the indicated times. Secreted AREG from A549 cells was detected by ELISA. Cell migration was observed by a wound healing assay. Chromatin immunoprecipitation was used to detect the c-Jun binding to the CCN2 promoter. RESULTS: AREG and FN expression colocalized in lung tissues from mice with ovalbumin-induced asthma by immunofluorescence staining. Moreover, TGF-ß caused the release of AREG from A549 cells into the medium. Smad3 siRNA down-regulated AREG expression. AREG also stimulated CCN2 and FN expression, JNK and c-Jun phosphorylation, and cell migration in A549 cells. AREG small interfering (si) RNA inhibited TGF-ß-induced expression of CCN2, FN, and cell migration. Furthermore, AREG-induced CCN2 and FN expression were inhibited by EGFR siRNA, a JNK inhibitor (SP600125), and an activator protein-1 (AP-1) inhibitor (curcumin). EGFR siRNA attenuated AREG-induced JNK and c-Jun phosphorylation. Moreover, SP600125 downregulated AREG-induced c-Jun phosphorylation. CONCLUSION: These results suggested that AREG mediates the TGF-ß-induced EMT in human lung epithelial cells through EGFR/JNK/AP-1 activation. Understanding the role of AREG in the EMT could foster the development of therapeutic strategies for airway remodeling in severe asthma.

Branched-chain amino acid transaminase 1 inhibition attenuates childhood asthma in mice by effecting airway remodeling and autophagy.

Childhood asthma is a common chronic childhood disease. Branched-chain amino acid transaminase 1 (BCAT1) was reported to be upregulated in chronic airway diseases, while its role in childhood asthma is unclear. Asthma mouse models were established in neonatal mice by 10 µg ovalbumin (OVA) intraperitoneal injection and 3% OVA inhalational challenge. In OVA-challenged mice, BCAT1 levels were upregulated. BCAT1 inhibitor alleviated airway structure and inflammation by suppressing IgE, OVA-specific IgE and inflammatory cytokine release and inflammatory cell infiltration. BCAT1 inhibitor alleviated airway remodeling by inhibiting goblet cell hyperplasia, mucus secretion and the expression of α-SMA and collagen I/III. The BCAT1 inhibitor prevented OVA-enhanced autophagy by decreasing Beclin-1, Atg5 and LC3I/II and increasing p65 levels. In IL-13-stimulated BEAS-2B cells, rapamycin promoted inflammatory cytokine release and autophagy after BCAT1 inhibitor administration. Our research revealed that BCAT1 was upregulated in neonatal asthmatic mice and that a BCAT1 inhibitor might restrain airway inflammation and remodeling by decreasing autophagy, which offered a novel mechanistic understanding of childhood asthma.

Deficiency of leukocyte-specific protein 1 (LSP1) alleviates asthmatic inflammation in a mouse model.

BACKGROUND: Asthma is a major cause of morbidity and mortality in humans. The mechanisms of asthma are still not fully understood. Leukocyte-specific protein-1 (LSP-1) regulates neutrophil migration during acute lung inflammation. However, its role in asthma remains unknown. METHODS: An OVA-induced mouse asthma model in LSP1-deficient (Lsp1-/-) and wild-type (WT) 129/SvJ mice were used to test the hypothesis that the absence of LSP1 would inhibit airway hyperresponsiveness and lung inflammation. RESULTS: Light and electron microscopic immunocytochemistry and Western blotting showed that, compared with normal healthy lungs, the levels of LSP1 were increased in lungs of OVA-asthmatic mice. Compared to Lsp1-/- OVA mice, WT OVA mice had higher levels of leukocytes in broncho-alveolar lavage fluid and in the lung tissues (P < 0.05). The levels of OVA-specific IgE but not IgA and IgG1 in the serum of WT OVA mice was higher than that of Lsp1-/- OVA mice (P < 0.05). Deficiency of LSP1 significantly reduced the levels of IL-4, IL-5, IL-6, IL-13, and CXCL1 (P < 0.05) but not total proteins in broncho-alveolar lavage fluid in asthmatic mice. The airway hyper-responsiveness to methacholine in Lsp1-/- OVA mice was improved compared to WT OVA mice (P < 0.05). Histology revealed more inflammation (inflammatory cells, and airway and blood vessel wall thickening) in the lungs of WT OVA mice than in those of Lsp1-/- OVA mice. Finally, immunohistology showed localization of LSP1 protein in normal and asthmatic human lungs especially associated with the vascular endothelium and neutrophils. CONCLUSION: These data show that LSP1 deficiency reduces airway hyper-responsiveness and lung inflammation, including leukocyte recruitment and cytokine expression, in a mouse model of asthma.

Circular RNA has Circ 001372-Reduced Inflammation in Ovalbumin-Induced Asthma Through Sirt1/NFAT5 Signaling Pathway by miRNA-128-3p.

In this study, we sought to investigate the prospective role of circ 001372 in modifying inflammation in ovalbumin-induced asthma. In the vivo model of asthma, the serum of circ 001372 was reduced. Down-regulation of circ 001372 increased inflammation reaction (TNF-α, IL-1ß, IL-6, and IL-18) and induced COX-2 and iNOS protein expression in vitro model through activation of NFAT5 and suppression of Sirt1. Up-regulation of circ 001372 decreased inflammation reaction (TNF-α, IL-1ß, IL-6, and IL-18) in vitro model through inactivation of NFAT5 and induction of Sirt1 by miRNA-128-3p. The miRNA-128-3p lowered the effects of circ 001372 on inflammation in vitro model. The Sirt1 inhibitor reduced the effects of circ 001372 on inflammation in vitro model. Our results revealed the serum of circ 001372 against inflammation in ovalbumin-induced asthma through Sirt1/NFAT5 by miRNA-128-3p.

Fructus Amomi extract attenuates nasal inflammation by restoring Th1/Th2 balance and down-regulation of NF-κB phosphorylation in OVA-induced allergic rhinitis.

Fructus Amomi Cardamomi (FA) is the mature fruit of Amomum villosum Lour (family Zingiberaceae) and is commonly used in Chinese traditional medicine to treat various gastrointestinal disorders. FA's possible benefits as an allergic rhinitis (AR) treatment, however, have not been examined. We used an ovalbumin (OVA)-induced AR mouse model to identify any anti-allergic effects associated with the administration of 200 mg/kg FA or dexamethasone (Dex) 2.5 mg/kg by oral administration. The results of our testing confirm that FA ameliorated nasal symptoms and alleviated nasal epithelium swelling, reduced the goblet cell hyperplasia and eosinophil cell infiltration in the nasal epithelium, and inhibited lung tissue inflammation and Dex as well. Significantly decreased Th2 cytokine (interleukin (IL)-1ß, IL-4, and IL-5) expression, and a correspondingly significant increase in Th1 cytokine (IL-12, interferon (IFN)-γ) production, was observed in nasal lavage fluid (NALF) taken from mice that received FA or Dex treatment. FA also reduced the presence of OVA-specific immunoglobulin (Ig) E, OVA-specific IgG1, and histamine levels in serum, and inhibited mast cell degranulation in vitro. In addition, these effects were involved with the reduction in NF-κB phosphorylation. These results suggest that FA restores Th1/Th2 balance and inhibits NF-κB phosphorylation and mast cell degranulation, thereby achieving a notable anti-inflammatory effect. Accordingly, it has the potential to be used as an efficacious therapeutic treatment for AR.

Resveratrol Treatment Prevents Increase of Mast Cells in Both Murine OVA Enteritis and IL-10-/- Colitis.

Mast cells are involved in allergic and other inflammatory diseases. The polyphenol resveratrol is known for its anti-inflammatory properties and may be used as nutraceutical in mast cell associated diseases. We analyzed the effect of resveratrol on mast cells in vivo in ovalbumin-induced allergic enteritis as well as experimental colitis in IL-10-/- mice which received resveratrol via drinking water. Treatment with resveratrol prevented the increase in mast cells in both allergic enteritis and chronic colitis in duodenum as well as in colon. Further, it delayed the onset of diseases symptoms and ameliorated diseases associated parameters such as tissue damage as well as inflammatory cell infiltration in affected colon sections. In addition to the findings in vivo, resveratrol inhibited IgE-dependent degranulation and expression of pro-inflammatory cytokines such as TNF-α in IgE/DNP-activated as well as in LPS-activated bone marrow-derived mast cells. These results indicate that resveratrol may be considered as an anti-allergic and anti-inflammatory plant-derived component for the prevention or treatment of mast cell-associated disorders of the gastrointestinal tract.