OVA-induced Asthma Mouse Model Research Articles

Huatan Tongluo Decoction Inhibits Inflammatory Infiltration and Airway Remodeling by Attenuating TGF-β1/Smad2/3 and Oxidative Stress-mediated NF-kB/HIF-1α/MMPs Signaling Pathway in Chronic Asthma Mice

Background: Asthma is a common chronic respiratory disorder characterized by inflammation and remodeling of the airways. Aims: This study aimed to identify the inhibitory effects of Huatan Tongluo decoction (HTTLD) on airway inflammation and associated remodeling mechanisms. Methods: Mice were immunized with ovalbumin (OVA) for 8 weeks to generate chronic asthma mouse models (CAS), which were randomly divided into 4 groups administrated with pachyman, dexamethasone (DEX), HTTLD, and without anything (CAS model), while mice who administrated saline were assigned as the control group. Hematoxylin-eosin (H&E) and Masson trichrome were used to determine inflammatory infiltration and airway remodeling (fiber deposition). Inflammatory cytokines, including VEGF, PDGF, and TGF-β1, were analyzed using ELISA. The gene transcriptions and expressions of MMP-9, TIMP-1, VEGF, HIF-1α, NF-kB, and β-actin were evaluated using RT-PCR and Western blot, while the expression of p-Smad2/3 was determined by Western blot. Results: HTTLD inhibited inflammatory infiltration and airway remodeling (reducing airway wall thickness and decreasing fiber deposition) of lung tissues in the CAS mouse model. HTTLD markedly attenuated levels of TGF-β1, VEGF, and PDGF compared to those of mice in the CAS model group (p < 0.05). HTTLD significantly reduced the secretion of matrix metalloproteinases (MMP-9 and TIMP-1) and the expression of NF-kB/HIF-1α compared to mice in the CAS model group (p < 0.05). HTTLD prominently downregulated phosphorylated levels of the Smad2/3 molecule (ratio of p-Smad3/2/Smad2/3) compared to mice in the CAS group (p < 0.05). Conclusion: HTTLD inhibited inflammatory infiltration and airway remodeling in an OVA-induced chronic asthma mouse model by attenuating the TGF-β1/Smad2/3 signaling pathway and suppressing the oxidative stress-mediated NF-kB/HIF-1α/MMPs signaling pathway.

Toll-like receptor 4 mediated autophagy regulates airway smooth muscle cells behavior

Objectives Airway remodeling, a prominent feature of asthma, involves aberrant proliferation, apoptosis, and migration of airway smooth muscle cells (ASMCs). Toll-like receptors (TLRs) are implicated in the regulation of the autophagy pathway. In this study, we aimed to investigate the influence of Toll-like receptor 4 (TLR4) on autophagy and its underlying mechanism in ASMC proliferation, apoptosis, and migration. Methods Histopathological changes in the lungs of asthmatic mice assessed by Hematoxylin-Eosin (HE) and Masson staining. Cell proliferation, apoptosis and migration were evaluated utilizing CCK8, Edu, Flow cytometry and wound heading assays. The effectiveness of siRNA transfection and the expression of TLR4, autophagy, and proliferation-related proteins after siRNA treatment were examined through RT-PCR and Western blot (WB). Conclusion We observed an increase in TLR4 expression and autophagy in a mouse model of OVA-induced asthma. In vitro experiments showed that siRNA-mediated inhibition of TLR4 suppressed autophagy, proliferation, and migration of ASMCs, whereas TLR4 activation by lipopolysaccharide (LPS) had the opposite effect. Furthermore, the autophagy inhibitor 3-Methyladenine (3MA) inhibited ASMCs proliferation and migration while promoting apoptosis. Significantly, our study demonstrated that autophagy inhibition reversed the promotion effect of LPS on ASMC proliferation and migration. These findings suggest that TLR4 may modulate ASMC behavior through the regulation of autophagy.

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.

VEGF and EGFR signaling pathways are involved in the baicalein attenuation of OVA-induced airway inflammation and airway remodeling in mice

BackgroundAlthough Traditional Chinese Medicine (TCM) has been used for treating asthma for centuries, the understanding of its mechanism of action is still limited. Thus, the purpose of this study was to explore the possible therapeutic effects, and underlying mechanism of baicalein in the treatment of asthma.MethodsFreely availabled atabases (e.g. OMIM, TTD, Genecards, BATMAN-TCM, STITCH 5.0, SEA, SwissTargetPrediction) and software (e.g. Ligplot 2.2.5 and PyMoL) were used for disease drug target prediction and molecular docking by network pharmacology. The efficacy and mechanism of action of baicalein in the treatment of asthma were validated using an ovalbumin (OVA)-induced asthma mouse model and molecular biology techniques.ResultsA total of 1655 asthma-related genes and 161 baicalein-related targets were identified from public databases. Utilizing common databases and software for network pharmacology and molecular docking analysis, seven potential target proteins for the therapeutic effects of baicalein on asthma were selected, including v-akt murine thymoma viral oncogene homolog 1 (AKT1), vascular endothelial growth factor A (VEGFA), epidermal growth factor receptor (EGFR), proto-oncogene tyrosine-protein kinase Src (SRC), mitogen-activated protein kinase 3 (MAPK3), matrix metallopeptidase 9 (MMP9), and MAPK1. In vivo, baicalein treatment via intraperitoneal injection at a dose of 50 mg/kg significantly reduced airway inflammation, collagen deposition, smooth muscle thickness, lung interleukin (IL)-4 and IL-13 levels, peripheral blood immunoglobulin (Ig)E levels, as well as the count and ratio of eosinophils in bronchoalveolar lavage fluid (BALF) in an OVA-induced asthma mouse model. Further validation by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blotting analysis revealed that the VEGF and EGFR signaling pathways involving VEGFA, MAPK1, MAPK3, and EGFR were inhibited by baicalein in the asthma mouse model.ConclusionBaicalein attenuates airway inflammation and airway remodeling through inhibition of VEGF and EGFR signaling pathways in an OVA-induced asthma mouse model. This will provide a new basis for the development of baicalein as a treatment for asthma and highlights the potential of network pharmacology and molecular docking in drug discovery and development.

Lipocalin 2 (LCN2) Knockdown Regulates Treg/Th17 Balance to Improve Asthma in Mice.

Asthma substantially affects the quality of life and health of children. Lipocalin 2 (LCN2) is an immune-related protein, which is predicted to be highly expressed in asthma. Here, we investigated the role of LCN2 in ovalbumin (OVA)-induced asthma mouse model. We knocked down LCN2 in an asthma mouse model and performed histopathological analysis using hematoxylin and eosin (H&E) staining assay. Differentiated cells were assessed using Diff-Quick staining assay. We investigated the regulatory T (Treg) cell/ T helper 17 (Th17) cell balance using flow cytometry and enzyme-linked immunosorbent assay (ELISA). Inflammatory factors were measured using quantitative real-time reverse transcription PCR (qRT-PCR). The involved pathways were assessed using Western blotting. LCN2 was upregulated in patients with asthma. OVA promoted pathological deterioration in the lungs, increased IgE levels in the plasma, and elevated the number of differentiated inflammatory cells, whereas LCN2 knockdown abrogated the OVA-induced effects. Additionally, the Treg/Th17 imbalance and increased inflammatory cytokine levels were improved by LCN2 knockdown in OVA-treated mice. Moreover, LCN2 knockdown reversed the activation of the janus kinase (JNK) pathway. LCN2 knockdown improved the Treg/Th17 balance, alleviated inflammation, and inactivated the JNK pathway in OVA-induced asthma mouse model, suggesting that LCN2 may be a novel therapeutic target for asthma in children.

Expression of constitutively active TβRI leads to attenuation of ovalbumin-induced allergic airway inflammation associated with augmented M2 polarization of alveolar macrophage

BackgroundTransforming growth factor-β (Tgf-β) plays an important role in the pathogenesis of asthma through the regulation of T cells and airway epithelium. Its functions in alveolar macrophage (AM) during allergic airway inflammation remain unknown. MethodsA murine asthma model was induced with ovalbumin (ova) in TβRICA/Fsp1-Cre transgenic mice expressing constitutively active Tgf-β receptor type I (TβRICA) under the control of Fsp1-Cre transgene. Cells in the bronchoalveolar lavage (BAL) were collected to study immune cell infiltration in the lungs. Cytokine levels in BAL fluid were measured by enzyme-linked immunoassay (ELISA). Lungs were sectioned and stained with hematoxylin and eosin, periodic acid-Schiff, and trichrome for histopathologic evaluation. AMs were assessed by flow cytometry and were sorted for quantitative polymerase chain reaction analysis. ResultsOur data indicated that TβRICA transcripts were induced in AMs of TβRICA/Fsp1-Cre mice. Following the ova challenges, TβRICA/Fsp1-Cre mice exhibited reduced cellular infiltration of the airway, reduced pulmonary fibrosis, and reduced bronchial mucus secretion as compared to ova-challenged wild-type mice. An alternatively activated macrophage (M2) polarization was significantly elevated in the lungs of ova-challenged TβRICA/Fsp1-Cre mice as reflected by increased numbers of AMs expressing M2 subtype marker, CD163, in the lungs and enhanced expression of CCR2 and CD206 in AMs. Moreover, TβRICA/Fsp1-Cre AMs showed augmented expression of transcription factors, Foxo1, and IRF4, which are known to be positive regulators for M2 polarization. ConclusionsExpression of TβRICA in AMs promoted M2 polarization and ameliorated allergic airway inflammation in an ova-induced asthma mouse model.

Corilagin attenuates airway inflammation and collagen deposition in ovalbumin-induced asthmatic mice

Objective: To investigate the effects of corilagin on inflammation and collagen deposition in ovalbumin (OVA)-induced asthma mouse model and uncover the mechanism. Methods: We constructed a mouse model of OVA-induced asthma. Enzyme-linked-immunosorbent serologic assays were conducted to detect the effects of corilagin on cytokines and Immunoglobulin E (IgE) production. Hematoxylin and eosin staining was used to show pathological features in lung tissues. Masson trichrome assay was used to examine collagen deposition. In addition, the lung function was detected by mouse lung function apparatus. Immunoblot was used to confirm the mechanism. Results: Corilagin alleviates OVA-induced cytokine and IgE production. In addition, corilagin alleviates OVA-induced pathological changes and collagen deposition in lung tissues. Corilagin also suppressed airway resistance and lung function in mice. Mechanically, corilagin activated the adenosine monophosphate-activated protein kinase (AMPK) pathway in lung tissues. Conclusion: Corilagin attenuates airway inflammation and collagen deposition in OVA-induced asthmatic mice via AMPK pathway.

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.

Phaeanthus vietnamensis Ban Ameliorates Lower Airway Inflammation in Experimental Asthmatic Mouse Model via Nrf2/HO-1 and MAPK Signaling Pathway.

Asthma is a chronic airway inflammatory disease listed as one of the top global health problems. Phaeanthus vietnamensis BÂN is a well-known medicinal plant in Vietnam with its anti-oxidant, anti-microbial, anti-inflammatory potential, and gastro-protective properties. However, there is no study about P. vietnamensis extract (PVE) on asthma disease. Here, an OVA-induced asthma mouse model was established to evaluate the anti-inflammatory and anti-asthmatic effects and possible mechanisms of PVE. BALB/c mice were sensitized by injecting 50 μg OVA into the peritoneal and challenged by nebulization with 5% OVA. Mice were orally administered various doses of PVE once daily (50, 100, 200 mg/kg) or dexamethasone (Dex; 2.5 mg/kg) or Saline 1 h before the OVA challenge. The cell infiltrated in the bronchoalveolar lavage fluid (BALF) was analyzed; levels of OVA-specific immunoglobulins in serum, cytokines, and transcription factors in the BALF were measured, and lung histopathology was evaluated. PVE, especially PVE 200mg/kg dose, could improve asthma exacerbation by balancing the Th1/Th2 ratio, reducing inflammatory cells in BALF, depressing serum anti-specific OVA IgE, anti-specific OVA IgG1, histamine levels, and retrieving lung histology. Moreover, the PVE treatment group significantly increased the expressions of antioxidant enzymes Nrf2 and HO-1 in the lung tissue and the level of those antioxidant enzymes in the BALF, decreasing the oxidative stress marker MDA level in the BALF, leading to the relieving the activation of MAPK signaling in asthmatic condition. The present study demonstrated that Phaeanthus vietnamensis BÂN, traditionally used in Vietnam as a medicinal plant, may be used as an efficacious agent for treating asthmatic disease.

Corilagin attenuates airway inflammation and collagen deposition in ovalbumin-induced asthmatic mice.

To investigate the effects of corilagin on inflammation and collagen deposition in ovalbumin (OVA)-induced asthma mouse model and uncover the mechanism. We constructed a mouse model of OVA-induced asthma. Enzyme-linked-immunosorbent serologic assays were conducted to detect the effects of corilagin on cytokines and Immunoglobulin E (IgE) production. Hematoxylin and eosin staining was used to show pathological features in lung tissues. Masson trichrome assay was used to examine collagen deposition. In addition, the lung function was detected by mouse lung function apparatus. Immunoblot was used to confirm the mechanism. Corilagin alleviates OVA-induced cytokine and IgE production. In addition, corilagin alleviates OVA-induced pathological changes and collagen deposition in lung tissues. Corilagin also suppressed airway resistance and lung function in mice. Mechanically, corilagin activated the adenosine monophosphate-activated protein kinase (AMPK) pathway in lung tissues. Corilagin attenuates airway inflammation and collagen deposition in OVA-induced asthmatic mice via AMPK pathway.

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.

Lactoferrin Ameliorates Ovalbumin-Induced Asthma in Mice through Reducing Dendritic-Cell-Derived Th2 Cell Responses.

Asthma is a chronic respiratory disease with symptoms such as expiratory airflow narrowing and airway hyperresponsiveness (AHR). Millions of people suffer from asthma and are at risk of life-threatening conditions. Lactoferrin (LF) is a glycoprotein with multiple physiological functions, including antioxidant, anti-inflammatory, antimicrobial, and antitumoral activities. LF has been shown to function in immunoregulatory activities in ovalbumin (OVA)-induced delayed type hypersensitivity (DTH) in mice. Hence, the purpose of this study was to investigate the roles of LF in AHR and the functions of dendritic cells (DCs) and Th2-related responses in asthma. Twenty 8-week-old male BALB/c mice were divided into normal control (NC), ovalbumin (OVA)-sensitized, and OVA-sensitized with low dose of LF (100 mg/kg) or high dose of LF (300 mg/kg) treatment groups. The mice were challenged by intranasal instillation with 5% OVA on the 21st to 27th day after the start of the sensitization period. The AHR, cytokines in bronchoalveolar lavage fluid, and pulmonary histology of each mouse were measured. Serum OVA-specific IgE and IgG1 and OVA-specific splenocyte responses were further detected. The results showed that LF exhibited protective effects in ameliorating AHR, as well as lung inflammation and damage, in reducing the expression of Th2 cytokines and the secretion of allergen-specific antibodies, in influencing the functions of DCs, and in decreasing the level of Th2 immune responses in a BALB/c mouse model of OVA-induced allergic asthma. Importantly, we demonstrated that LF has practical application in reducing DC-induced Th2 cell responses in asthma. In conclusion, LF exhibits anti-inflammation and immunoregulation activities in OVA-induced allergic asthma. These results suggest that LF may act as a supplement to prevent asthma-induced lung injury and provide an additional agent for reducing asthma severity.

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

BackgroundAsthma 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.MethodsAn 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.ResultsLight 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.ConclusionThese data show that LSP1 deficiency reduces airway hyper-responsiveness and lung inflammation, including leukocyte recruitment and cytokine expression, in a mouse model of asthma.

Blockade of NLRP3/Caspase-1/IL-1β Regulated Th17/Treg Immune Imbalance and Attenuated the Neutrophilic Airway Inflammation in an Ovalbumin-Induced Murine Model of Asthma

Asthma is a heterogeneous inflammatory disorder of the airways, and multiple studies have addressed the vital role of the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3)/caspase-1/interleukin-1β (IL-1β) pathway in asthma, but its impact on ovalbumin- (OVA-) induced neutrophilic asthma remains unclear. Here, we explored this pathway's effect on airway inflammation in neutrophilic asthma to clarify whether blocking this signaling could alleviate asthmatic airway inflammation. Using an established OVA-induced neutrophilic asthma mouse model, we provided asthmatic mice with a highly selective NLRP3 inhibitor, MCC950, and a specific caspase-1 inhibitor, Ac-YVAD-cmk. Our results indicated that asthmatic mice exhibited increased airway hyperresponsiveness, neutrophil infiltration, and airway mucus hypersecretion, upregulated retinoid-related orphan receptor-γt (RORγt) mRNA expression, and downregulated fork head box p3 (Foxp3) mRNA expression, which was concurrent with NLRP3 inflammasome activation and upregulation of caspase-1, IL-1β, and IL-18 expression in lung. Treatment of NLRP3 inflammasome inhibitors significantly attenuated airway hyperresponsiveness, airway inflammation, and reversed T helper 17 (Th17)/regulatory T (Treg) cell imbalance in asthmatic mice. We propose that the NLRP3/caspase-1/IL-1β pathway plays an important role in the pathological process of neutrophilic asthma and provides evidence that blocking this pathway could potentially be a treatment strategy to ameliorate airway inflammation in asthma after validation with future experimental and clinical studies.

Dual Effect of Low-Molecular-Weight Bioregulators of Bacterial Origin in Experimental Model of Asthma.

Asthma is one of the most common noncommunicable diseases, affecting over 200 million people. A large number of drugs control asthma attacks, but there is no effective therapy. Identification of reasons for asthma and preventing this disease is a relevant task. The influence of bacterial components is necessary for the normal development of the immune system and the formation of an adequate immune response to antigens. In the absence of microorganisms or their insufficient exposure, the prerequisites are formed for excessive reactivity to harmless antigens. In the present study, we analyzed cellular and humoral factors in a standard mouse model of OVA-induced asthma modified by 5-fold intraperitoneal injection of bacterial cell wall fragments of glucosaminylmuramyl dipeptide (GMDP) 5 μg/animal or 1 μg lipopolysaccharide (LPS) per animal for 5 days before sensitization by ovalbumin (OVA). Preliminary administration of LPS or GMDP to animals significantly reduced goblet cells as well as the number of neutrophils, lymphocytes, and eosinophils in bronchoalveolar lavage, wherein GMDP corrected neutrophilia to a 2-fold degree, and LPS reduced the severity of eosinophilia by 1.9 times. With OVA administration of GMDP or LPS at the sensitization stage, an increase in the total number of bronchoalveolar lavage cells due to neutrophils, macrophages, lymphocytes, and eosinophils in relation to the group with asthma without GMDP or LPS was observed. The administration of GMDP or LPS to normal mice without asthma for 5 days had no statistically significant effect on the change in the number and population composition of cells in bronchoalveolar lavage in comparison with the control group receiving PBS. As a result of a study in a mouse model of asthma, a dual effect of LPS and GMDP was established: the introduction of LPS or GMDP before sensitization reduces neutrophilia and eosinophilia, while the introduction of LPS or GMDP together with an allergen significantly increases neutrophilia and eosinophilia. The study of the immunoglobulin status shows that in normal-asthma mice, GMDP and LPS slightly increase IgA in bronchoalveolar lavage; at the same time, in the asthma model, injections of GMDP or LPS before sensitization contribute to a significant decrease in IgA (2.6 times and 2.1 times, respectively) in BALF and IgE (2.2 times and 2.0 times, respectively) in blood serum. In an experimental model of asthma, the effect of GMDP and LPS was multidirectional: when they are repeatedly administered before sensitization, the bacterial components significantly reduce the severity of the allergic process, while in the case of a joint injection with an allergen, they increase the influx of macrophages, lymphocytes, and neutrophils into the lungs, which can aggravate the course of pathological process. Thus, the insufficient effect of antigens of a bacterial nature, in particular, with prolonged use of antibiotics can be compensated for by substances based on low-molecular-weight bioregulators of bacterial origin to establish the missing signals for innate immunity receptors, whose constant activation at a certain level is necessary to maintain homeostasis.

Allergen-Specific Treg Cells Upregulated by Lung-Stage S. japonicum Infection Alleviates Allergic Airway Inflammation.

Schistosoma japonicum infection showed protective effects against allergic airway inflammation (AAI). However, controversial findings exist especially regarding the timing of the helminth infection and the underlying mechanisms. Most previous studies focused on understanding the preventive effect of S. japonicum infection on asthma (infection before allergen sensitization), whereas the protective effects of S. japonicum infection (allergen sensitization before infection) on asthma were rarely investigated. In this study, we investigated the protective effects of S. japonicum infection on AAI using a mouse model of OVA-induced asthma. To explore how the timing of S. japonicum infection influences its protective effect, the mice were percutaneously infected with cercaria of S. japonicum at either 1 day (infection at lung-stage during AAI) or 14 days before ovalbumin (OVA) challenge (infection at post–lung-stage during AAI). We found that lung-stage S. japonicum infection significantly ameliorated OVA-induced AAI, whereas post–lung-stage infection did not. Mechanistically, lung-stage S. japonicum infection significantly upregulated the frequency of regulatory T cells (Treg cells), especially OVA-specific Treg cells, in lung tissue, which negatively correlated with the level of OVA-specific immunoglobulin E (IgE). Depletion of Treg cells in vivo partially counteracted the protective effect of lung-stage S. japonicum infection on asthma. Furthermore, transcriptomic analysis of lung tissue showed that lung-stage S. japonicum infection during AAI shaped the microenvironment to favor Treg induction. In conclusion, our data showed that lung-stage S. japonicum infection could relieve OVA-induced asthma in a mouse model. The protective effect was mediated by the upregulated OVA-specific Treg cells, which suppressed IgE production. Our results may facilitate the discovery of a novel therapy for AAI.

Atractylon Treatment Attenuates Pulmonary Fibrosis via Regulation of the mmu_circ_0000981/miR-211-5p/TGFBR2 Axis in an Ovalbumin-Induced Asthma Mouse Model.

Asthma-induced pulmonary fibrosis (PF) is an important public health concern that has few treatment options given its poorly understood etiology; however, the epithelial to mesenchymal transition (EMT) of pulmonary epithelial cells has been implicated to play an important role in inducing PF. Although previous studies have found atractylon (Atr) to have anti-inflammatory effects, whether Atr has anti-PF abilities remains unknown. The purpose of the current study was to validate the protective efficiency of Atr in both an animal model of ovalbumin (OVA)-induced asthma and an EMT model induced by transforming growth factor-β1 (TGF-β1) using TC-1 cells. The results of this study revealed that Atr treatment suppressed OVA-induced PF via fibrosis-related protein expression. Atr treatment suppressed OVA-induced circRNA-0000981 and TGFBR2 expression but promoted miR-211-5p expression. In vivo studies revealed that Atr suppressed TGF-β1-induced EMT and fibrosis-related protein expression via suppressing circRNA-0000981 and TGFBR2 expression. The results also suggested that the downregulation of circRNA-0000981 expression suppressed TGFBR2 by sponging miR-211-5p, which was validated by a luciferase reporter assay. Collectively, the findings of the present study suggest that Atr treatment attenuates PF by regulating the mmu_circ_0000981/miR-211-5p/TGFBR2 axis in an OVA-induced asthma mouse model.

The Protective Effects of Maresin 1 in the OVA-Induced Asthma Mouse Model.

Asthma is a chronic inflammatory disease that cannot be cured. Maresin 1 (MaR1) is a specific lipid synthesized by macrophages that exhibits powerful anti-inflammatory effects in various inflammatory diseases. The goal of this study was to evaluate the effect of MaR1 on allergic asthma using an ovalbumin- (OVA-) induced asthma model. Thirty BALB/c mice were randomly allocated to control, OVA, and MaR1 + OVA groups. Mice were sacrificed 24 hours after the end of the last challenge, and serum, bronchoalveolar lavage fluid (BALF), and lung tissue were collected for further analysis. Western blotting was used to measure the protein level of IκBα, the activation of the NF-κB signaling pathway, and the expression of NF-κB downstream inflammatory cytokines. Quantitative real-time polymerase chain reactions (qRT-PCRs) were used to evaluate the expression levels of COX-2 and ICAM-1 in lung tissues. We found that high doses of MaR1 were most effective in preventing OVA-induced inflammatory cell infiltration and excessive mucus production in lung tissue, reducing the number of inflammatory cells in the BALF and inhibiting the expression of serum or BALF-associated inflammatory factors. Furthermore, high-dose MaR1 treatment markedly suppressed the activation of the NF-κB signaling pathway, the degradation of IκBα, and the expression of inflammatory genes downstream of NF-κB, such as COX-2 and ICAM-1, in the OVA-induced asthma mouse model. Our findings indicate that MaR1 may play a critical role in OVA-induced asthma and may be therapeutically useful for the management of asthma.

TMT-based quantitative proteomics reveals suppression of SLC3A2 and ATP1A3 expression contributes to the inhibitory role of acupuncture on airway inflammation in an OVA-induced mouse asthma model

Asthma is a chronic airway inflammatory disease and acupuncture is frequently used in patients suffering from asthma in clinic. However, the regulatory mechanism of acupuncture treatment in asthma is not fully elucidated. We sought to investigate the effectiveness of acupuncture on asthma and the associated regulatory mechanism. An ovalbumin (OVA)-induced mouse asthma model was established and the effect of acupuncture on airway hyperresponsiveness (AHR), mucus hypersecretion and inflammation was assessed. Tandem mass tag (TMT)-based quantitative proteomics analysis of lung tissue and bioinformatics analysis were performed. Our results revealed that the OVA-induced mouse asthma model was successfully established with the significantly elevated AHR to methacholine (Mch), and acupuncture was effective in attenuation of AHR to Mch, peribronchial and perivascular inflammation and mucus production. The inflammatory cells around the airways, mucous secretion as well as levels of IgE, CCL5, CCL11, IL-17A in bronchoalveolar lavage fluid (BALF) and IL-4, IL-5 and IL-13 levels in serum were siginificantly inhibited by acupuncture. TMT-based quantitative proteomics analysis found that a total of 6078 quantifiable proteins were identified, and 564 (334 up-regulated and 230 down regulated) differentially expressed proteins (DEPs) were identified in OVA-induced asthma model group (A) versus normal control group (NC). Acupuncture treatment resulted in 667 DEPs (416 up-regulated and 251 down regulated) compared with A group, and 86 overlapping DEPs were identified in NC, A and AA groups. Among the 86 overlapping DEPs, we identified 41 DEPs regulated by acupuncture. Based on the above data, we performed a systematic bioinformatics analysis of the 41 DEPs, and results showed that these 41 DEPs were predominantly related to 4 KEGG pathways including SNARE interactions in vesicular transport, ferroptosis, endocrine and other factor-regulated calcium reabsorption, and protein digestion and absorption. DEPs of SLC3A2 and ATP1A3 expression levels were verified by immumohistochemical staining. Mice in OVA-induced asthma model group had elevated SLC3A2 and ATP1A3 expression and acupuncture had the ability to downregulate SLC3A2 and ATP1A3 protein expression. Furthermore, acupuncture reduced the MDA level and increased the GSH and SOD levels in the lung tissue. Taken together, our data suggested that acupuncture was effective in treating asthma by attenuation of AHR, mucus secretion and airway inflammation, and the mechanism was associated with regulation of ferroptosis, SLC3A2 and ATP1A3 protein expression as well as oxidative stress. Results from our experiments revealed the anti-inflammatory effect of acupuncture in OVA-induced mouse asthma model, leading to a more effective approach to be chosen by patients in clinic.

Multiple Components Rapidly Screened from Perilla Leaves Attenuate Asthma Airway Inflammation by Synergistic Targeting on Syk.

BackgroundPerilla frutescens (L.) Britt., a classic medicinal plant, has been demonstrated to have anti-inflammatory and anti-allergic effects in asthma. Perilla leaves extract (PLE) exerted significant therapeutic effect against allergic asthma inflammation through Syk inhibition. But the active chemical ingredients from PLE are complex and unclear, it is difficult to fully elucidate its pharmacological mechanisms.MethodsA method was established for rapid screening and characterization of active ingredients from PLE that targeted Syk, with which three potential active ingredients were identified. By using OVA-induced allergic asthma mouse model in vivo, OVA-induced human PBMCs inflammation model and DNP-IgE/BSA-induced RBL-2H3 cells model in vitro, the effects and mechanisms of PLE and its active components were evaluated.ResultsUsing Syk-affinity screening method, roseoside (RosS), vicenin-2 (Vic-2) and rosmarinic acid (RosA) were identified from PLE. In vitro, PLE and its ingredients showed significant inhibitory activities against Syk, with their mixture (Mix, prepared by RosS, Vic-2 and RosA in accordance with their ratio in Syk-conjugated beads bound fraction) showing a stronger inhibitory activity. RosS, Vic-2 and RosA also showed significant effects on allergic asthma, and a synergistic effect of Mix was observed. Moreover, treatment with PLE, RosS, Vic-2, RosA, and Mix significantly inhibited the expression and phosphorylation of Syk, PKC, NF-κB p65, and cPLA2 in allergic mice lung tissue and in RBL-2H3 cells.ConclusionPLE may alleviate allergic airway inflammation partly through the multiple components synergistic targeting on Syk and its downstream inflammatory pathway.