Human adenovirus type 4 (HAdV-4) associated acute respiratory tract infection in children & genetic characteristics of HAdV-4 in China: a prospective multicenter study.

BACKGROUND: Human adenovirus (HAdV) is an important pathogen causing acute respiratory infection (ARI) in children. Many countries, including China, have experienced sporadic or outbreaks related to HAdV-4, and death cases were reported. However, there is little research on HAdV-4 and the epidemic situation of HAdV-4 in China is little known. This study was designed to comprehend the prevalence and genetic characteristics of HAdV-4 in ARI children in China. METHODS: Respiratory tract samples from ARI children hospitalized in six hospitals of Northern and Southern China from 2017 to 2020 were collected for HAdV detection and typing. Clinical information was collected from HAdV-4 positive patients for clinical characteristics and epidemiological analysis. The main capsid proteins and the whole genome sequences were amplified and sequenced for bioinformatics analysis. RESULTS: There were 2847 ARI children enrolled, and 156 (5.48%) HAdV positive samples were detected. Eleven HAdV-4 positive samples were identified, accounting for 0.39% of the total samples and 7.05% of the HAdV positive samples. The main manifestations were fever and cough. Two children had conjunctivitis. Two children were diagnosed with severe pneumonia and developed respiratory failure. One of them developed hemophagocytic syndrome and checked in pediatric intensive care unit (PICU). This child had ventricular septal defect. All the children recovered. The isolated strains of HAdV-4 obtained in this study and the reference strains from China located in the same phylogenetic branch (HAdV-4a), while the prototype strain and vaccine strains formed another branch (HAdV-4p). Upon comparison with the prototype strain, there were a few amino acid mutations existing in three major capsid proteins. According to recombination analysis, no new recombination was found. CONCLUSIONS: The detection rate of HAdV-4 in children hospitalized with ARI was 0.39% in the total samples and 7.05% of all HAdV positive samples. HAdV-4 isolates obtained in this study and other reference strains from China belonged to the HAdV-4a subtype. Our data provided reference for the monitoring, prevention and control of HAdV-4, as well as the research and development of vaccines and drugs.

Radical S-adenosyl methionine domain-containing 2, a potential target of D-tryptophan in asthma treatment, regulates T helper cell type 1/2 balance.

Asthma is a common chronic respiratory disease. D-tryptophan (D-TRP) can inhibit allergic airway inflammation and T helper cell type 2 (Th2) immune response. RNA-sequencing results have indicated that radical S-adenosyl methionine domain-containing 2 (RSAD2) might be a potential molecular target of D-TRP in asthma treatment. Herein, we established a mouse model of asthma using ovalbumin (OVA) via intraperitoneal injection and inhalational challenge. Gain- and loss-of-function studies of RSAD2 were performed in mice following the intratracheal delivery of lentiviral vectors (3 × 106 TU/mL). Naïve CD-4+ T cells were isolated from the spleen and used to explore the effects of RSAD2 on Th2 cell differentiation. RSAD2 expression was higher in the asthma group than in the control group. RSAD2 knockdown alleviated inflammatory cell infiltration and reduced the number of goblet cells. Low RSAD2 expression decreased the levels of IgE, IL-25, IL-33, and TSLP, and it reduced the number of inflammatory cells in the bronchoalveolar lavage fluid. RSAD2 silencing suppressed Th2-related cytokine levels (such as IL-4, IL-5, and IL-13) and increased Th1-related cytokine levels (such as IFN-γ). Additionally, RSAD2 knockdown inhibited the phosphorylation of JAK1, JAK3, and STAT6, and downregulated GATA-3 expression. RSAD2 overexpression increased inflammatory cell infiltration and mucus secretion in the lung tissues of mice pretreated with D-TRP. D-TRP pretreatment reduced OVA-specific IgE content and IL-4 and IL-5 levels, and it increased the IFN-γ levels; however, RSAD2 overexpression reversed these effects. In conclusion, RSAD2 knockdown can mitigate OVA-induced asthma by regulating the Th2 immune response via JAK/STAT6 pathway inhibition.

Growth differentiation factor 15 protects the airway by inhibiting cell pyroptosis in obese asthmatic mice through the phosphoinositide 3-kinase/AKT pathway.

Obese asthma is a form of refractory asthma with inflammation as the underlying mechanism. The specific mechanism of action of anti-inflammatory growth differentiation factor 15 (GDF15) in obese asthma is unclear. The purpose of this study was to explore the effect of GDF15 on cell pyroptosis in obese asthma and to determine its mechanism of airway protection. Male C57BL6/J mice were fed with a high-fat diet, sensitized, and challenged with ovalbumin. Recombinant human (rh)GDF15 was administered 1 h before the challenge. GDF15 treatment significantly reduced airway inflammatory cell infiltration, mucus hypersecretion and airway resistant, and decreased cell counts and inflammatory factors in bronchoalveolar lavage fluid. Serum inflammatory factors decreased, and the increased levels of NLR family pyrin domain containing 3 (NLRP3), caspase-1, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), and gasdermin-D (GSDMD-N) in obese asthmatic mice were inhibited. Furthermore, the suppressed phosphoinositide 3-kinase (PI3K)/AKT signal pathway was activated after rhGDF15 treatment. The same result was obtained by overexpression of GDF15 in human bronchial epithelial cells induced by lipopolysaccharide (LPS) in vitro, and the effect of GDF15 was reversed after the application of a PI3K pathway inhibitor. Thus, GDF15 could protect the airway by inhibiting cell pyroptosis in obese asthmatic mice through the PI3K/AKT signaling pathway.

A multicentre study on the incidence of respiratory viruses in children with community-acquired pneumonia requiring hospitalization in the setting of the zero-COVID policy in China.

BACKGROUND: Stringent nonpharmaceutical interventions (NPIs) have been implemented worldwide to combat the COVID-19 pandemic, and the circulation and seasonality of common respiratory viruses have subsequently changed. There have been few multicentre studies or comparisons of the prevalence of respiratory viruses accounting for community-acquired pneumonia (CAP) in hospitalized children between the pre-COVID period and the period after community and school reopening in the setting of the zero-COVID policy. METHODS: We included 1543 children with CAP who required hospitalization from November 1, 2020 to April 30, 2021 (period 1), and 629 children with the same conditions from November 1, 2018, to April 30, 2019 (period 2), in our study. All respiratory samples from these patients were screened for six respiratory viruses (respiratory syncytial virus [RSV], adenovirus [ADV], influenza A virus [Flu A], influenza B virus [Flu B], parainfluenza virus type 1 [PIV1], and parainfluenza virus type 3 [PIV3]) using a multiplex real-time PCR assay. RESULTS AND CONCLUSIONS: The median ages of the enrolled patients at the time of diagnosis were 1.5 years and 1.0 years for period 1 and period 2, respectively. In period 1, viral pathogens were detected in 50.3% (776/1543) of the enrolled patients. The most frequently identified viral pathogen was RSV (35.9%, 554/1543), followed by PIV3 (9.6%, 148/1543), PIV1 (3.6%, 56/1543), ADV (3.4%, 52/1543), Flu A (1.0%, 16/1543), and Flu B (0.8%, 13/1543). The total detection rates of these six viruses in the peak season of CAP were at the pre-COVID level. The prevalence of Flu A decreased dramatically, and circulation activity was low compared to pre-COVID levels, while the incidence of PIV3 increased significantly. There were no significant differences in the detection rates of RSV, ADV, Flu B, and PIV1 between the two periods. Our results showed that respiratory viruses accounted for CAP in hospitalized children at pre-COVID levels as communities and schools reopened within the zero-COVID policy, although the prevalence aetiology spectrum varied.

Ferrostatin-1 and 3-Methyladenine Ameliorate Ferroptosis in OVA-Induced Asthma Model and in IL-13-Challenged BEAS-2B Cells.

Ferroptosis was reported to be involved in the occurrence and development of asthma. However, the potential mechanism underlying the role of ferroptosis in asthma remains unclear. In this study, we established the mouse asthma model following the ovalbumin (OVA) method in C57BL/6 mice and the cell model with IL-13 induction in bronchial epithelial cells (BEAS-2B cells). Treatment of ferrostatin-1 (Ferr-1) and 3-methyladenine (3-MA) decreased iron deposition in IL-13-induced BEAS-2B cells and lung tissues of asthma mice, opposite to that in bronchoalveolar lavage fluid (BALF). Meanwhile, excessive lipid peroxidation asthma model in vivo and in vitro was alleviated by Ferr-1 or 3-MA treatment. In addition, Ferr-1 and 3-MA inhibited the expression of LC-3 in these cells and lung tissues of mice. Moreover, Ferr-1 and 3-MA also suppressed the production of inflammatory cytokines (IL-1ß, IL-6, and TNF-α) and oxidative stress factors (ROS and MDA), while promoting the level of SOD, in vivo and in vitro. Furthermore, application of Ferr-1 exhibited a greater inhibitory effect on iron release and lipid peroxidation in IL-13-induced BEAS-2B cells and asthma mice than 3-MA, accompanied with a weaker effect on ferritinophagy than 3-MA. Collectively, Ferr-1 and 3-MA ameliorated asthma in vivo and in vitro through inhibiting ferroptosis, providing a new strategy for the clinical treatment of asthma.

Human bone marrow-mesenchymal stem cell-derived exosomal microRNA-188 reduces bronchial smooth muscle cell proliferation in asthma through suppressing the JARID2/Wnt/ß-catenin axis.

The functions of exosomes in allergic diseases including asthma have aroused increasing concerns. This paper focuses on the effects of exosomes derived from human bone marrow-mesenchymal stem cells (hBM-MSCs) on the proliferation of bronchial smooth muscle cells in asthma and the mechanism involved. Exosomes were extracted from hBM-MSCs and identified. Human BSMCs were induced with transforming growth factor (TGF)-ß1 to mimic an asthma-like condition in vitro and then treated with exosomes. A mouse model with asthma was induced by ovalbumin (OVA) and treated with exosomes for in vivo study. The hBM-MSC-derived exosomes significantly reduced the abnormal proliferation and migration of TGF-ß1-treated BSMCs. microRNA (miR)-188 was the most enriched miRNA in exosomes according the microarray analysis, and JARID2 was identified as a mRNA target of miR-188. Either downregulation of miR-188 or upregulation of JARID2 blocked the protective effects of exosomes on BSMCs. JARID2 activated the Wnt/ß-catenin signaling pathway. In the asthmatic mice, hBM-MSC-derived exosomes reduced inflammatory cell infiltration, mucus production, and collagen deposition in mouse lung tissues. In conclusion, this study suggestes that hBM-MSC-derived exosomes suppress proliferation of BSMCs and lung injury in asthmatic mice through the miR-188/JARID2/Wnt/ß-catenin axis. This study may provide novel insights into asthma management.

Long noncoding RNA PTTG3P/miR-192-3p/CCNB1 axis is a potential biomarker of childhood asthma.

BACKGROUND: Increasing evidence suggests that long non-coding RNAs (lncRNAs) affect the regulation of immune responses, airway inflammation, and other pathological processes involved in asthma. LncRNA PTTG3P is associated with the development of various tumors, but its role in childhood asthma remains unknown. In this study, we investigated the functions of the lncRNA PTTG3P in the progression of childhood asthma. METHODS: Twenty-six healthy children and 26 asthmatic children were monitored for disease progression for 2 years. We obtained blood samples during the chronic phase of disease for lncRNA/mRNA expression microarray analysis. A competitive endogenous RNA network (PTTG3P/miR-192-3p/CCNB1) was identified using bioinformatics analyses. Real-time qPCR and western blot were used to quantify gene and protein expression levels, respectively. Cell counting kit­8 and transwell assays were used to evaluate the proliferation and migration of bronchial epithelial (16HBE) cells. Double luciferase reporter gene assay was used to validate the predictive targets in PTTG3P, miR-192-3p, and CCNB1. RESULTS: PTTG3P was highly expressed in the peripheral blood of asthmatic children. Knocking down PTTG3P inhibited epithelial-mesenchymal transition, proliferation, and migration of 16HBE cells. PTTG3P promoted progression of childhood asthma by targeting the miR-192-3p/CCNB1 axis. CONCLUSIONS: Childhood asthma was associated with the PTTG3P/miR-192-3p/CCNB1 axis. This study provides potential diagnostic and treatment biomarkers for childhood asthma.

A Multicenter Study of Viral Aetiology of Community-Acquired Pneumonia in Hospitalized Children in Chinese Mainland.

Community-acquired pneumonia (CAP) is one of the leading causes of morbidity and mortality in children worldwide. In this study, we aimed to describe the aetiology of viral infection of pediatric CAP in Chinese mainland. During November 2014 to June 2016, the prospective study was conducted in 13 hospitals. The hospitalized children under 18 years old who met the criteria for CAP were enrolled. The throat swabs or nasopharyngeal aspirates (NPAs) were collected which were then screened 18 respiratory viruses using multiplex PCR assay. Viral pathogens were present in 56.6% (1539/2721) of the enrolled cases, with the detection rate of single virus in 39.8% of the cases and multiple viruses in 16.8% of the cases. The most frequently detected virus was respiratory syncytial virus (RSV) (15.2%, 414/2721). The highest detection rate of virus was in < 6-month-age group (70.7%, 292/413). RSV, human metapneumovirus (HMPV), human parainfluenza viruses (HPIVs) and influenza B virus (Flu B) showed the similar prevalence patterns both in north and south China, but HPIVs, Flu A, human bocavirus (HBoV), human adenovirus (HAdV) and human coronaviruses (HCoVs) showed the distinct circulating patterns in north and south China. Human enterovirus/human rhinovirus (HEV/HRV) (27.6%, 27/98), HBoV (18.4%, 18/98), RSV (16.3%, 16/98) and HMPV (14.3%, 14/98) were the most commonly detected viruses in severe pneumonia cases with single virus infection. In conclusion, viral pathogens are frequently detected in pediatric CAP cases and may therefore play a vital role in the aetiology of CAP. RSV was the most important virus in hospitalized children with CAP in Chinese mainland.

Baicalin relieves Mycoplasma pneumoniae infection­induced lung injury through regulating microRNA­221 to inhibit the TLR4/NF­κB signaling pathway.

Mycoplasma pneumoniae (MP) is a common pathogen that can cause respiratory infections. MP pneumonia (MPP) leads to numerous complications, including lung injury and even death. The present study aimed to investigate the protective effects of Baicalin treatment on MP infection­induced lung injury and the molecular mechanism underlying these effects. Briefly, after mice were infected intranasally by MP and treated with Baicalin (80 mg/kg), serum levels of MP­immunoglobulin M (IgM) were detected by ELISA. The expression levels of C­reactive protein (CRP) in lung tissue were detected by immunohistochemistry and the bronchoalveolar lavage fluid (BALF) was examined by ELISA. Inflammatory factors and inflammatory cells in the BALF were assessed. The expression levels of microRNA (miR)­221 in lung tissue were examined by reverse transcription­quantitative PCR and pathological changes in lung tissue were detected by H&E staining. Cell apoptosis was evaluated by TUNEL assay and the protein expression levels of TLR4, MyD88 and NF­κB were detected by western blotting. Baicalin treatment significantly reduced serum levels of MP­IgM and CRP expression in lung tissue during MP infection. In addition, Baicalin decreased the levels of IL­1ß, IL­6, IL­18 and TNF­α in the BALF, and the number of inflammatory cells. Baicalin also reduced the inflammatory infiltration in lung tissue induced by MP infection, improved the pathological changes detected in lung tissue, reduced apoptosis, and downregulated the protein expression levels of TLR4, MyD88 and NF­κB. Furthermore, Baicalin treatment downregulated the expression of miR­221 and the protective effects of Baicalin were attenuated by miR­221 overexpression. In conclusion, Baicalin has a therapeutic effect on mice with MP infection­induced lung injury, which may be related to inhibition of miR­221 expression and regulation of the TLR4/NF­κB signaling pathway.

Fructose-1,6-bisphosphatase aggravates oxidative stress-induced apoptosis in asthma by suppressing the Nrf2 pathway.

Asthma is a chronic airway disease that causes excessive inflammation, oxidative stress, mucus production and bronchial epithelial cell apoptosis. Fructose-1,6-bisphosphatase (Fbp1) is one of the rate-limiting enzymes in gluconeogenesis and plays a critical role in several cancers. However, its role in inflammatory diseases, such as asthma, is unclear. Here, we examined the expression, function and mechanism of action of Fbp1 in asthma. Gene Expression Omnibus (GEO) data sets revealed that Fbp1 was overexpressed in a murine model of asthma and in interleukin (IL)-4- or IL-13-stimulated bronchial epithelial cells. We confirmed the findings in an animal model as well as Beas-2B and 16HBE cells. In vitro investigations revealed that silencing of Fbp1 reduced apoptosis and the proportion of cells in the G2/M phase, whereas overexpression led to increases. Fbp1 knock-down inhibited oxidative stress by activating the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway, whereas Fbp1 overexpression aggravated oxidative stress by suppressingthe Nrf2 pathway. Moreover, the Nrf2 pathway inhibitor ML385 reversed the changes caused by Fbp1 inhibition in Beas-2B and 16HBE cells. Collectively, our data indicate that Fbp1 aggravates oxidative stress-induced apoptosis by suppressing Nrf2 signalling, substantiating its potential as a novel therapeutic target in asthma.

Significance of RNA N6-Methyladenosine Regulators in the Diagnosis and Subtype Classification of Childhood Asthma Using the Gene Expression Omnibus Database.

RNA N6-methyladenosine (m6A) regulators play important roles in a variety of biological functions. Nonetheless, the roles of m6A regulators in childhood asthma remain unknown. In this study, 11 significant m6A regulators were selected using difference analysis between non-asthmatic and asthmatic patients from the Gene Expression Omnibus GSE40888 dataset. The random forest model was used to screen five candidate m6A regulators (fragile X mental retardation 1, KIAA1429, Wilm's tumor 1-associated protein, YTH domain-containing 2, and zinc finger CCCH domain-containing protein 13) to predict the risk of childhood asthma. A nomogram model was established based on the five candidate m6A regulators. Decision curve analysis indicated that patients could benefit from the nomogram model. The consensus clustering method was performed to differentiate children with asthma into two m6A patterns (clusterA and clusterB) based on the selected significant m6A regulators. Principal component analysis algorithms were constructed to calculate the m6A score for each sample to quantify the m6A patterns. The patients in clusterB had higher m6A scores than those in clusterA. Furthermore, we found that the patients in clusterA were linked to helper T cell type 1 (Th1)-dominant immunity while those in clusterB were linked to Th2-dominant immunity. In summary, m6A regulators play nonnegligible roles in the occurrence of childhood asthma. Our investigation of m6A patterns may be able to guide future immunotherapy strategies for childhood asthma.

WU Polyomavirus Infection in Children With Acute Lower Respiratory Tract Infections in China, 2017 to 2019: Case Reports and Multicentre Epidemiological Survey.

WU polyomavirus (WUPyV) is a novel member of the family Polyomaviridae recently detected in respiratory tract specimens. So far, it has not been proven whether WUPyV is a real causative agent for respiratory diseases. In this study, we described two patients with fatal infection who had WUPyV detected in their nasopharyngeal swabs. Furthermore, we conducted a multicentre study in six hospitals from different districts of China. WUPyV was detected by real-time polymerase chain reaction assays, and the clinical and molecular epidemiological characteristics of WUPyV strains among hospitalized children with acute lower respiratory tract infections all around China from 2017 to 2019 were analysed. Two complete WUPyV genome sequences were assembled from fatal patients' airway specimens. Phylogenetic tree analysis revealed that they were most closely related to strains derived from Fujian and Chongqing, China, in 2008 and 2013, respectively. In 2017-2019, a total of 1,812 samples from children with acute lower respiratory tract infections were detected for WUPyV, of which 11 (0.6%) were positive. Children aged ≤5 were more susceptible to WUPyV infection. A total of 81.8% of WUPyV-positive patients were coinfected with other viruses, of which rhinovirus enjoyed the highest frequency. The main clinical symptoms of infected patients include fever, coughing and sputum expectoration. Most patients were diagnosed with pneumonia, followed by bronchial surgery. Three patients manifested severe infection, and all patients improved and were discharged. Our results show that WUPyV persistently circulates in China. Further investigations on the clinical role and pathogenicity of WUPyV are necessary.

Downregulation of miRNA-451a Promotes the Differentiation of CD4+ T Cells towards Th2 Cells by Upregulating ETS1 in Childhood Asthma.

Children exposed to common aeroallergens may develop asthma that progresses into adulthood. Inflammation regulated by T helper 2 (Th2) cells, a specific subpopulation of CD4+ T lymphocytes, is involved in asthmatic injury. Herein, our microarray data indicated that microRNA-451a-5p (miRNA-451a) expression decreased by 4.6-fold and ETS proto-oncogene 1 (ETS1) increased by 2.2-fold in the peripheral blood lymphocytes isolated from asthmatic children (n = 4) as compared to control individuals (n = 4). The negative correlation between miRNA-451a and ETS1 was further validated in 40 CD4+ T cell samples (10 healthy vs. 30 asthmatic samples). In vitro, naïve CD4+ T cells isolated from control individuals were cultured under Th2 cell polarizing condition. miRNA-451a expression decreased while ETS1 increased in CD4+ T cells in the setting of Th2 cell polarization. Moreover, miRNA-451a knockdown enhanced Th2 cell polarization - cells positive for both GATA3 (GATA binding protein 3, a Th2-transcription factor) and CD4 increased, and the generation of Th2 cell cytokines, interleukin (IL)5 and IL13, increased. In contrast, miRNA-451a overexpression inhibited Th2 cell differentiation. Interestingly, dual-Luciferase assay proved ETS1 as a novel target of miRNA-451a. Moreover, enforced expression of ETS1 partially restored miRNA-451a-induced inhibition of IL5 and IL13, and increased the GATA3+CD4+ cell population. Collectively, our work demonstrates that downregulation of miRNA-451a upregulates ETS1 expression in CD4+ T cells, which may contribute to Th2 cell differentiation in pediatric asthma.

Epigallocatechin gallate ameliorates airway inflammation by regulating Treg/Th17 imbalance in an asthmatic mouse model.

Epigallocatechin gallate (EGCG) is a polyphenol that is found in green tea that has been shown to ameliorate airway inflammation in an ovalbumin-sensitized asthmatic mouse model. The purpose of this study was to investigate whether the immunomodulatory and anti-inflammatory effects of EGCG by regulating the regulatory T cell (Treg)/Th 17 cells balance in this model. Female BALB/c mice were sensitized and challenged with ovalbumin by intraperitoneal injection. EGCG was administered to asthmatic mice intraperitoneally 1 h before each OVA challenge. Airway hyperresponsiveness (AHR) was measured, and lung inflammatory infiltrations were assessed by hematoxylin and eosin (HE) staining. Serum OVA-specific IgE levels, Interleukin-10 (IL-10) levels and Interleukin-17A (IL-17A) levels in the bronchoalveolar lavage fluid (BALF), serum, and splenocyte culture supernatants were measured by ELISA. Flow cytometry was used to assess the effects of EGCG on the frequency of CD4+CD25+Foxp3+Treg cells in the splenocytes and real-time PCR method was used to measure the expression of Forkhead box P3 (Foxp3) mRNA and retinoid-related orphan receptor gammat (RORγt) mRNA in the lung tissue. The results showed that administration of EGCG significantly decreased AHR and OVA specific IgE in the serum, increased IL-10 levels in the BALF, serum, and splenocyte culture supernatant, and the frequency of CD4+CD25+Foxp3+Treg cells in the splenocytes in asthmatic mice. Administration of EGCG also ameliorated airway inflammation and eosinophil infiltrations in asthmatic mice. These results suggested that EGCG likely ameliorated OVA-induced airway inflammation by increasing the production of IL-10, the number of CD4+CD25+Foxp3+Treg cells and expression of Foxp3 mRNA in the lung tissue, and it could be an effective agent for treating asthma.

Epigallocatechin gallate improves airway inflammation through TGF­ß1 signaling pathway in asthmatic mice.

The present study aimed to investigate the effect of epigallocatechin gallate (EGCG) on airway inflammation in mice with bronchial asthma, and the regulatory mechanism of transforming growth factor (TGF)­ß1 signaling pathway, so as to provide theoretical basis for research and development of a novel drug for clinical treatment. Mouse models of bronchial asthma were established and injected with dexamethasone and EGCG via the caudal vein. 7 days later, bronchoalveolar tissue was collected for hematoxylin and eosin staining. Determination of airway resistance (AWR) and lung function in mice was detected. Serum was separated for cytometric bead array to detect the changes in inflammatory factors. Bronchoalveolar lavage fluid was collected for eosinophil and neutrophil counts. Fresh blood was obtained for flow cytometry to determine the percentages of Th17 and Treg cells. Bronchovesicular tissue was utilized for western blot assay and reverse transcription­quantitative polymerase chain reaction to determine the proteins and transcription factors in the TGF­ß1 pathway. EGCG 20 mg/kg significantly reduced asthmatic symptoms, lung inflammatory cell infiltration, and the inflammatory factor levels of interleukin (IL)­2, IL­6 and tumor necrosis factor (TNF)­α. In addition, it increased the levels of inflammatory factors, including IL­10, diminished the percentage of Th17 cells, increased the percentage of Treg cells, and decreased the expression of TGF­ß1 and phosphorylated (p)­Smad2/3 expression. Following the inhibition of the TGF­ß1 receptor, the anti­inflammatory effect of EGCG disappeared, and the expression of TGF­ß1 and p­Smad2/3 increased. EGCG attenuated airway inflammation in asthmatic mice, decreased the percentage of Th17 cells and increased the percentage of Treg cells. The anti­inflammatory effect of EGCG is achieved via the TGF­ß1 signaling pathway.

Vasoactive intestinal peptide inhibits airway smooth muscle cell proliferation in a mouse model of asthma via the ERK1/2 signaling pathway.

Asthma is a heterogeneous clinical syndrome characterized by airway inflammation, hyper-responsiveness and remodeling. Airway remodeling is irreversible by current antiasthmatic drugs, and it is the main cause of severe asthma. Airway smooth muscle cells (ASMCs) act as the main effector cells for airway remodeling; the proliferation and hypertrophy of which are involved in airway remodeling. Caveolin (Cav)- 1 is present on the surface of ASMCs, which is involved in cell cycle and signal transduction regulation, allowing ASMCs to change from proliferation to apoptosis. The extracellular signal-regulated kinase (ERK)1/2 signaling pathway is a common pathway regulated by various proliferative factors, which demonstrates a regulatory role in airway remodeling of asthma. There have been many studies on the correlation between vasoactive intestinal peptide (VIP) and airway reactivity and inflammation in asthma, but the functions and related mechanisms of ASMCs remain unclear. In this study, we established an airway remodeling model in asthmatic mice, and concluded that VIP inhibits airway remodeling in vivo. The in vitro effect of VIP on interleukin-13-induced proliferation of ASMCs was studied by examining the effects of VIP on expression of ERK1/2, phospho-ERK1/2 and Cav-1 in ASMCs, as well as changes in cell cycle distribution. VIP inhibited phosphorylation of the ERK1/2 signaling pathway and expression of Cav-1 on ASMCs and decreased the proportion of S phase cells in the cell cycle, thus inhibiting the proliferation of ASMCs. This study provides a novel therapeutic mechanism for the treatment of asthma.

Vitamin D reduces inflammatory response in asthmatic mice through HMGB1/TLR4/NF­κB signaling pathway.

The present study aimed to investigate the effects of vitamin D (VD) on inflammatory responses in asthmatic mice and the underlying mechanism, providing a theoretical basis for clinical application of targeted drug therapy, and the development of novel drugs against asthma. Mouse models of asthma were established. Hematoxylin­eosin staining was performed to observe the pathological changes of the lung tissue. Pulmonary function tests were conducted to determine airway resistance in asthmatic mice. ELISA was performed to measure the serum levels of inflammatory factors. Western blot analysis and reverse transcription­quantitative polymerase chain reaction were performed to determine the changes in apoptosis­inducing factors, and high mobility group box 1 protein (HMGB1)/Toll­like receptor­4 (TLR4)/nuclear factor (NF)­κB signaling pathway­related proteins. VD reduced infiltrated inflammatory factors, attenuated the airway resistance of asthmatic mice, decreased serum levels of interleukin (IL)­1ß, IL­6, tumor necrosis factor (TNF)­α, increased serum levels of IL­10, decreased apoptotic factor Bcl­2­associated X and caspase­3 expression, downregulated HMGB1 and TLR4, NF­κB and phosphorylated­NF­κB p65 expression. When TLR4 expression was inhibited, the anti­inflammatory effects of VD were attenuated, and HMGB1, TLR4, NF­κB and p­NF­κB p65 expression was increased. VD was able reduce the inflammatory response of asthmatic mice and apoptosis in lung tissue through the HMGB1/TLR4/NF­κB signaling pathway.

Epigallocatechin-3-gallate inhibits inflammation and epithelial­mesenchymal transition through the PI3K/AKT pathway via upregulation of PTEN in asthma.

Asthma is a chronic disease associated with hyperresponsiveness, obstruction and remodeling of the airways. Epithelial-mesenchymal transition (EMT) has an important role in these alterations and may account for the accumulation of subepithelial mesenchymal cells, thus contributing to airway hyperresponsiveness and remodeling. Epigallo-catechin­3­gallate (EGCG), which is a type of polyphenol, is the most potent ingredient in green tea, and exhibits antibacterial, antiviral, antioxidative, anticancer and chemopreventive activities. Recently, numerous studies have investigated the protective effects of EGCG against asthma and other lung diseases. In the present study, the role of EGCG in ovalbumin (OVA)­challenged asthmatic mice was determined. In addition, the inhibitory effects of EGCG against transforming growth factor (TGF)­ß1­induced EMT and migration of 16HBE cells, and the underlying mechanisms of the phosphatidylinositol 3­kinase/protein kinase B (PI3K/Akt) signaling pathway, were investigated by immunofluorescence, Transwell, wound healing assay and western blot analysis, respectively. The results indicated that EGCG may suppress inflammation and inflammatory cell infiltration into the lungs of OVA­challenged asthmatic mice, and may also inhibit EMT via the PI3K/Akt signaling pathway through upregulating the expression of phosphatase and tensin homolog (PTEN) in vivo and in vitro. The present study also revealed the anti­migratory effects of EGCG in TGF­ß1­induced 16HBE cells, thus suggesting it may reduce airway remodeling. The present study provides a novel insight into understanding the protective effects of EGCG on airway remodeling in asthma, and indicates that EGCG may be useful as an adjuvant therapy for bronchial asthma.

Curcumin ameliorates asthmatic airway inflammation by activating nuclear factor-E2-related factor 2/haem oxygenase (HO)-1 signalling pathway.

Previous studies have shown that curcumin alleviates asthma in vivo. However, the relationship between curcumin and the nuclear factor-E2-related factor 2 (Nrf2)/haem oxygenase (HO)-1 pathway in asthma treatment remains unknown. The aim of the present study was to investigate the mechanisms of curcumin involved in the amelioration of airway inflammation in a mouse asthma model. Curcumin was administrated to asthmatic mice, and bronchoalveolar lavage fluid was collected. Inflammatory cell infiltration was measured by Giemsa staining. Immunoglobulin E production in bronchoalveolar lavage fluid was measured by enzyme-linked immunosorbent assay. Histological analyses were evaluated with haematoxylin-eosin and periodic acid-Schiff staining. Airway hyperresponsiveness was examined by whole-body plethysmography. Nuclear factor-E2-related factor 2, HO-1, nuclear factor-κB and inhibitory κB/p-inhibitory κB levels in lung tissues were detected by western blot, and Nrf2 activity was measured by electrophoretic mobility shift assay. Tumour necrosis factor-α, interleukin (IL)-1ß, and IL-6 levels in the small interfering RNA-transfected cells were detected by enzyme-linked immunosorbent assay. Curcumin treatment significantly reduced immunoglobulin E production, attenuated inflammatory cell accumulation and goblet cell hyperplasia, and ameliorated mucus secretion and airway hyperresponsiveness. Nuclear factor-E2-related factor 2 and HO-1 levels in lung tissues were significantly increased. Meanwhile, Nrf2 activity was enhanced. Nuclear factor-κB and p-inhibitory κB levels were elevated in the lung tissue of ovalbumin-challenged mice. Both were restored to normal levels after curcumin treatment. Haem oxygenase-1 and nuclear Nrf2 levels were enhanced in dose- and time-dependent manners in curcumin-treated RAW264.7 cells. Curcumin blocked lipopolysaccharide-upregulated expression of tumour necrosis factor-α, IL-1ß, and IL-6. After the cells were transfected with HO-1 or Nrf2 small interfering RNA, lipopolysaccharide-induced pro-inflammation cytokine expression was significantly restored. In summary, curcumin might alleviate airway inflammation in asthma through the Nrf2/HO-1 pathway, potentially making it an effective drug in asthma treatment.