RATIONALE:Lingguizhugan decoction (LGZG) is a traditional Chinese formula that has been commonly used in obstructive sleep apnea (OSA) for relieving lung inflammation. However, the active substance of LGZG and the specific mechanism remain unclear. This study aims to identify the bioactive components of LGZG and subsequently elucidate the underlying therapeutic mechanisms against OSA based on mass spectrometry analysis, network pharmacology, transcriptomics, and experimental verification. METHODS:Ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) was used to identify the main ingredients of LGZG. The OSA animal model was induced by chronic intermittent hypoxia (CIH) for 5 weeks in C57BL/6 mice. Transcriptome sequencing and network pharmacology were used to analyze potential mechanisms, which were further validated by molecular docking. HE staining was used for detecting lung inflammation. Immunohistochemistry (IHC), ELISA, and Western blot were employed to investigate protein expression, while quantitative real-time PCR (RT-qPCR) was used to determine gene expressions. Tissue reactive oxygen species (ROS) levels were measured by the DCFH-DA probe method. RESULTS:LGZG inhibited CIH-induced pulmonary inflammatory infiltration, protein concentration in bronchoalveolar lavage fluid (BALF), and suppressed IL-17 and IL-1β gene expression. UHPLC-HRMS identified 482 compounds in the LGZG aqueous decoction. Network pharmacology analysis revealed that IL-6 and HIF-1α pathway were the major targets. Subsequently, transcriptomics analysis revealed that LGZG affected functions associated with ROS production and polymorphonuclear cells. LGZG suppressed the CIH-induced expression of neutrophil elastase and reduced MPO production in BALF. Furthermore, LGZG inhibited the IL-6 expression and secretion, and reduced CIH-induced ROS production. LGZG inhibited the CIH-induced activation of HIF-1α pathway. Moreover, molecular docking identified compounds in LGZG that could directly interact with the core targets IL-6, MPO, CYBB, and HIF-1α. CONCLUSIONS:LGZG alleviates CIH-induced pulmonary inflammation, neutrophil infiltration, and IL-6 secretion in mice. These effects are associated with the suppression of ROS production and inhibition of the HIF-1α signaling pathway.

OBJECTIVE:Exposure to benzo(a)pyrene (BaP) negatively affects lung inflammation in patients with asthma. However, there is a lack of systematic research on the key mechanisms of BaP toxicity in asthma. METHODS:In this study, BaP target genes were predicted using ChEMBL, SEA, and PharmMapper, and asthma-related genes were retrieved from GeneCards, OMIM, and TTD. A protein-protein interaction network was constructed in Cytoscape software using overlapping genes. The causal relationships between core genes and asthma risk were evaluated using Mendelian randomization. BALB/c mice were treated with ovalbumin (OVA) with or without BaP. Single-cell RNA sequencing (scRNA-seq) was performed on mouse lung tissues, and hematoxylin and eosin (HE) staining was used to assess lung inflammation. Molecular docking and molecular dynamics simulations were performed to assess BaP-target interactions. Virtual knockout was performed using scTenifoldKnk. The CIBERSORT algorithm was applied to analyze immune cell infiltration. RESULTS:A total of 23 potential core targets of BaP-induced asthma were initially screened using network toxicology, followed by functional enrichment analysis. The identified target genes were mainly involved in leukocyte migration, immunity, and phosphodiesterase activity, and were enriched in cytokine-cytokine receptor interaction, NF-kappa B signaling pathway, and TNF signaling pathway. Further analysis of the initially selected target genes using machine learning and Mendelian randomization revealed that only IL1R1 remains relevant to asthma, and as IL1R1 expression increases, the risk of asthma onset and exacerbation also increases. Afterward, single-cell sequencing data were obtained from mouse lung tissues (two OVA and two OVA + BaP), and the results verified that BaP induces IL1R1 expression in lymphocytes and vascular endothelial cells. Additionally, molecular docking and molecular dynamics simulations demonstrated that BaP and IL1R1 have strong binding capacity. Virtual knockout of IL1R1 regulated inflammatory responses and immune cell analysis, suggesting that IL1R1 induced mast cell activation. CONCLUSIONS:This study elucidates the mechanism by which BaP exacerbates asthma by modulating the key target IL1R1, offering novel insights into BaP-induced asthma pathogenesis and identifying IL1R1 as a promising biomarker for risk assessment and therapeutic intervention.

Clear cell renal cell carcinoma (ccRCC) is a highly heterogeneous malignancy with complex molecular features, posing significant challenges for accurate prognosis prediction and treatment stratification. Lactylation, an emerging epigenetic regulatory mechanism, plays a pivotal role in tumor progression and immune microenvironment remodeling. This study aims to develop a lactylation-related gene signature (LRGS) and systematically evaluate its prognostic value and potential for guiding clinical interventions in ccRCC. We developed a machine learning-derived lactylation signature using lactylation-related genes (LRGs) selected through bulk and single-cell RNA sequencing data. The model’s performance was subsequently validated using internal (TCGA-KIRC) and external (E-MTAB-1980) cohorts. The associations of LRGS with clinical characteristics, immune cell infiltration, and drug response were further analyzed. The functions of the key risk gene SHC1 were investigated using in vitro experiments. Ultimately, an 11-gene LRGS was identified that effectively predicts the prognosis of ccRCC patients. This model demonstrated robust predictive capability in an external dataset (AUC > 0.7). Univariate and multivariate analyses indicated that the model’s risk score was an independent predictor of clinical outcomes. The high-risk group exhibited significant infiltration of Tregs and a higher tumor immune dysfunction score, indicating a poorer responsiveness to immunotherapy. Finally, cell experiments confirmed that inhibiting SHC1 expression suppressed the proliferation and migration of ccRCC cells. In conclusion, we successfully constructed a novel LRGS, which provides a new perspective for risk stratification and personalized treatment of ccRCC.

Airway remodeling, involving fibroblast activation and collagen deposition, is a hallmark of asthma. The pollutant Benzo(a)pyrene (BaP), an aryl hydrocarbon receptor (AhR) activator, is associated with increased asthma severity, but its role in remodeling remains unclear. This study investigates whether BaP exacerbates remodeling by enhancing epithelial-fibroblast crosstalk through an AhR-mediated pathway and identifies key mediators. We found BaP exposure significantly upregulated growth differentiation factor 15 (GDF15) expression and secretion from airway epithelial cells in an AhR-dependent manner, as AhR directly bound to the GDF15 promoter. Consequently, conditioned media from BaP-treated epithelial cells promoted fibroblast activation, marked by elevated α-smooth muscle actin (α-SMA) and type I collagen (COL1A1) levels, which were reduced by GDF15 knockdown. In vivo, BaP co-exposure worsened allergen-induced airway remodeling, with greater collagen deposition and higher GDF15, α-SMA, and COL1A1 expression. Consistently, recombinant GDF15 directly activated lung fibroblasts and stimulated collagen production. Critically, inhibition of AhR by epigallocatechin gallate (EGCG) attenuated BaP-induced GDF15 expression and remodeling in vivo, a effect associated with reduced AhR expression. In conclusion, these findings demonstrate that BaP aggravates airway remodeling via epithelial-fibroblast crosstalk mediated by the AhR-GDF15 axis, highlighting a potential target for treating pollutant-exacerbated asthma.

Growing evidence implicates gut microbiota (GM) dysbiosis in Alzheimer’s disease (AD) pathogenesis via the gut-brain axis. Dysbiosis contributes to neuroinflammation, amyloid-β deposition, tau hyperphosphorylation, blood-brain barrier disruption, and cognitive decline. Synbiotics (combinations of probiotics and prebiotics) offer a promising strategy to modulate GM, potentially ameliorating these AD hallmarks through multiple mechanisms including enhanced production of neuroprotective short-chain fatty acids (SCFAs), reduced inflammation, improved gut barrier integrity, and immunomodulation. This review critically evaluates the current evidence on the therapeutic potential of synbiotics for AD. It aims to synthesize findings from preclinical and clinical studies regarding the efficacy of synbiotics in improving cognitive function and AD pathology, elucidate the underlying biological mechanisms including GM modulation, SCFA production, immune regulation, and gut-brain signaling, and identify key challenges and future research directions for translating GM-targeted interventions into effective AD therapies. Synbiotics demonstrate significant potential, particularly in early AD, by improving cognitive domains, reducing neuroinflammation and AD biomarkers, and modulating beneficial microbial metabolites. However, challenges include confounding factors, unresolved questions about causality, inconsistent results in advanced disease, and insufficient large-scale human trials. Future success hinges on rigorous longitudinal randomized controlled trials integrating multi-omics approaches, advanced in vitro models, and personalized strategies considering baseline microbiota and host genetics. While not a standalone cure, synbiotics represent a valuable component within multi-target therapeutic approaches aimed at modulating the gut-brain axis to slow AD progression.

BACKGROUND:Obstructive sleep apnea (OSA) is characterized by chronic intermittent hypoxia (CIH) and subsequent respiratory tract inflammation. While air pollution is identified as an OSA risk factor and can influence disease severity, its specific pulmonary effects and underlying mechanisms remain unclear. METHODS:We established a murine model of OSA in C57BL/6 mice using 5-week CIH, with and without 2-day diesel exhaust particle (DEP) exposure and used 16HBE bronchial epithelial cells in vitro. We conducted RNA sequencing on lung tissues to analyse gene expression and utilized hematoxylin and eosin (HE) staining to assess mice lung inflammation. Additionally, immunohistochemistry was performed to evaluate protein expression levels. Chemokines in bronchoalveolar lavage fluid (BALF) were identified using a panel, followed by ELISA quantification. RESULTS:Compared with the control group, CIH resulted in heightened lung inflammation, increased BALF protein concentrations, and elevated expression of hypoxia-related genes. The CIH + DEP group exhibited even more pronounced elevations in these indicators. RNA sequencing identified that CIH upregulated neutrophil-related cytokine genes in lung tissue. Specifically, DEP amplified this upregulation. Furthermore, myeloperoxidase expression, indicative of neutrophil activation, significantly increased in the lung tissue and BALF of the CIH group. The CIH + DEP group showed an even more substantial rise. Subsequent protein screening and ELISA confirmed marked elevations of CXCL5 and CCL5 in both CIH and DEP groups. In 16HBE cells, DEP aggravated CIH-induced secretion of CXCL5 and CCL5, and p-p38 protein expression, which was ameliorated by the p38 MAPK inhibitor SB 203580. CONCLUSION:DEP aggravated CIH-induced lung inflammation and neutrophil infiltration, which was likely mediated by CXCL5 and CCL5.

Background Asthma is a chronic airway inflammatory disease characterized by airway inflammation, mucus hypersecretion, airway hyper-reactivity. Sanzi Yangqin Decoction (SZYQD) is widely prescribed for asthma treatment. Its anti-asthma activities have been reported in animal model, but the exact mechanism and targets of SZYQD in asthma treatment have not been fully elucidated. Methods A network pharmacological approach was used to predict the active components, targets, and signalling pathways of SZYQD in asthma, including potential target prediction, protein‒protein interaction (PPI) network construction and analysis, and Gene Ont (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The active ingredients were identified from the SZYQD, and were molecular docked according to the results of network pharmacology. A mouse model of asthma induced by ovalbumin (OVA) and lipopolysaccharide (LPS) was constructed to evaluate the therapeutic effect of SZYQD. Furthermore, the effects of SZYQD and its active ingredients were tested in vitro for regulating inflammation and MUC5AC expression (two main pathophysiologic abnormalities of asthma) in macrophages and airway epithelial cells by using Real-time PCR and western blotting. Results A total of 28 active ingredients and 111 HUB genes were screened in the relevant databases, including three key ingredients (luteolin, β-carotene, and Sinapine) and nine core target genes (JUN, CTNNB1, IL10, TP53, AKT1, STAT3, TNF, IL6 and EGFR). KEGG and GO analysis indicated that the potential anti-asthmatic mechanisms of SZYQD were related to PI3K-Akt signalling pathway and response to lipopolysaccharide, etc. In the in vivo asthmatic model, our findings demonstrated that SZYQD exerted a protective effect against asthmatic mice induced by OVA and LPS through the inhibition of inflammation and mucus overproduction. Consistently, cell experiments showed that the SZYQD extract or the key active ingredients luteolin significantly decreased lipopolysaccharide (LPS)-induced IL-6 expression and activation of the NF-κB pathway in macrophages. In addition, SZYQD extract or luteolin inhibited activation of the AKT pathway and expression of MUC5AC induced by EGF in airway epithelial cells. Conclusion The anti-asthmatic mechanism of SZYQD might be associated with inhibiting inflammation and airway mucus hypersecretion by regulating the NF-κB and AKT signalling pathways as predicted by network pharmacology, which provides more evidence for the application of SZYQD in asthma treatment.

Ursolic acid (UA), a natural pentacyclic triterpenoid, is known to exhibit various biological activities and anticancer effects. However, the underlying anticancer mechanism is not fully understood to date. The present study aimed to investigate the antimetastatic effect of UA through ADP-ribosylation factor like GTPase 4C (ARL4C) in colon cancer. A lung metastasis model of colon cancer in nude mice was established through tail vein injection. A Cell Counting Kit-8 assay was used to investigate the proliferation of colon cancer cells. Transwell assays were used to detect cell migration and invasion. The expression levels of proteins including ARL4C, matrix metallopeptidase 2 (MMP2), phosphorylated (p)-AKT and p-mTOR were measured using western blot analysis. Immunohistochemistry was used to determine the protein expression level in tissues. ARL4C ubiquitination levels were analysed using immunoprecipitation and western blotting. The results indicated that UA inhibits the metastasis of colon cancer in vivo and in vitro. The expression of ARL4C in human colon cancer tissue was significantly higher than that in adjacent tissues and its high expression level was associated with lymph node metastases and tumour stage. UA treatment significantly decreased ARL4C and MMP2 protein levels and inhibited the AKT/mTOR signalling pathway. Overexpression of ARL4C reversed the inhibitory effect of UA on the invasion and migration of HCT-116 and SW480 cells, as well as the expression and secretion of MMP2 protein. In addition, UA and an AKT signalling pathway inhibitor (LY294002) induced the ubiquitination of the ARL4C protein, which was reversed by a proteasome inhibitor (MG-132). Collectively, it was revealed in the present study that UA served as a novel solution to relieve colon cancer metastasis by inducing the ubiquitination-mediated degradation of ARL4C by modulating the AKT signalling pathway. Thus, UA may be a promising treatment option to prolong the survival of patients with colon cancer metastasis.

Objective To observe the effects of Qixian Decoction（composed of Astragali Radix,Herba Epimedii,Morindae Officinalis Radix,etc.） on airway inflammation and mucus secretion in asthmatic mice exposed to the atmospheric pollutant benzopyrene（BaP）,and to explore the molecular mechanisms.Methods Female BALB/c mice were randomly divided into blank group,asthma group[ovalbumin（OVA）sensitization],BaP group（OVA+BaP）and Qixian Decoction group(OVA+BaP+Qixian Decoction 37.5 g·kg -1 ),with 10 mice in each group.The mouse asthma model was replicated using the OVA sensitization method,and exposed to atmospheric pollution was simulated by BaP nasal drip method.The levels of lung inflammation and mucus secretion were detected by HE and AB-PAS staining;the mRNA expressions of interleukin 4（IL-4）and interleukin 13（IL-13）in lung tissue were detected by qRT-PCR,and the protein expressions of mucin 5AC（MUC5AC）and phosphorylated ERK（p-ERK）in lung tissue were detected by immunohistochemistry;the lung tissue reactive oxygen species（ROS）levels were measured by the DCFH-DA probe method.Results Compared with the blank group,the inflammatory cells around airway and blood vessels in asthma group were significantly increased;the airway mucosa was interrupted and discontinuous,and the inflammation score was significantly increased（P＜0.01）;the mRNA expressions of IL-4 and IL-13 in mice lung tissue were significantly up-regulated（P＜0.05）;the airway mucus secretion of mice was significantly increased（P＜0.01）;the protein expressions of MUC5AC and p-ERK in airway of mice were significantly up-regulated（P＜0.01）;the ROS level in mouse lung tissue was significantly increased（P＜0.05）.Compared with the asthma group,the inflammatory cell infiltration in BaP group was further aggravated,the alveolar septum was significantly broken,and the inflammatory score was significantly increased（P＜0.01）;the mRNA expressions of IL-4 and IL-13 in mouse lung tissue were significantly up-regulated（P＜0.01）;the airway mucus in mice was significantly increased（P＜0.01）;the protein expressions of MUC5AC and p-ERK in mice airway were significantly up-regulated（P＜0.05,P＜0.01）;and the ROS level in mice lung tissue was significantly increased（P＜0.01）.Compared with the BaP group,the inflammatory cell infiltration and airway mucosal continuity in the mice in the Qixian Decoction group were significantly improved,and the inflammation score was significantly reduced（P＜0.01）;the mRNA expressions of IL-4 and IL-13 in mice lung tissue were significantly down-regulated（P＜0.01）;the airway mucus secretion in mice was significantly reduced（P＜0.01）;the protein expressions of MUC5AC and p-ERK in mice airway were significantly down-regulated（P＜0.05,P＜0.01）;the ROS level in mice lung tissue was significantly reduced（P＜0.01）.Conclusion Qixian Decoction has effects in relieving airway inflammation and mucus secretion in mice with asthma exacerbated by atmospheric pollutants BaP and the anti-lung damage effects were possibly related to its inhibition of ROS and ERK pathways.

OBJECTS:Benzo(a)pyrene (BaP), an environmental pollutant, is present in high concentrations in urban smog and cigarette smoke and has been reported to promote high mucin 5AC (MUC5AC) expression. Epithelium-derived inflammatory cytokines are considered an important modulator of mucus oversecretion and MUC5AC overexpression. Here, we investigated whether the effect of BaP on MUC5AC overexpression was associated with cytokine autocrine activity in vivo and in vitro.METHODS:In vivo, BALB/c mice were treated with ovalbumin (OVA) in the presence or absence of BaP. Allergy-induced mucus production was assessed by Alcian Blue Periodic acid Schiff (AB-PAS) staining. The human airway epithelial cell line NCI-H292 was used in vitro. MUC5AC and transforming growth factor (TGF)-α mRNA levels were assessed with real-time quantitative PCR. The concentration of cytokines was measured by ELISA. The MUC5AC, p-ERK, ERK, p-EGFR and EGFR proteins were detected by Western blotting in cells or by immunohistochemistry in mouse lungs. Small-interfering RNAs were used for gene silencing.RESULTS:TGF-α was overproduced in the supernatant of NCI-H292 cells treated with BaP. Knockdown of TGF-α expression inhibited the BaP-induced increase in MUC5AC expression and subsequent activation of the EGFR-ERK signalling pathway. Knocking down aryl hydrocarbon receptor (AhR) expression or treatment with an ROS inhibitor (N-acetyl-L-cysteine) could relieve the TGF-α secretion induced by BaP in epithelial cells. In an animal study, coexposure to BaP with OVA increased mucus production, MUC5AC expression and ROS-EGFR-ERK activation in the lung as well as TGF-α levels in bronchoalveolar lavage fluid (BALF). Furthermore, the concentration of TGF-α in BALF was correlated with MUC5AC mRNA levels. Additionally, TGF-α expression was found to be positively correlated with MUC5AC expression in the airway epithelial cells of smokers. Compared with non-smoker asthma patients, TGF-α serum levels were also elevated in smoker asthma patients.CONCLUSION:Autocrine TGF-α was associated with BaP-induced MUC5AC expression in vitro and in vivo. BaP induced TGF-α secretion by activating AhR and producing ROS, which led to activation of the EGFR-ERK pathway.

Skeletal muscle wasting is a clinically remarkable phenotypic feature of pulmonary arterial hypertension (PAH) that increases the risk of mortality. Growth differentiation factor 11 (GDF11), centrally involved in PAH pathogenesis, has an inhibitory effect on skeletal muscle growth in other conditions. However, whether GDF11 is involved in the pathogenesis of skeletal muscle wasting in PAH remains unknown. We showed that serum GDF11 levels in patients were increased following PAH. Skeletal muscle wasting in the MCT-treated PAH model is accompanied by an increase in circulating GDF11 levels and local catabolic markers (Fbx32, Trim63, Foxo1, and protease activity). In vitro GDF11 activated phosphorylation of STAT3. Antagonizing STAT3, with Stattic, in vitro and in vivo, could partially reverse proteolytic pathways including STAT3/socs3 and iNOS/NO in GDF11-meditated muscle wasting. Our findings demonstrate that GDF11 contributes to muscle wasting and the inhibition of its downstream molecule STAT3 shows promise as a therapeutic intervention by which muscle atrophy may be directly prevented in PAH.

Obstructive sleep apnea (OSA) is the most common sleep-related disorder with high prevalence, more complications, great hazard, contributing to reduced quality of life.OSA can cause multiple organs impairment and complications, however, the current research on pulmonary hypertension (PH) is rare.OSA-related pulmonary hypertension (PH) is common in clinical practice, present little is known about the pathophysiology and treatment of OSA-related PH, and the prognosis is poor if left untreated.Based on the current results about OSA-related PH at home and abroad, this paper mainly discusses the prevalence of OSA-related PH, and its possible mechanism and current treatment.

综述近年来中药治疗哮喘气道炎症机制的研究进展.中药复方或单味中药及其提取物可通过适应性免疫调节、抑制抗原呈递、固有免疫调节、抑制氧化应激、提高抗氧化能力、抑制NLRP3炎症小体形成、抑制炎症反应通路等,实现治疗哮喘的抗炎效应.同时通过网络药理学、高效液相色谱、基因组学、蛋白质组学等研究显示有效成分间的相互作用,进一步揭示中药多靶点多通路治疗哮喘气道炎症的机制.

BACKGROUND:Polygonum cuspidatum is a Chinese medicine commonly used to treat phlegm-heat asthma. However, its anti-asthmatic active ingredients and mechanism are still unknown. The aim of this study was to predict the active ingredients and pathways of Polygonum cuspidatum and to further explore the potential molecular mechanism in asthma by using network pharmacology.METHODS:The active ingredients and their targets related to Polygonum cuspidatum were seeked out with the TCM systematic pharmacology analysis platform (TCMSP), and the ingredient-target network was constructed. The GeneCards, DrugBank and OMIM databases were used to collect and screen asthma targets, and then the drug-target-disease interaction network was constructed with Cytoscape software. A target protein-protein interaction (PPI) network was constructed using the STRING database to screen key targets. Finally, GO and KEGG analyses were used to identify biological processes and signaling pathways. The anti-asthmatic effects of Polygonum cuspidatum and its active ingredients were tested in vitro for regulating airway smooth muscle (ASM) cells proliferation and MUC5AC expression, two main symptoms of asthma, by using Real-time PCR, Western blotting, CCK-8 assays and annexin V-FITC staining.RESULTS:Twelve active ingredients in Polygonum cuspidatum and 479 related target proteins were screened in the relevant databases. Among these target proteins, 191 genes had been found to be differentially expressed in asthma. PPI network analysis and KEGG pathway enrichment analysis predicted that the Polygonum cuspidatum could regulate the AKT, MAPK and apoptosis signaling pathways. Consistently, further in vitro experiments demonstrated that Polygonum cuspidatum and resveratrol (one active ingredient of Polygonum cuspidatum) were shown to inhibit ASM cells proliferation and promoted apoptosis of ASM cells. Furthermore, Polygonum cuspidatum and resveratrol inhibited PDGF-induced AKT/mTOR activation in ASM cells. In addition, Polygonum cuspidatum decreased H2O2 induced MUC5AC overexpression in airway epithelial NCI-H292 cells.CONCLUSION:Polygonum cuspidatum could alleviate the symptoms of asthma including ASM cells proliferation and MUC5AC expression through the mechanisms predicted by network pharmacology, which provides a basis for further understanding of Polygonum cuspidatum in the treatment of asthma.

Objectives: Benzo(a)pyrene (BaP) is a ubiquitous air pollutants, and BaP exposure leads to a risk of respiratory diseases. The oversecretion of airway mucus and high expression of mucin 5AC (MUC5AC) are associated with common respiratory disorders caused by air pollution. We aimed to investigate the effect of BaP on MUC5AC expression, especially the mechanisms by which BaP induces MUC5AC gene expression. Methods: The human airway epithelial cell NCI-H292 was used to test the effects of BaP on the expression of MUC5AC in vitro. MUC5AC mRNA and protein expression were assessed with real-time quantitative PCR, immunochemistry, and western blotting. A luciferase assay was conducted to detect the activity of the promoter. The total cellular ROS and mitochondrial ROS were measured by corresponding probes. Small-interfering RNAs were used for gene silencing. AhR-overexpressing cell lines were constructed by transfection with AhR overexpression lentivirus. Results: We found that BaP stimulation upregulated the MUC5AC mRNA and protein levels and activated the ERK pathway. Suppressing ERK with U0126 (an ERK inhibitor) or knocking down ERK with siRNA decreased BaP-induced MUC5AC expression. The luciferase activity transfected with the MUC5AC promoter and cAMP-response element (CRE) was increased after BaP treatment, whereas CREB siRNA suppressed the BaP-induced overexpression of MUC5AC. In addition, BaP increased mitochondrial ROS production, and Mito-TEMP, a mitochondrial ROS inhibitor, inhibited BaP-induced MUC5AC expression and ERK activation. BaP increased the mRNA levels of CYP1A1 and CYP1B1, while Alizarin, a CYP1s inhibitor, suppressed the effects of BaP, including the MUC5AC overexpression, ERK activation and mitochondrial ROS generation. BaP induced the translocation of aryl hydrocarbon receptor (AhR) from the cytoplasm to the nucleus. SiRNA-mediated knockdown or chemical inhibition of AhR decreased the BaP-induced expression of MUC5AC, while the overexpression of AhR significantly enhanced the BaP-induced expression of MUC5AC. ITE, an endogenous AhR ligand, also upregulated the mRNA and protein expression of MUC5AC. Furthermore, resveratrol treatment inhibited the BaP-induced MUC5AC overexpression, AhR translocation, mitochondrial ROS production and ERK pathway activation. Conclusion: Here, we highlighted the crucial role of AhR/mitochondrial ROS/ERK pathway activation in BaP-induced MUC5AC overexpression and identified resveratrol as a promising drug to reduce BaP-induced MUC5AC overexpression.

Background: MUC5AC was recently identified to play important roles in the proliferation and metastasis of malignant mucinous lung tumor cells. Resveratrol (Res), a natural compound with anticancer effects in lung cancer cells, has been reported to inhibit mucin production in airway epithelial cells. This study aimed to investigate the inhibitory effect of Res on MUC5AC expression in lung mucinous adenocarcinoma cells and the potential mechanisms. Methods: Mucus-producing A549 human lung carcinoma cells were used to test the effects of Res on SPDEF and MUC5AC expression. Gene and protein expression was assessed by real-time quantitative PCR (qPCR), immunofluorescence and western blotting assays. SPDEF lentivirus was used to upregulate SPDEF expression levels in mucus-producing A549 human lung carcinoma cells. Cell proliferation was assessed by Cell Counting Kit-8 (CCK8) assay. Results: Res decreased MUC5AC expression in an SPDEF-dependent manner in mucus-producing A549 human lung carcinoma cells, and this change was accompanied by decreased ERK expression and AKT pathway activation. Moreover, SPDEF was found to be overexpressed in lung adenocarcinoma (LUAD), especially in mucinous adenocarcinoma. In-vitro functional assays showed that overexpression of SPDEF reduced the chemosensitivity of A549 cells to cisplatin (DDP). In addition, Res treatment increased A549 cell chemosensitivity to DDP by inhibiting the SPDEF-MUC5AC axis. Conclusion: Our results indicate that the SPDEF-MUC5AC axis is associated with DDP sensitivity, and that Res decreases SPDEF and MUC5AC expression by inhibiting ERK and AKT signaling in A549 cells, which provides a potential pharmacotherapy for the prevention and therapeutic management of mucinous adenocarcinoma.

目的 探讨芪仙汤提取物对苯并芘(BaP)诱导的气道上皮细胞黏蛋白(MUC)5AC的抑制作用及其可能的机制.方法 以人支气管黏液上皮细胞NCI-H292细胞株为研究对象,用BaP刺激细胞构建MUC5AC升高模型(模型组),分别用芪仙汤提取物240 μg/mL(低浓度组)和480 μg/mL(高浓度组)处理细胞株,空白组予二甲基亚砜(DMSO)处理.采用CCK-8法检测芪仙汤提取物处理后细胞活性的变化,实时荧光定量PCR检测MUC5AC和细胞色素P450 1A1(CYP1A1)mRNA水平,免疫印迹法检测MUC5AC、CYP1A1、细胞外信号调节激酶(ERK)1/2、磷酸化ERK(p-ERK)1/2、相对分子质量为90 000核糖体S6激酶(RSK)和磷酸化p90 RSK(p-p90 RSK)蛋白质表达水平,荧光探针检测细胞内活性氧(ROS)水平.结果 浓度≤480 μg/mL的芪仙汤提取物对NCI-H292细胞株活力无显著影响.与模型组相比,芪仙汤低、高浓度组MUC5AC蛋白质和MUC5AC mRNA水平均显著降低(P＜0.05或0.01),ROS荧光强度值均显著降低(P＜0.05或0.01),CYP1A1 mRNA水平均显著降低(P值均＜0.01),p-ERK 1/2和p-p90 RSK蛋白质水平亦显著降低(P值均＜0.01).结论 芪仙汤可下调BaP诱导的人气道上皮细胞MUC5AC基因和蛋白质高水平表达,其机制与抑制芳香烃受体活化介导的ROS和ERK通路激活相关.

Background Pulmonary rehabilitation is a crucial part of the nonpharmacological treatment of stable chronic obstructive pulmonary disease (COPD), but management remains problematic. WeChat could serve as a useful tool in patient management. Baduanjin is a popular exercise in China that is usually applied in pulmonary rehabilitation, which has been confirmed to be effective in improving lung function and life quality. Objective This study aimed to explore the efficiency of WeChat in the management of Baduanjin exercise in COPD patients. Methods A total of 200 patients from the respiratory department of Putuo Hospital participated in the Baduanjin rehabilitation project from September 2018 to October 2019, and were randomly assigned to the WeChat and control groups and followed up using the WeChat platform or telephone for 12 weeks. The frequency of Baduanjin exercise, lung function (percentage of forced expiratory volume in 1 second predicted, FEV1% predicted), and COPD assessment test (CAT) scores were collected and compared between the two groups. The number of message exchanges and a satisfaction survey on the WeChat platform were used to assess the feasibility of WeChat management outside the hospital. Results The Baduanjin exercise frequency significantly differed between the control group and WeChat group (F=33.82, P<.001) and across various time points (F=214.87, P<.001). After the follow-up on WeChat, there were fewer patients not performing Baduanjin exercise. The FEV1% predicted value significantly differed before and after Baduanjin exercise in the control group (Z=−3.686, P<.001) and the WeChat group (Z=−6.985, P<.001). A significant difference in the FEV1% predicted value was observed after Baduanjin exercise between the two groups (Z=−3.679, P<.001). The CAT score significantly differed before and after Baduanjin exercise in the control group (Z=−4.937, P<.001) and the WeChat group (Z=−5.246, P<.001). A significant difference in the CAT score was observed after Baduanjin exercise between the two groups (Z=−5.246, P<.001). The number of completed Baduanjin exercises, lung function, and CAT scores in active patients were higher than those in nonactive patients. All satisfaction survey items were scored with more than 4 points. Among the items, the highest score (mean 4.54, SD 0.77) was for continued WeChat management, followed by the effective management of Baduanjin exercise (mean 4.46, SD 0.87). The patients in the WeChat group showed much higher enthusiasm for and compliance with Baduanjin exercise, resulting in better life quality and lung function. The patients were very satisfied with the WeChat management because of the obvious curative effect and home feeling. Conclusions The WeChat platform provided a feasible, effective, and sustainable management plan for Baduanjin rehabilitation. Trial Registration Chinese Clinical Trial Registry ChiCTR1900028248; http://www.chictr.org.cn/showprojen.aspx?proj=46995

Purpose: Arl4c is overexpressed in several cancer tissues and is involved in cancer development. Nevertheless, the exact mechanism that regulates Arl4c expression in lung cancer has not been fully elucidated. The aim of this study was to investigate the regulatory mechanism of Arl4c and to explore potential chemotherapeutic drugs targeting Arl4c. Methods: Immunohistochemistry was used to examine Arl4c expression levels in human lung adenocarcinoma cancer specimens. Protein expression was detected by western blot. Overexpression of Arl4c-Flag protein was used to detect the ubiquitination of Arl4c. A short interfering RNA against Arl4c was used for gene silencing. Results: Arl4c was overexpressed in lung cancer tissues, and knockdown of Arl4c expression by siRNA decreased lung cancer A549 and 95-D cell proliferation. In addition, Arl4c expression was downregulated via inhibition of the AKT pathway in A549 and 95-D cells, whereas exposure to benzo (a) pyrene (a carcinogen in smoke) increased Arl4c expression in 16HBE cells via AKT activation. Finally, we found that chemotherapy drug hydroxycamptothecin (HCPT) could decrease Arl4c expression levels by inhibiting the activation of the AKT pathway in A549 and 95-D cells. Moreover, accumulation of ubiquitinated Arl4c protein was increased by HCPT and LY294002 (an AKT inhibitor) treatment whereas the proteasome inhibitor MG-132 attenuated the inhibitory effect of HCPT and LY294002 on Arl4c expression. Conclusion: Here, we highlighted the AKT pathway as an important regulatory pathway for Arl4c expression in lung cancer cells and identified HCPT as a promising drug for lung adenocarcinoma treatment that functioned by targeting Arl4c expression.

Background: Loss of the epithelial barrier is characterized by a reduction in E-cadherin expression and is a hallmark of asthma. Qi-xian decoction (QXT) is a Chinese medicinal formula that has been used to effectively treat asthma. This study aimed to investigate the effect of QXT on E-cadherin expression in human lung epithelial 16HBE cells and ovalbumin-challenged mice and to explore the underlying molecular mechanism. Methods: Ovalbumin (OVA)-induced mice were used as a model of asthma. Real-time PCR and Western blotting were utilized to examine mRNA and protein levels. Lung tissue reactive oxygen species (ROS) levels were evaluated using dichloro-dihydro-fluorescein diacetate (DCFH-DA). Serum superoxide dismutase (SOD) and the total antioxidant capacity (TAOC) were measured via enzyme-linked immunosorbent assay (ELISA)-based analyses. 16HBE cells were utilized to explore the effect of QXT or hydrogen peroxide (H2O2) on the expression of E-cadherin in vitro. Results: We found that QXT treatment increased E-cadherin expression and decreased extracellular-signal-regulated kinase (ERK) phosphorylation levels in the lung tissues of OVA-challenged mice. QXT also downregulated ROS levels and increased serum SOD and TAOC levels in OVA-challenged mice. In vitro studies demonstrated that increased ROS generation induced by H2O2 resulted in decreased E-cadherin expression levels in 16HBE cells, which was attenuated by inhibition of ERK signaling. Moreover, the H2O2-induced downregulation of E-cadherin expression, increased ROS generation, and ERK activation in 16HBE cells were restored by treatment with QXT water or ethanol extract. Conclusions: These data demonstrate that one mechanism by which QXT protects against asthma is to restore E-cadherin expression in vivo and in vitro by inhibiting ROS-mediated ERK activation.