Chronic obstructive pulmonary disease (COPD) is the third leading cause of death worldwide and is characterized by persistent inflammation, microbiota dysbiosis, and excessive neutrophil extracellular trap (NET) formation. Rosavin, a natural phenylpropanoid glycoside, exhibits anti-inflammatory and immunomodulatory activities, but its therapeutic potential in COPD remains unclear. A COPD rat model was induced by intratracheal lipopolysaccharide instillation combined with chronic passive cigarette smoke exposure. Rosavin (50 or 100 mg/kg) or the NF-κB inhibitor BAY 11-7082 (3 mg/kg) was administered for treatment. Pulmonary function tests (FEV1/FVC, PEF, and airway resistance), histopathological evaluation (HE and PAS staining), bronchoalveolar lavage fluid (BALF) inflammatory cell counts, ELISA-based cytokine assays, and oxidative stress markers (MDA, MPO, and SOD) were systematically assessed. NET formation was evaluated using MPO-DNA ELISA, Western blotting, and immunofluorescence for CitH3 and MPO/IL-17 colocalization. In addition, lung microbiota composition was analyzed by 16 S rRNA gene sequencing. Cigarette smoke extract (CSE)–stimulated BEAS-2B cells were used to assess the direct effects of Rosavin on NF-κB activation in vitro. Rosavin significantly inhibited NF-κB activation, improved lung function, and reduced structural damage, oxidative stress, and inflammatory cytokines in COPD rats. It also suppressed NET formation, including IL-17-enriched NETs, by downregulating MPO, NE, and CitH3. In BEAS-2B cells, Rosavin similarly reduced CSE-induced NF-κB activation and cytokine release. Microbiota profiling showed decreased diversity and enrichment of Fusobacterium nucleatum in COPD rats, whereas Rosavin restored beneficial taxa such as Lactobacillus spp. BAY 11-7082 produced comparable effects, supporting NF-κB inhibition as a key mechanism. Rosavin ameliorates COPD-associated pathology through integrated mechanisms involving NF-κB inhibition, reduction of IL-17-enriched NET formation, and modulation of lung microbiota composition. These findings identify Rosavin as a promising multi-target therapeutic candidate for COPD.

BACKGROUND:Asthma is a chronic inflammatory airway disease characterized by epithelial barrier dysfunction, mucus hypersecretion, and type 2-skewed immune responses. Macrophages play a critical role in shaping airway inflammation and tissue remodeling, yet the molecular mechanisms regulating macrophage activation in allergic asthma remain incompletely understood. The vitamin D receptor (VDR) is an immune-modulatory transcription factor expressed in macrophages, but its macrophage-specific function and regulatory mechanisms in allergic airway inflammation remain unclear. METHODS:We investigated the role of macrophage VDR signaling using a chronic house dust mite (HDM)-induced asthma model combined with myeloid-specific Vdr conditional knockout mice. Airway inflammation, epithelial remodeling, and macrophage polarization were assessed by histological, molecular, and flow cytometric analyses. Mechanistic studies were performed to identify nuclear localization sequences (NLSs) of VDR and to evaluate their interaction with the nuclear import adaptor KPNA1. RESULTS:HDM exposure increased VDR expression in macrophages both in vivo and in vitro. Myeloid-specific deletion of Vdr significantly attenuated airway inflammation, reduced inflammatory cell recruitment, and alleviated epithelial barrier disruption and mucus metaplasia. Mechanistically, VDR promoted macrophage M2-like polarization in response to HDM stimulation. We further identified a previously unrecognized C-terminal nuclear localization sequence spanning amino acids 387-412 that is required for VDR nuclear translocation and macrophage polarization. Structural modeling and biochemical analyses indicated that phosphorylation within this region enhances the interaction between VDR and the nuclear import adaptor KPNA1, thereby facilitating nuclear import of VDR. CONCLUSIONS:These findings identify macrophage-intrinsic VDR signaling as an important regulator of allergic airway inflammation and reveal a phosphorylation-dependent mechanism controlling VDR nuclear trafficking and macrophage polarization.

Background Sarcopenia is a well-established prognostic factor in advanced lung cancer; however, most studies rely on single time-point measurements, and few have captured longitudinal changes in skeletal muscle mass during treatment. Consequently, the prognostic value of dynamic SMI changes remains poorly characterized. This study aimed to characterize longitudinal SMI trajectories and assess their association with PFS. Methods This retrospective cohort study included 316 patients with advanced lung cancer. SMI was quantified using serial chest CT scans at the T12 level at baseline, 3, 6, and 12 months. The rate of SMI change was calculated as percentage change per month. Optimal cut-off values were determined using maximally selected rank statistics. Progression-free survival (PFS) was analyzed using Kaplan–Meier and Cox regression models. Results During follow-up, 111 of 316 patients (35.1%) experienced disease progression, with a 1-year PFS rate of 61.6%. Baseline SMI was significantly associated with PFS (HR = 0.895, P < 0.001); however, its prognostic effect attenuated over time and became non-significant beyond one year (P = 0.682). In contrast, SMI declined in 310 patients (98.1%), following a non-linear "fast–slow–fast" trajectory. Early-phase SMI decline was strongly associated with PFS (HR = 0.849, P < 0.001), with an optimal cut-off of 0.89% per month. Each 1% per month reduction in the rate of SMI decline (i.e., slower muscle loss) was associated with a 15.1% lower risk of disease progression. Longitudinal SMI change remained consistently associated with PFS throughout follow-up. Conclusions Longitudinal decline in skeletal muscle mass provides clinically meaningful prognostic information beyond baseline measurements in advanced lung cancer. Serial SMI assessment may serve as a useful tool for risk stratification and disease monitoring.

While neutrophils represent a prominent myeloid component in non-small cell lung cancer (NSCLC), the specific immunosuppressive functions of N2-polarized neutrophils and their mechanistic interactions with CD8⁺ T cells remain incompletely characterized. Furthermore, the development of clinically applicable models for prognostic stratification and immunotherapy response prediction, grounded in these molecular interactions, represents a critical unmet need. We integrated large-scale single-cell RNA sequencing datasets to delineate the tumor immune microenvironment, performing pathway enrichment and cell-cell communication analyses. Key molecular features derived from these interactions were employed to construct a deep neural network model. This model was trained and validated on bulk RNA sequencing cohorts to predict immunotherapy response. Additionally, we developed the N2_Neu-CD8⁺ Tex Loop Score (NTLS) for prognostic assessment and evaluated its pan-cancer applicability. Our analysis revealed a previously uncharacterized positive feedback loop between N2 neutrophils and exhausted CD8⁺ T cells (Tex). Neutrophil-derived ICAM1 engages with ITGAL/ITGB2 on CD8⁺ T cells, suppressing their NF-κB signaling and reinforcing the exhausted phenotype. In a feed-forward manner, Tex-derived CCL5 signals via CCR1 on N2 neutrophils, activating their NF-κB pathway and further upregulating ICAM1 expression. This ICAM1–Integrin and CCL5-CCR1 axis creates a self-sustaining immunosuppressive circuit. A deep learning model, built upon genes central to this loop, accurately predicted immunotherapy outcomes in NSCLC and melanoma. The derived NTLS score proved effective for prognostic stratification and was validated across multiple independent cohorts and cancer types. This study defines a pathogenic positive feedback loop, driven by ICAM1–Integrin and CCL5–CCR1 interactions, through which N2 neutrophils and Tex cells cooperatively establish an immunosuppressive niche that drives immunotherapy resistance. The computational models we developed, based on this molecular circuitry, offer robust tools for patient stratification and hold significant translational promise.

Background:This study aimed to explore the regulation of Ras-related C3 botulinum toxin substrate1 (Rac1) on the intestinal barrier function in colitis and explore its molecular mechanism of regulation on tight junctions. Methods:A dextran sulfate sodium (DSS)-induced colitis mouse model was used. The diseases activity index (DAI) was calculated daily. Epithelial permeability was measured. Colon sections were stained with hematoxylin and eosin, and the histological severity was analysed. Reverse transcription polymerase chain reaction (RT-PCR) was used to analyse the messenger ribonucleic acid (mRNA) level of Rac1, nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1), and occludin in the colon. Western blot was used to detect occludin protein expression. Results:Colitis mice showed increased DAI and histological scores, reduced colon length, and impaired epithelial permeability, which were significantly alleviated by the administration of Rac1 inhibitor NSC23766. The level of inflammatory genes including interleukin 6 (IL-6), myeloperoxidase and NOX1 in the colon tissue were elevated in colitis mice, while the administration of NSC23766 remarkably reduced the expression of these genes. Western blot analysis showed that the occludin protein level was suppressed by DSS, while NSC23766 treatment restored the expression of occludin in DSS mice. Conclusions:Rac1 inhibitor NSC23766 attenuates symptoms, colonic inflammation, and intestinal permeability in a DSS-induced colitis model. These effects may be attributed to the suppression of inflammatory responses and DSS-induced damage of intestinal integrity.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, irreversible interstitial lung disease with limited effective treatment options. Dihydroartemisinin (DHA), a water-soluble derivative of artemisinin primarily known for its antimalarial properties, has shown potential antifibrotic effects, although its mechanisms remain unclear. To investigate the therapeutic effects and underlying molecular mechanisms of DHA in IPF. Network pharmacology and single-cell transcriptomic analyses were used to identify DHA-related target molecules. Molecular docking was performed to assess the binding affinity between DHA and candidate proteins. A bleomycin-induced mouse model of pulmonary fibrosis and a TGF-β-stimulated primary lung fibroblast model were used for in vivo and in vitro validation, respectively. DHA inhibited fibroblast proliferation and activation by downregulating the calcium signaling pathway. Protein–protein interaction analysis identified CALM1, CAMK,PPP3CA, and NFAT1 as central targets. DHA demonstrated strong binding affinity to these targets in docking analyses. Histopathological staining revealed significant attenuation of bleomycin-induced pulmonary fibrosis, with reduced expression of COL1A1, α-SMA, and Fibronectin. In vitro experiments demonstrated that DHA significantly inhibited the proliferation and migration of primary fibroblasts while concurrently downregulating COL1A1, α-SMA, and Fibronectin expression. Notably, the CAMK inhibitor exhibited effects comparable to those of DHA, suggesting a potential shared mechanistic pathway in modulating fibroblast activity. DHA alleviates pulmonary fibrosis by inhibiting the calcium signaling pathway, thereby suppressing fibroblast proliferation and activation.

OBJECTIVES:Reusable colonoscopes pose a risk of iatrogenic infections due to improper disinfection and maintenance, prompting the development of disposable colonoscopes. However, direct comparisons between disposable and reusable colonoscopes remain limited. This pilot study aimed to evaluate the technical performance of disposable colonoscopes compared to reusable ones for routine colon examinations. METHODS:This randomized controlled, noninferiority study was conducted at two endoscopy centers. Patients requiring colonoscopy were randomly assigned to either the disposable or reusable colonoscope group. The primary outcome was the successful completion rate of colonoscopy between the groups, with a noninferior margin of -10%. Secondary outcomes included image characteristics, technical maneuverability, colonoscopy performance measures, and adverse events. RESULTS:A total of 116 patients underwent colonoscopy (58 in each group). The successful completion rate of colonoscopy was 100% in both groups (difference: 0% [95% confidence interval -6.21% to 6.21%]), confirming noninferiority. Although the disposable colonoscope group had lower performance scorings in imaging characteristics, technical maneuverability, and longer operating time compared to the reusable colonoscope group, no significant differences were observed in cecal intubation rate, polyp detection rate, polyp characteristics, or adverse event rate. Additionally, experienced endoscopists achieved proficiency with disposable colonoscopes after approximately 10 cases, requiring minimal training. CONCLUSION:With further technical advancements, disposable colonoscopes may serve as a safe and viable alternative to reusable colonoscopes for routine colon examinations in certain clinical scenarios.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease without efficient treatment. Fuzi has anti-inflammatory and immunomodulatory properties. However, the bioactive compounds and mechanisms of fuzi in the treatment of UC are not completely understood. The active components of fuzi were retrieved from Traditional Chinese Medicine Database System Pharmacology and Analysis Platform; PharmMapper was used to predict the targets of the active components of fuzi; UC-related disease targets were obtained from Online Mendelian Inheritance in Man and Genecards databases, and Venny 2.1 was used to obtain common targets; Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were performed on the common targets using R 4.0.2. STRING and Cytoscape 3.9.0 was used to construct a protein-protein interaction (PPI) network for the intersection targets. We then determined the role of the candidate molecule from fuzi, Higenamine (Hig), in a mouse model of dextran sulfate sodium (DSS)-induced colitis. In total, 21 active components and 420 corresponding targets of fuzi were obtained, of which 224 common targets were identified by intersecting with UC-related targets. The GO, KEGG, and PPI results suggested that fuzi and Hig may target RAC-alpha serine/threonine-protein kinase (AKT) to regulate the phosphoinositide-3-kinase (PI3K)/AKT pathway in UC. Animal experiments have shown that Hig treatment greatly reduced DSS-induced colitis, as measured by the disease activity index score, colonic inflammation, and intestinal barrier integrity. Mechanistically, Hig downregulated the DSS-induced PI3K-AKT signaling pathway by inhibiting AKT phosphorylation. Altogether, Hig alleviated DSS-induced colitis in mice, possibly by inhibiting colon inflammation and improving the intestinal barrier by regulating the PI3K-AKT signaling pathway. The active component Hig from fuzi is likely to play a role in the treatment of UC.

Background: Extracellular high mobility group box 1 (HMGB1) is a key mediator in driving allergic airway inflammation and contributes to asthma. Yet, mechanism of HMGB1 secretion in asthma is poorly defined. Pulmonary metabolic dysfunction is recently recognized as a driver of respiratory pathology. However, the altered metabolic signatures and the roles of metabolic to allergic airway inflammation remain unclear. Methods: Male C57BL/6 J mice were sensitized and challenged with toluene diisocyanate (TDI) to generate a chemically induced asthma model. Pulmonary untargeted metabolomics was employed. According to results, mice were orally administered allopurinol, a xanthine oxidase (XO) inhibitor. Human bronchial epithelial cells (16HBE) were stimulated by TDI-human serum albumin (HSA). Results: We identified the purine metabolism was the most enriched pathway in TDI-exposed lungs, corresponding to the increase of xanthine and uric acid, products of purine degradation mediated by XO. Inhibition of XO by allopurinol ameliorates TDI-induced oxidative stress and DNA damage, mixed granulocytic airway inflammation and Th1, Th2 and Th17 immunology as well as HMGB1 acetylation and secretion. Mechanistically, HMGB1 acetylation was caused by decreased activation of the NAD+-sirtuin 1 (SIRT1) axis triggered by hyperactivation of the DNA damage sensor poly (ADP-ribose)-polymerase 1 (PARP-1). This was rescued by allopurinol, PARP-1 inhibitor or supplementation with NAD+ precursor in a SIRT1-dependent manner. Meanwhile, allopurinol attenuated Nrf2 defect due to SIRT1 inactivation to help ROS scavenge. Conclusions: We demonstrated a novel regulation of HMGB1 acetylation and secretion by purine metabolism that is critical for asthma onset. Allopurinol may have therapeutic potential in patients with asthma.

This study aimed to compare the effectiveness of chemotherapy in different histological types of pancreatic cancer using data collected from the Surveillance, Epidemiology, and End Results (SEER) database. Patients who were diagnosed with pancreatic cancer between 2004 and 2015 were selected from the SEER database. Propensity score matching (PSM) was employed to minimize the selection bias. The Kaplan-Meier survival curves and the log-rank test were utilized to compare the overall survival (OS) and cancer-specific survival (CSS) among different groups. Of the 7,653 pancreatic cancer patients, both OS and CSS were higher in the chemotherapy group than those in the non-chemotherapy group (p < 0.001). After PSM, 2381 pairs were generated. The Kaplan-Meier survival curved indicated that both OS and CSS for pancreatic ductal adenocarcinoma (PDAC), pancreatic adenosquamous carcinoma (PASC), and pancreatic mucin-producing adenocarcinoma (PMPAC) (p < 0.001) in the chemotherapy group were superior to those in the non-chemotherapy group, while there was no significant difference in pancreatic mucinous adenocarcinoma (PMAC) (p = 0.2586). Compared with PASC and PMPAC, PDAC exhibited longer OS and CSS. The results of statistical analysis showed that PASC tumors were mainly poorly differentiated, and the majority of patients with PMPAC had distant metastasis. Chemotherapy could prolong pancreatic cancer patients' survival, especially for patients with advanced disease. PMPAC patients had a higher rate of metastasis, accompanying with the worse survival.

Nicotinamide adenine dinucleotide (NAD+) is an essential element in cellular metabolism that regulates fundamental biological processes. Growing evidence suggests that a decline in NAD+ is a common pathological factor in various diseases and aging. However, its role in airway epithelial barrier function in response to asthma remains underexplored. The current study aims to explore the efficacy of restoring cellular NAD+ concentration through supplementation with the NAD+ precursor, nicotinamide mononucleotide (NMN), in the treatment of allergic asthma and to investigate the role of SIRT3 in mediating the effects of NAD+ precursors. In this research, NMN alleviated airway inflammation and reduced mucus secretion in house dust mite (HDM)-induced asthmatic mice. It also mitigated airway epithelial barrier disruption in HDM-induced asthma in vitro and in vivo. But inhibition of SIRT3 expression abolished the effects of NMN. Mechanistically, HDM induced SIRT3 SUMOylation and proteasomal degradation. Mutation of these two SIRT3 SUMO modification sites enhanced the stability of SIRT3. Additionally, SIRT3 was targeted by SENP1 which acted to de-conjugate SUMO. And down-regulation of SENP1 expression in HDM-induced models was reversed by NMN. Collectively, these findings suggest that NMN attenuates airway epithelial barrier dysfunction via inhibiting SIRT3 SUMOylation in asthma. Blockage of SIRT3 SUMOylation emerges as for the treatment of allergic asthma.

Lead (Pb) is a non-biodegradable environmental pollutant that can lead to neurotoxicity by inducing neuroinflammation. Microglial activation plays a key role in neuroinflammation, and microglial migration is one of its main features. However, whether Pb affects microglial migration has not yet been elucidated. Herein, the effect of Pb on microglial migration was investigated using BV-2 microglial cells and primary microglial cells. The results showed that cell activation markers (TNF-α and CD206) in BV-2 cells were increased after Pb treatment. The migration ability of microglia was inhibited by Pb. Both store-operated calcium entry (SOCE) and the Ca2+ release-activated Ca2+ (CRAC) current were downregulated by microglia treatment with Pb in a dose-dependent manner. However, there was no statistical difference in the protein levels of stromal interaction molecule (STIM) 1, STIM2, or Ca2+ release-activated Ca2+ channel protein (Orai) 1 in microglia. The external Ca2+ influx and cell migration ability were restored to a certain extent after overexpression of either STIM1 or its CRAC activation domain in microglia. These results indicated that Pb inhibits microglial migration by downregulation of SOCE and impairment of the function of STIM1.

To the Editor: Asthma is a complex chronic inflammatory lung disease, and its pathogenesis is not fully understood. It is believed that airway epithelium primarily acts as a defensive barrier to protect the lung from allergens, avoiding the asthma. The integrity of the airway epithelial barrier is dependent on tight junctions comprised of zona occludens (ZO) 1 to 3, occludin, and claudin 1 to 5, and adherens junctions mainly formed by E-cadherin, β-catenin, and junctional adhesion molecule, which maintain apicobasal polarity of bronchial epithelial cells. Our team has previously confirmed that house dust mite (HDM) disrupts the airway epithelial barrier in vivo and in vitro and that thymic stromal lymphopoietin (TSLP) plays an essential role in this process.[1] Autophagy, associated with various human diseases, is a highly conserved and lysosome-dependent process that selectively degrades and recycles the targeted organelles within lysosomes. The role of autophagy in allergic asthma is still under debate. In this study, we hypothesized that the autophagy process plays a critical role in HDM-induced asthma, and HDM induces autophagy-related genes (ATGs), which are associated with epithelial barrier disruption; and we utilized HDM to stimulate human bronchial epithelial cells (HBECs) to evaluate the role of autophagy in HDM-induced asthma. The normal human bronchial epithelial cell line, HBE-135o (YaJi Biological, Shanghai, China), was administrated with 200 nmol/L rapamycin (Solarbio, Beijing, China) for 24 h or 400 IU/mL HDM (ALK-Abello A/S, H⊘rsholm, Denmark) for 3 to 48 h. The HBECs were also treated with small interfering RNA (Keygen, Jiangsu, China) for 12 h. Epithelial barrier integrity was assessed by measuring transepithelial electrical resistance (TEER) and fluorescein isothiocyanate-dextran (FITC-Dx) flux across the monolayers of cultured epithelial cells. Besides, an immunofluorescence (IF) assay was used for detecting the distribution of ZO-1, E-cadherin, and β-catenin. Western blotting was performed for detecting the expression of light chain 3a/b (LC3a/b), P62, autophagy related protein 5 (Atg5), Atg12, Atg7, Atg16L1. The co-immunoprecipitation (Co-IP) assay and proximity ligation assay (PLA) were utilized in our study for confirming the relationship between Atg5 and TSLP. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to test the messenger RNA. Statistical analysis was carried out using the GraphPad Prism 8 software package (GraphPad by Dotmatics, San Diego, CA, USA). The data were expressed as the mean ± standard error (SE). One-way analysis of variance followed by Bonferonni post-hoc test for multiple comparisons was used to compare differences between the groups. A value of P < 0.05 was considered statistically significant. FITC-Dx permeability and TEER results showed that inducing autophagy in HBECs with rapamycin or stimulating them with HDM both increased permeability and decreased electrical resistance [Figure 1A]. IF assay found the cell–cell junction proteins redistributed after being treated with HDM or rapamycin [Figure 1B]. Western blotting results showed that LC3a/b was upregulated significantly after stimulating HBECs with HDM for 12 and 24 h, while P62 expression remained unchanged [Figure 1C]. Besides, western blot results of the Atgs, including Atg5, Atg7, Atg16L1, and Atg12, found the expressions of Atg5 and Atg12 both increased after stimulating HBECs with HDM for 12 h [Figure 1D]. In order to clarify the roles of Atg5 and Atg12 in HDM-induced asthma, we used small interfering RNA to interfere with the expressions of Atg5 or Atg12 in HBECs, and found that interfering with Atg5 expression decreased the permeability and increased the TEER [Figure 1E], and also decreased the level of TSLP mRNA [Figure 1F]. However, the same results were not observed after Atg12 interfering treatment. In further experiment, IF suggested that interfering with Atg5 alleviated the redistribution of ZO-1, E-cadherin, and β-catenin induced by HDM [Figure 1G]. Co-IP assay and PLA showed that Atg5 can interact with TSLP in HDM-induced HBECs [Figures 1H and 1I]. These results showed that Atg5 gene expression was upregulated in asthma and was associated with airway epithelial barrier disruption.Figure 1: House dust mite disrupts the airway epithelial barrier by affecting the expression of thymic stromal lymphopoietin. (A) FITC-Dx permeability and TEER were used to determine the epithelial barrier integrity in HBECs after being treated with HDM or Rap for 24 h. (B) The distribution of ZO-1, E-cadherin, and β-catenin monitored by IF staining. Green represents ZO-1 and E-cadherin, red represents β-catenin, and blue represents the nucleus (confocal microscopic analysis, original magnification × 1000). (C) Western blotting of LC3a/b and P62 in HBECs stimulated with HDM for 3 to 48 h. (D) Western blotting of Atg7, Atg16L1, Atg12, and Atg5 in HBECs stimulated with HDM for 6 to 24 h. (E) FITC-Dx permeability and TEER were used to determine the epithelial barrier integrity after HBECs were treated with HDM plus siAtg5 or siAtg12. (F) TSLP mRNA level of HBECs after being treated with HDM and siAtg5 or siAtg12. (G) Distribution of ZO-1, E-cadherin, and β-catenin in HBECs after being treated with HDM plus siAtg5 or siAtg12. Green represents ZO-1 and E-cadherin, red represents β-catenin, and blue represents the nucleus (confocal microscopic analysis, original magnification × 1000). (H) The association between TSLP and Atg5 was measured using Co-IP assay. (I) The association between TSLP and Atg5 was measured using PLA (original magnification × 1000). Data are presented as the mean ± standard error; n ≥3. ∗ P <0.05, † P <0.01, ‡ P <0.0001 vs. control group, § P <0.05, || P <0.01 vs. HDM group. Atg 5/7/12/16L1: Autophagy-related gene 5/7/12/16L1; FITC-Dx: Fluorescein isothiocyanate-dextran; HBECs: Human bronchial epithelial cells; HDM: House dust mite; IF: Immunofluorescence; Co-IP: Co-immunoprecipitation; IgG: Immunoglobulin G; LC3a/b: Light chain 3a/b; mRNA: Messenger RNA; NC: Negative control; n.s.: No statistically significant difference; PBS: Phosphate buffered saline; PLA: Proximity ligation assay; Rap: Rapamycin; si: Small interfering RNA; TEER: Transepithelial electrical resistance; TSLP: Thymic stromal lymphopoietin; ZO-1: Zona occludens-1.Autophagy is a conserved process in which substrates are endocytosed by the double membrane vesicles and transported to the lysosomes for degradation. A recent research found that the autophagy receptor P62 can mediate the anti-inflammatory reaction in lipopolysaccharide (LPS)-induced macrophages; it has revealed a novel mechanism of macrophage inflammation.[2] As for asthma, multiple pieces of evidence indicate that autophagy plays a significant role. It is believed that autophagy is associated with epithelial barrier disruption,[3] but the mechanisms of autophagy affecting airway epithelial disruption in allergic asthma are not fully understood. In the current study, we have demonstrated that Atg5 expression increased in HBECs induced by HDM. Moreover, our results also suggested that HDM-induced Atg5 was associated with TSLP, which was found upregulated in HBEC treated with HDM. Hence, we considered that Atg5 may regulate the expression of inflammatory factor TSLP, which has not been reported before; our findings have filled the current blank. Han et al[4] found that TSLP increased autophagy-related factors, such as Beclin-1, LC3-II, P62, and Atg5, to activate autophagy in T cells via Janus Kinase 1 (JAK1)/JAK2/signal transducers and activators of transcription 5/c-Jun NH (2)-terminal kinases/phosphoinositide 3-kinase pathway in LPS-induced acute liver injury, indicating that TSLP and Atg5 have an interaction relationship, but there may be different regulatory mechanisms in different diseases. There are several limitations in our research. First, the Atg5-/- mice is necessary to be used for further investigation according to our results. And, a known method, air-liquid interface culture, for airway epithelial cells experiments is lacking in our research, and should be used in future studies to validate our results. In conclusion, HDM-induced Atg5 interacts with TSLP, an important factor that is responsible for airway inflammation, during the airway epithelial barrier disruption induced by HDM; we concluded that the Atg5 plays an important role in HDM-induced asthma, and Atg5 may serve as a therapeutic target in suppressing the airway epithelial barrier disruption to protect asthmatic patients from allergic antigens. Funding This study was supported by grants from the National Natural Science Foundation of China (Nos. 81970032 and 81870026). Conflicts of interest None.

Background:Neutrophils can be rapidly recruited and are largely abundant in the airways of patients with asthma. However, whether the polarization and chemotaxis of neutrophils in patients with asthma are abnormal, and the underlying mechanisms, have not been clarified. Pseudopods formation is the initial step of neutrophils' polarization, ezrin, radixin and moesin (ERM) play an important role in the polarization of neutrophils. As an important signaling molecule in cell physiological processes, Ca2+ has been shown to be involved in the polarity changes of neutrophils. This study thus aimed to explore polarization and chemotaxis of neutrophils in patients with asthma and the underlying mechanism.Methods:Fresh neutrophils were isolated using standard separation protocols. The polarization and chemotactic activity of neutrophils were observed using Zigmond chamber and Transwell migration assay under linear concentration gradients of N-formyl-methionine-leucine-phenylalanine (fMLP) or interleukin (IL)-8. The distribution of calcium, ERMs and F-actin in neutrophils were observed by confocal laser scanning microscope. The expression of the main components of ERMs (moesin and ezrin) was detected with reverse transcription-polymerase chain reaction (RT-PCR).Results:Compared with those in the healthy control group, the polarization and chemotaxis of neutrophils in the venous blood of patients with asthma were significantly increased, and the expression and distribution of cytoskeletal proteins F-actin and ezrin were abnormal. The expression and function of key components of store-operated calcium entry (SOCE), stromal interaction molecule 1 (STIM1), STIM2, and Orai1 of neutrophils in patients with asthma were significantly increased.Conclusions:The polarization and chemotaxis of neutrophils in the venous blood of patients with asthma are increased. This may be due to the abnormal expression and distribution of ERM and F-actin as a result of abnormal SOCE function.

The potential role of polycomb chromobox 4 (Cbx4), as a small ubiquitin-like ligase (SUMO) E3 ligase, in the development and exacerbation of asthma remains unclear. Hypoxia inducible factor-1 (HIF-1) is a key transcription factor in the cellular response to hypoxia and contributes to the pathogenesis and progression of a range of diseases, including asthma. Here, we aimed to investigate the interaction of Cbx4 with Hypoxia inducible factor-1α (HIF-1α) and the potent mechanism of action in asthma progression. In present study, in vitro and ex vivo results demonstrated that Cbx4 interacts with HIF-1α protein through its SUMO E3 ligase activity and enhances the sumoylation, which increases HIF-1 transactivation through Cbx4 and promotes the differentiation of Th9 cells, then in turn promotes the process of asthma. Treatment of inhibitors targeting SUMO E3 ligase activity of Cbx4 or HIF-1α can effectively reduce HIF-1α activation and differentiation of Th9 cells, which further attenuates the asthma in mouse model. Current results collectively demonstrated Cbx4 can govern HIF-1α to involve in Th9 cell differentiation promoting asthma by its SUMO E3 ligase activity, providing a new direction for clinical treatment of asthma.

Background and purpose: Mitochondrial dysfunction is an essential part of the pathophysiology of asthma, and potential treatments that target the malfunctioning mitochondria have attracted widespread attention. We have previously demonstrated that aberrant epithelial 8-catenin signaling played a crucial role in a toluene diisocyanate (TDI)-induced steroid-insensitive asthma model. The objective of this study was to determine if the mitochondrially targeted antioxidant mitoquinone(MitoQ) regulated the activation of 8-catenin in TDI-induced asthma.Method: Mice were sensitized and challenged with TDI to generate a steroid-insensitive asthma model. Human bronchial epithelial cells (16HBE) were exposed to TDI-human serum albumin (HSA) and ethidium bromide(EB) to simulate the TDI-induced asthma model and mitochondrial dysfunction.Results: MitoQ dramatically attenuated TDI-induced AHR, airway inflammation, airway goblet cell metaplasia, and collagen deposition and markedly protected epithelial mitochondrial functions by preserving mass and diminishing the production of reactive oxygen species (ROS). MitoQ administration stabilized 8-catenin destruction complex from disintegration and inhibited the activation of 8-catenin. Similarly, YAP1, an important constituent of 8-catenin destruction complex, was inhibited by Dasatinib, which alleviated airway inflammation and the activation of 8-catenin, and restored mitochondrial mass. In vitro, treating 16HBE cells with EB led to the activation of YAP1 and 8-catenin signaling, decreased the expression of glucocorticoid receptors and upregulated interleukin (IL)-18, IL6 and IL-8 expression.Conclusion: Our results indicated that mitochondria mediates airway inflammation by regulating the stability of the 8-catenin destruction complex and MitoQ might be a promising therapeutic approach to improve airway inflammation and severe asthma. Availability of data and materials: The data that support the findings of this study are available from the corresponding author upon reasonable request. Some data may not be made available because of privacy or ethical restrictions.

Lung fibroblast activation is associated with airway remodeling during asthma progression. Stearoyl-CoA desaturase 1 (SCD1) plays an important role in the response of fibroblasts to growth factors. This study aimed to explore the effects of SCD1 on fibroblast activation induced by transforming growth factor-β1 (TGF-β1) and the role of the phosphatidylinositol-3-kinase-AKT serine-threonine protein kinase-mechanistic target of rapamycin (PI3K-Akt-mTOR) pathway on the regulation of SCD1 expression in airway remodeling. Female C57BL/6 mice were sensitized and challenged with house dust mites to generate a chronic asthma model. The inhibitor of SCD1 was injected i.g. before each challenge. The airway hyper-responsiveness to methacholine was evaluated, and airway remodeling and airway inflammation were assessed by histology. The effects of SCD1 on fibroblast activation were evaluated in vitro using an SCD1 inhibitor and oleic acid and via the knockdown of SCD1. The involvement of the PI3K-Akt-mTOR-sterol regulatory element-binding protein 1 (SREBP1) pathway in lung fibroblasts was investigated using relevant inhibitors. The expression of SCD1 was increased in fibroblasts exposed to TGF-β1. The inhibition of SCD1 markedly ameliorated airway remodeling and lung fibroblast activation in peripheral airways. The knockdown or inhibition of SCD1 resulted in significantly reduced extracellular matrix production in TGF-β1-treated fibroblasts, but this effect was reversed by the addition of exogenous oleic acid. The PI3K-Akt-mTOR-SREBP1 pathway was found to be involved in the regulation of SCD1 expression and lung fibroblast activation. The data obtained in this study indicate that SCD1 expression contributes to fibroblast activation and airway remodeling and that the inhibition of SCD1 may be a therapeutic strategy for airway remodeling in asthma.

Background: There are no large sample, epidemiological data describing initial asthma severity and change. We used a large health care database to examine asthma severity at initial diagnosis, and the changes in severity over the first year of management. Methods: The clinical data of patients diagnosed with asthma for the first time were collated from the SuValue electronic medical database. The following inclusion criteria were applied: (I) patients who were 14 years or older at the time of first diagnosis; (II) initial diagnosis occurred between Jan 2001 and Mar 2019; (III) patients were followed up for at least 12 months; (IV) patients had follow-up visits every 3 months. Disease severity at diagnosis and at each follow-up visit, medications prescribed were collated and analyzed. Results: A total of 7,654 adult patients with newly diagnosed asthma from tertiary hospitals (26.38%) and secondary hospitals (73.62%), who were followed up for at least 12 months, were included in this retrospective analysis. Approximately 54% of patients were females and the largest age group was over 60 years old (37.66%). Nearly 16% of patients were moderate to severe asthma initially. The proportions of patients with moderate and severe asthma decreased during the first 6 months, and remained stable thereafter. At the end of the 1-year follow-up period, 2.7% of patients had severe asthma. Patients with mild asthma tended to continue to have mild asthma in the following 3 months (>76.19%). However, of the patients with mild or moderate asthma at 3 months, 92.85% and 75.1%, respectively, experienced maintenance and reduction in severity and had mild asthma by 12 months. 1.26% and 3.15% of patients with mild or moderate asthma, respectively, progressed to severe asthma by 12 months. Conclusions: Patients with mild asthma did not progress but rather, remained stable with mild asthma over the year. A proportion of patients diagnosed with moderate and severe asthma remained stable over a 1-year period. Further studies should be conducted to examine the clinical features of newly diagnosed patients with severe asthma without reduction in severity in order to facilitate intensive treatment and reduce the disease burden for these patients.