NDRG1 promotes endothelial dysfunction and hypoxia-induced pulmonary hypertension by targeting TAF15.

Background: Pulmonary hypertension (PH) represents a threatening pathophysiologic state that can be induced by chronic hypoxia and is characterized by extensive vascular remodeling. However, the mechanism underlying hypoxia-induced vascular remodeling is not fully elucidated. Methods and Results: By using quantitative polymerase chain reactions, western blotting, and immunohistochemistry, we demonstrate that the expression of N-myc downstream regulated gene-1 (NDRG1) is markedly increased in hypoxia-stimulated endothelial cells in a time-dependent manner as well as in human and rat endothelium lesions. To determine the role of NDRG1 in endothelial dysfunction, we performed loss-of-function studies using NDRG1 short hairpin RNAs and NDRG1 over-expression plasmids. In vitro, silencing NDRG1 attenuated proliferation, migration, and tube formation of human pulmonary artery endothelial cells (HPAECs) under hypoxia, while NDRG1 over-expression promoted these behaviors of HPAECs. Mechanistically, NDRG1 can directly interact with TATA-box binding protein associated factor 15 (TAF15) and promote its nuclear localization. Knockdown of TAF15 abrogated the effect of NDRG1 on the proliferation, migration and tube formation capacity of HPAECs. Bioinformatics studies found that TAF15 was involved in regulating PI3K-Akt, p53, and hypoxia-inducible factor 1 (HIF-1) signaling pathways, which have been proved to be PH-related pathways. In addition, vascular remodeling and right ventricular hypertrophy induced by hypoxia were markedly alleviated in NDRG1 knock-down rats compared with their wild-type littermates. Conclusions: Taken together, our results indicate that hypoxia-induced upregulation of NDRG1 contributes to endothelial dysfunction through targeting TAF15, which ultimately contributes to the development of hypoxia-induced PH.

A reductive dehalogenative process for chemo- and stereoselective synthesis of 1,3-dienylsulfonyl fluorides.

A method for the mild and efficient synthesis of 1,3-dienylsulfonyl fluorides was developed via dehalogenation of α-halo-1,3-dienylsulfonyl fluorides in the presence of zinc powder and acetic acid, achieving exclusive chemo- and stereoselectivities. This protocol was successfully applied to the synthesis of heterocyclic dienylsulfonyl fluorides and polyene sulfonyl fluoride.

Evaluating carbon content in airway macrophages as a biomarker of personal exposure to fine particulate matter and its acute respiratory effects.

It remains unclear whether carbon content in airway macrophages (AM) can predict personal short-term exposure to fine particulate matter (PM2.5) air pollution and its respiratory health effects. We aimed to evaluate the pathway from personal PM2.5 exposure to adverse respiratory outcomes through AM carbon content. We designed a longitudinal panel study with 3 scheduled follow-ups among 113 non-smoking patients of chronic obstructive pulmonary disease in Shanghai, China, from April 2017 to January 2019. We quantified AM carbon content from induced sputum by image analysis, tested lung function and measured sputum levels of 4 pro-inflammatory cytokines and 2 anti-inflammatory cytokines. We applied the "meet in the middle" approach incorporating linear mixed-effect models to evaluate the associations from external PM2.5 exposure to respiratory outcomes through AM carbon content. Our results indicated that personal exposure to PM2.5 within 24 h was significantly associated with decreased forced expiratory volume in 1s and anti-inflammatory cytokines, as well as increased macrophages and pro-inflammatory cytokines. These changes were accompanied by increased areas of AM carbon and higher percentage of AM area occupied by carbon, both of which were associated with increased levels of pro-inflammatory cytokines and decreased levels of anti-inflammatory cytokines. Exposure to ambient black carbon and organic carbon in PM2.5 within 2 days was significantly associated with increased AM carbon area and percentage of AM area occupied by carbon. Our findings reinforced the causality in respiratory health effects of PM2.5 in which increased AM carbon content might serve as a valid exposure biomarker.

Lipopolysaccharide-Activated Bone Marrow-Derived Dendritic Cells Suppress Allergic Airway Inflammation by Ameliorating the Immune Microenvironment.

Background: Previous studies have shown that lipopolysaccharide (LPS)-activated bone marrow-derived dendritic cells (DClps) might induce tolerance in autoimmune and cancer models in vivo, whereas it remains unclear whether DClps could play a role in allergic disease model. Herein, we aimed to elucidate the potential effects of DClps on OVA-sensitized/challenged airway inflammation in a mouse model, which may help facilitate the application of specific tolerogenic dendritic cells (tolDC) in allergic asthma in the future. Methods: The phenotype and function of immature DC (DCia), DClps or IL-10-activated-DC (DC10) were determined. OVA-sensitized/challenged mice were treated with OVA-pulsed DCia or DClps or DC10. We assessed the changes of histopathology, serum total IgE level, pulmonary signal transducers and activators of transcription (STAT), pulmonary regulatory T cells (Tregs), and airway recall responses to OVA rechallenge, including proliferation and cytokine secretory function of pulmonary memory CD4+ T cells in the treated mice. Results: DClps exhibited low levels of CD80 and MHCII and increased levels of anti-inflammatory cytokines such as IL-10 and TGF-ß. Additionally, DClps treatment dramatically diminished infiltration of inflammatory cells, eosinophilia, serum IgE and STAT6 phosphorylation level, increased the number of pulmonary Tregs. In addition, DClps treatment decreased the proliferation of pulmonary memory CD4+ T cells, which further rendered the downregulation of Th2 cytokines in vitro. Conclusion: LPS stimulation may lead to a tolerogenic phenotype on DC, and thereby alleviated the Th2 immune response of asthmatic mice, possibly by secreting anti-inflammatory cytokines, inhibiting pulmonary memory CD4+ T cells, downregulating pulmonary STAT6 phosphorylation level and increasing pulmonary Tregs.

Induced sputum metabolomic profiles and oxidative stress are associated with chronic obstructive pulmonary disease (COPD) severity: potential use for predictive, preventive, and personalized medicine.

Chronic obstructive pulmonary disease (COPD) is a highly heterogeneous disease, and metabolomics plays a hub role in predictive, preventive, and personalized medicine (PPPM) related to COPD. This study thus aimed to reveal the role of induced sputum metabolomics in predicting COPD severity. In this pilot study, a total of 20 COPD patients were included. The induced sputum metabolites were assayed using a liquid chromatography-mass spectrometry (LC-MS/MS) system. Five oxidative stress products (myeloperoxidase (MPO), superoxide dismutase (SOD), glutathione (GSH), neutrophil elastase (NE), and 8-iso-PGF2α) in induced sputum were measured by ELISA, and the metabolomic profiles were distinguished by principal component analysis (PCA) and orthogonal projections to latent structures discriminant analysis (OPLS-DA). The Kyoto Encyclopedia of Genes and Genomes (KEGG) was used for pathway enrichment analysis, and a significant difference in induced sputum metabolomics was observed between moderate and severe COPD. The KEGG analysis revealed that the glycerophospholipid metabolism pathway was downregulated in severe COPD. Due to the critical role of glycerophospholipid metabolism in oxidative stress, significant negative correlations were discovered between glycerophospholipid metabolites and three oxidative stress products (SOD, MPO, and 8-iso-PGF2α). The diagnostic values of SOD, MPO, and 8-iso-PGF2α in induced sputum were found to exhibit high sensitivities and specificities in the prediction of COPD severity. Collectively, this study provides the first identification of the association between induced sputum metabolomic profiles and COPD severity, indicating the potential value of metabolomics in PPPM for COPD management. The study also reveals the correlation between glycerophospholipid metabolites and oxidative stress products and their value for predicting COPD severity. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13167-020-00227-w.

Adoptive transfer of bone marrow-derived dendritic cells (BMDCs) alleviates OVA-induced allergic airway inflammation in asthmatic mice.

Airway dendritic cells (DCs) are recognized as important factors in the mechanisms of allergic inflammatory diseases. Suppressor of cytokine signaling 3 (SOCS3) is involved in regulating the functions of T cells and macrophages, but the roles of SOCS3-expressing DCs in the pathogeneses of allergic inflammatory diseases are still controversial. We compared the effects of adoptively transferred SOCS3-/- and SOCS3+/+ bone marrow-derived DCs (BMDCs) on airway inflammation in ovalbumin (OVA)-sensitized asthmatic mice. Adoptive transfer of mature DCs (lipopolysaccharide [LPS]-induced DCs, DClps) with or without SOCS3 gene expression significantly ameliorated allergic airway inflammation. SOCS3-/- DCs slightly attenuated BMDC-induced immunogenic tolerance. DClps migrated to OVA-sensitized lungs with higher efficiency than immature DCs (DCim). DClps with or without SOCS3 greatly improved lung pathology scores and alleviated airway inflammatory cell infiltration after adoptive transfer into mice; they also increased interleukin-10 (IL-10) and transforming growth factor-ß (TGF-ß) production and inhibited signal transducer and activator of transcription (STAT) 4 and STAT6 signaling in the lungs after OVA sensitization. In conclusion, the BMDC adoptive transfer-induced immunogenic tolerance in OVA-sensitized mice might not be due to SOCS3 gene depletion. BMDC adoptive transfer may be developed into a new approach that alleviates asthma by modulating the balance between immune tolerance and inflammation.

PI3K inhibitor treatment ameliorates the glucocorticoid insensitivity of PBMCs in severe asthma.

BACKGROUND: Glucocorticoid (GC) insensitivity is an important feature of severe and fatal asthma. Oxidative stress can induce phosphoinositide-3-kinase (PI3K) activation, contributing to the development of GC insensitivity in chronic airway diseases. However, the underlying molecular mechanism of PI3K in the pathogenesis of severe asthma remains unknown. METHODS: We isolated peripheral blood mononuclear cells (PBMCs) from 34 participants (12 patients with mild/moderate asthma, 10 patients with severe asthma, and 12 control subjects). H2O2 was used to stimulate the human macrophage line U937 to mimic the oxidative stress status in severe asthma. The ability of candidate compounds, namely, azithromycin, PI3K inhibitors (BEZ235 and LY294002) and a p38 MAPK inhibitor (BIRB796), to ameliorate GC insensitivity in severe asthma was evaluated. RESULTS: PBMCs from patients with severe asthma exhibited dose-dependent and time-dependent GC insensitivity, which correlated with reduced activity of histone deacetylase 2 (HDAC2) (p < 0.05) and elevated expression of proinflammatory genes [nuclear factor-κB (NF-κB) and activator protein-1 (AP-1)] (p < 0.01) compared with these parameters in the control group. The PI3K inhibitors (BZE235 and LY294002) significantly restored the GC sensitivity of PBMCs from patients with severe asthma. In vitro, the PI3K inhibitors (BZE235 and LY294002) ameliorated GC insensitivity in H2O2/TNFα-induced IL-8 release from U937 cells by independently restoring the activity of HDAC2 or inhibiting the activation of transcription factors. CONCLUSIONS: This study demonstrates that PI3K inhibitors ameliorate GC insensitivity in severe asthma by restoring HDAC2 activity and inhibiting the phosphorylation of nuclear signaling transcription factors.

Clinical and pulmonary function changes in cough variant asthma with small airway disease.

BACKGROUND: It is known that small airway disease is present across all asthma severities; however, its prevalence and clinical characteristics in cough variant asthma (CVA) have not been fully illuminated. METHODS: A total of 77 CVA patients with preserved proximal airway function (FEV1/FVC > 70%) were enrolled in this study. The correlation between forced expiratory flow at 50% (FEF50%) and FEF25-75% in the CVA population was first evaluated. FEF50% was determined to be an easy and feasible parameter for identifying small airway disease. CVA with small airway disease is defined as FEF50% < 70%, whereas CVA with normal small airways is identified as FEF50% > 70%. Demographic features, clinical characteristics, lung function and induced sputum test results were determined at the initial visit and at the final visit 1 year later. RESULTS: FEF50% is a good marker for small airway disease. The cutoff value of 70% is more sensitive than the previously published 60% for identifying more patients with small airway problems early. Nearly half of the CVA population (45.4%) in our cohort had small airway disease. In both group, symptoms improved greatly after anti-asthmatic treatment. Interestingly, the changes in symptom scores [Asthma Control Test (ACT) and ACQ] were even greater in the CVA with small airway disease group than in the control group because of the higher medication usage in this subpopulation in real life. However anti-asthmatic therapy can not reverse small airway dysfunction. At last visit, FEF50% of CVA with small airway diseases was 57.2% ± 10.5%, still much lower than the control group (FEF50% = 92.6% ± 16.5%). CONCLUSIONS: In our cohort, nearly half of the CVA population had small airway disease. Their demographic features, clinical characteristics, airway eosinophils and drug responsiveness were quite similar between two groups, which means these indices can not be used as markers to identify small airway obstruction. We found FEF50% is an easy and feasible marker for early identification. Regular anti-asthmatic medication helped to improve clinical scores in patients with small airway disease, but the obstruction could not be reversed over 1-year period.

Preventative tracheal administration of interleukin-27 attenuates allergic asthma by improving the lung Th1 microenvironment.

BACKGROUND: Interleukin-27 (IL-27) modulates CD4+ T-cell differentiation and function. The aim of this study is to investigate the effect and molecular mechanisms of IL-27 on the development of asthma. METHODS: IL-27 was intranasally administered in an ovalbumin-induced asthma model, and lung mononuclear cells and different Th cell classes were detected by fluorescence-activated cell sorting. The effect and mechanisms of IL-27 on human bronchial epithelial (HBE) cells were investigated by measuring changes in chemotactic factors, cytokines, transcription factors, and signaling pathways. RESULTS: We found that intranasal administration of IL-27 could attenuate airway inflammation and hyperresponsiveness, upregulate the type 1 T helper (Th1)-T memory (Tm) cells and regulatory T (Treg) cells subgroups of lung tissue lymphocytes, and diminish the levels of type 2 T helper (Th2) cytokines. IL-27 upregulated the expression of C-C motif chemokine ligand 2 (CCL2), CCL3, and CCL4 in HBE cells and promoted the production of chemotactic factors to attract monocyte recruitment. Recruited monocytes secondarily secreted IL-27 to influence HBE cells in a positive feedback cycle. After IL-27 intervention, signal transducer and activator of transcription 1 (STAT1) phosphorylation increased, while STAT4 and STAT6 phosphorylation declined. CONCLUSIONS: Preventative intranasal administration of IL-27 can recruit more IL-27-secreted monocytes to the airway and change the different T-cell classes in lung. The improved Th1 environment helps to alleviate Th2-mediated allergic asthma by repairing the STAT1 pathway but not the STAT4 pathway.

Structural and functional characterisation of FOXO/Acan-DAF-16 from the parasitic nematode Angiostrongylus cantonensis.

Fork head box transcription factors subfamily O (FoxO) is regarded to be significant in cell-cycle control, cell differentiation, ageing, stress response, apoptosis, tumour formation and DNA damage repair. In the free-living nematode Caenorhabditis elegans, the FoxO transcription factor is encoded by Ce-daf-16, which is negatively regulated by insulin-like signaling (IIS) and involved in promoting dauer formation through bringing about its hundreds of downstream genes expression. In nematode parasites, orthologues of daf-16 from several species have been identified, with functions in rescue of dauer phenotypes determined in a surrogate system C. elegans. In this study, we identified the FoxO encoding gene, Acan-daf-16, from the parasitic nematode Angiostrongylus cantonensis, and determined the genomic structures, transcripts and functions far more thorough in longevity, stress resistance and dauer formation. Acan-daf-16 encodes two proteins, Acan-DAF-16A and Acan-DAF-16B, consisting of 555 and 491 amino acids, respectively. Both isoforms possess the highly conserved fork head domains. Acan-daf-16A and Acan-daf-16B are expressed from distinct promoters. The expression patterns of Acan-daf-16 isoforms in the C. elegans surrogate system showed that p Acan-daf-16a:gfp was expressed in all cells of C. elegans, including the pharynx, and the expression of p Acan-daf-16b:gfp was restricted to the pharynx. In addition to the same genomic organization to the orthologue in C. elegans, Ce-daf-16, both Acan-DAF-16 isoforms could restore the C. elegans daf-16(mg54) mutation in longevity, dauer formation and stress resistance, in spite of the partial complementation of Acan-DAF-16B isoform in longevity. These findings provide further evidence of the functional conservation of DAF-16s between parasitic nematodes and the free-living nematode C. elegans.