DACH1 Attenuates Airway Inflammation in Chronic Obstructive Pulmonary Disease by Activating NRF2 Signaling.

Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease of the airways characterized by impaired lung function induced by cigarette smoke (CS). Reduced DACH1 (dachshund homolog 1) expression has a detrimental role in numerous disorders, but its role in COPD remains understudied. This study aimed to elucidate the role and underlying mechanism of DACH1 in airway inflammation in COPD by measuring DACH1 expression in lung tissues of patients with COPD. Airway epithelium-specific DACH1-knockdown mice and adenoassociated virus-transfected DACH1-overexpressing mice were used to investigate the role of DACH1 and the potential for therapeutic targeting in experimental COPD caused by CS. Furthermore, we discovered a potential mechanism of DACH1 in inflammation induced by CS extract stimulation in vitro. Compared with nonsmokers and smokers without COPD, patients with COPD had reduced DACH1 expression, especially in the airway epithelium. Airway epithelium-specific DACH1 knockdown aggravated airway inflammation and lung function decline caused by CS in mice, whereas DACH1 overexpression protected mice from airway inflammation and lung function decline. DACH1 knockdown and overexpression promoted and inhibited IL-6 and IL-8 secretion, respectively, in 16HBE human bronchial epidermal cells after CS extract stimulation. NRF2 (nuclear factor erythroid 2-related factor 2) was discovered to be a novel downstream target of DACH1, which binds directly to its promoter. By activating NRF2 signaling, DACH1 induction reduced inflammation. DACH1 levels are lower in smokers and nonsmoking patients with COPD than in nonsmokers. DACH1 has protective effects against inflammation induced by CS by activating the NRF2 signaling pathway. Targeting DACH1 is a potentially viable therapeutic approach for COPD treatment.

Blood glucose and lipids are associated with sarcoidosis: findings from observational and mendelian randomization studies.

BACKGROUND: Several researches have demonstrated that patients with sarcoidosis accompanied with the abnormality in blood glucose and/or lipids, however, the causal relationship between them remains uncertain. To elucidate the potential association and causality of blood glucose and lipids with sarcoidosis, we conducted a propensity score matching (PSM)-based observational study combined with mendelian randomization (MR) analysis. METHODS: All subjects in this study were retrospectively collected from Tongji Hospital during 2010 and 2023. 1:1 PSM was employed to control the potential confounders as appropriate. Univariable and multivariable logistic regression analyses were performed to estimate the associations of sarcoidosis with fasting glucose, high density lipoprotein cholesterol (HDLC), low density lipoprotein cholesterol (LDLC), total cholesterol (TC), and total triglyceride (TG). The further subtype analysis was also conducted. Afterwards, a bidirectional MR analysis based on public data deeply explored the causality among the 5 candidate traits and sarcoidosis, for which the inverse-variance weighted (IVW) method was utilized as the main inferring approach. RESULTS: In the observational study, a total number of 756 subjects were enrolled, with 162 sarcoidosis patients and 594 non-sarcoidosis participants, while 160 pairs of subjects were matched after PSM. Multivariable logistic regression analysis indicated that HDLC (OR: 0.151; 95% CI: 0.056-0.408; P < 0.001) and TC (OR: 3.942; 95% CI: 2.644-5.877; P < 0.001) were strongly associated with sarcoidosis. Subtype analysis showed that low HDLC was independently correlated to risk of lesions in bronchus and lungs, and mediastinal lymph nodes, while high TC was to cervical lymph nodes. In MR analysis, high fasting glucose, low HDLC, and high TC were identified as the causal factors of sarcoidosis. CONCLUSION: HDLC and TC had the potential to influence the risk of sarcoidosis, which could be regarded as predictors and may provide new diagnostic and therapeutic targets for sarcoidosis.

Human epididymis protein 4 aggravates airway inflammation and remodeling in chronic obstructive pulmonary disease.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a progressive disease characterized by chronic inflammation and airway remodeling. Human epididymis protein 4 (HE4) plays a critical role in various inflammatory or fibrotic diseases. However, the role of HE4 in COPD remains unidentified. METHODS: HE4 expression was determined in the lung tissues from COPD patients and cigarette smoke (CS)-exposed mice using immunohistochemical staining, qPCR, or western blot. The plasma level of HE4 was detected by ELISA. The regulations of HE4 in the expressions of CS extract (CSE)-induced inflammatory cytokines in human bronchial epithelial cells (HBE) were investigated through knockdown or overexpression of HE4. The role of secretory HE4 (sHE4) in the differentiation and proliferation in human pulmonary fibroblast cells (HPF) was explored via qPCR, western blot, CCK8 assay or 5-ethynyl-2'-deoxyuridine (EdU) staining. The probe of related mechanism in CSE-induced HE4 increase in HBE was conducted by administrating N-acetylcysteine (NAC). RESULTS: HE4 was up-regulated in both the lung tissue and plasma of COPD patients relative to controls, and the plasma HE4 was negatively associated with lung function in COPD patients. The same enhanced HE4 expression was verified in CS-exposed mice and CSE-induced HBE, but CSE failed to increase HE4 expression in HPF. In vitro experiments showed that reducing HE4 expression in HBE alleviated CSE-induced IL-6 release while overexpressing HE4 facilitated IL-6 expression, mechanistically through affecting phosphorylation of NFκB-p65, whereas intervening HE4 expression had no distinctive influence on IL-8 secretion. Furthermore, we confirmed that sHE4 promoted fibroblast-myofibroblast transition, as indicated by promoting the expression of fibronectin, collagen I and α-SMA via phosphorylation of Smad2. EdU staining and CCK-8 assay demonstrated the pro-proliferative role of sHE4 in HPF, which was further confirmed by enhanced expression of survivin and PCNA. Pretreatment of NAC in CSE or H2O2-induced HBE mitigated HE4 expression. CONCLUSIONS: Our study indicates that HE4 may participate in airway inflammation and remodeling of COPD. Cigarette smoke enhances HE4 expression and secretion in bronchial epithelium mediated by oxidative stress. Increased HE4 promotes IL-6 release in HBE via phosphorylation of NFκB-p65, and sHE4 promotes fibroblastic differentiation and proliferation.

DNA hypomethylation-mediated upregulation of GADD45B facilitates airway inflammation and epithelial cell senescence in COPD.

INTRODUCTION: Chronic obstructive pulmonary disease (COPD) is a heterogeneous disease typically characterized by chronic airway inflammation, with emerging evidence highlighting the driving role of cellular senescence-related lung aging. Accelerated lung aging and inflammation mutually reinforce each other, creating a detrimental cycle that contributes to disease progression. Growth arrest and DNA damage-inducible (GADD45) family has been reported to involve in multiple biological processes, including inflammation and senescence. However, the role of GADD45 family in COPD remains elusive. OBJECTIVES: To investigate the role and mechanism of GADD45 family in COPD pathogenesis. METHODS: Expressions of GADD45 family were evaluated by bioinformatic analysis combined with detections in clinical specimens. The effects of GADD45B on inflammation and senescence were investigated via constructing cell model with siRNA transfection or overexpression lentivirus infection and animal model with Gadd45b knockout. Targeted bisulfite sequencing was performed to probe the influence of DNA methylation in GADD45B expression in COPD. RESULTS: GADD45B expression was significantly increased in COPD patients and strongly associated with lung function, whereas other family members presented no changes. GADD45B upregulation was confirmed in mice exposed by cigarette smoke (CS) and HBE cells treated by CS extract as well. Moreover, experiments involving bidirectional modulation of GADD45B expression in HBE cells further substantiated its positive regulatory role in inflammatory response and cellular senescence. Mechanically, GADD45B-facilitated inflammation was directly mediated by p38 phosphorylation, while GADD45B interacted with FOS to promote cellular senescence in a p38 phosphorylation-independent manner. Furthermore, Gadd45b deficiency remarkably alleviated inflammation and senescence of lungs in CS-exposed mice, as well as improved emphysema and lung function. Eventually, in vivo and vitro experiments demonstrated that GADD45B overexpression was partially mediated by CS-induced DNA hypomethylation. CONCLUSION: Our findings have shed light on the impact of GADD45B in the pathogenesis of COPD, thereby offering a promising target for intervention in clinical settings.

Metabolism-Related Gene TXNRD1 Regulates Inflammation and Oxidative Stress Induced by Cigarette Smoke through the Nrf2/HO-1 Pathway in the Small Airway Epithelium.

Chronic obstructive pulmonary disease (COPD), a small airway disease, is regarded as a metabolic disorder. To further uncover the metabolic profile of COPD patients, it is necessary to identify metabolism-related differential genes in small airway epithelium (SAE) of COPD. Metabolism-related differential genes in SAE between COPD patients and nonsmokers were screened from GSE128708 and GSE20257 datasets. KEGG, GO, and PPI analyses were performed to evaluate the pathway enrichment, term enrichment, and protein interaction of candidate metabolism-related differential genes, respectively. RT-PCR was used to verify the mRNA expression of the top ten differential genes. Western blotting was used to evaluate the protein expression of TXNRD1. TXNRD1 inhibitor auranofin (AUR) was used to assess the impact of TXNRD1 on oxidative stress and inflammation induced by cigarette smoke extraction (CSE). Twenty-four metabolism-related differential genes were selected. ALDH3A1, AKR1C3, CYP1A1, AKC1C1, CPY1B1, and TXNRD1 in the top ten genes were significantly upregulated after CSE simulation for 24 h in human bronchial epithelial (16HBE) cells. Among them, CYP1A1 and TXNRD1 also have a significant upregulation in primary SAE after simulation of CSE for 24 h. The overexpression of protein TXNRD1 has also been detected in 16HBE cells, primary SAE stimulated with CSE, and mouse lung exposed to cigarette smoke (CS). Additionally, inhibition of TXNRD1 with 0.1 µM AUR alleviated the expression of IL-6 and reactive oxygen species (ROS) induced by CSE by activating the Nrf2/HO-1 pathway in 16HBE cells. This study identified twenty-four metabolism-related differential genes associated with COPD. TXNRD1 might participate in the oxidative stress and inflammation induced by CS by regulating the activation of the Nrf2/HO-1 pathway.

Identification and validation of genetic signature associated with aging in chronic obstructive pulmonary disease.

Aging plays an essential role in the development for chronic obstructive pulmonary disease (COPD). The aim of this study was to identify and validate the potential aging-related genes of COPD through bioinformatics analysis and experimental validation. Firstly, we compared the gene expression profiles of aged and young COPD patients using two datasets (GSE76925 and GSE47460) from Gene Expression Omnibus (GEO), and identified 244 aging-related different expressed genes (DEGs), with 132 up-regulated and 112 down-regulated. Then, by analyzing the data for cigarette smoke-induced COPD mouse model (GSE125521), a total of 783 DEGs were identified between aged and young COPD mice, with 402 genes increased and 381 genes decreased. Additionally, functional enrichment analysis revealed that these DEGs were actively involved in COPD-related biological processes and function pathways. Meanwhile, six genes were identified as the core aging-related genes in COPD after combining the human DEGs and mouse DEGs. Eventually, five out of six core genes were validated to be up-regulated in the lung tissues collected from aged COPD patients than young COPD patients, namely NKG7, CKLF, LRP4, GDPD3 and CXCL9. Thereinto, the expressions of NKG7 and CKLF were negatively associated with lung function. These results may expand the understanding for aging in COPD.

Cigarette Smoke Promotes Interleukin-8 Production in Alveolar Macrophages Through the Reactive Oxygen Species/Stromal Interaction Molecule 1/Ca2+ Axis.

Chronic obstructive pulmonary disease (COPD), primarily attributed to cigarette smoke (CS), is characterized by multiple pathophysiological changes, including oxidative stress and inflammation. Stromal interaction molecule 1 (STIM1) is a Ca2+ sensor that regulates Ca2+ entry in different types of cells. The present study aimed to explore the relationship between CS-induced oxidative stress and inflammation, as well as the functional role of STIM1 thereinto. Our results showed that the reactive oxygen species (ROS)/STIM1/Ca2+ axis played a critical role in CS-induced secretion of interleukin (IL)-8 in human alveolar macrophages. Specifically, smokers with COPD (SC) showed higher levels of ROS in the lung tissues compared with healthy non-smokers (HN). STIM1 was upregulated in the lung tissues of COPD patients. The expression of STIM1 was positively associated with ROS levels and negatively correlated with pulmonary function. The expression of STIM1 was also increased in the bronchoalveolar lavage fluid (BALF) macrophages of COPD patients and PMA-differentiated THP-1 macrophages stimulated by cigarette smoke extract (CSE). Additionally, CSE-induced upregulation of STIM1 in PMA-differentiated THP-1 macrophages was inhibited by pretreatment with N-acetylcysteine (NAC), a ROS scavenger. Transfection with small interfering RNA (siRNA) targeting STIM1 and pretreatment with NAC alleviated CSE-induced increase in intracellular Ca2+ levels and IL-8 expression. Furthermore, pretreatment with SKF-96365 and 2-APB, the inhibitors of Ca2+ influx, suppressed CSE-induced secretion of IL-8. In conclusion, our study demonstrates that CSE-induced ROS production may increase the expression of STIM1 in macrophages, which further promotes the release of IL-8 by regulating Ca2+ entry. These data suggest that STIM1 may play a crucial role in CSE-induced ROS production and inflammation, and participate in the pathogenesis of COPD.

Glycogen synthase kinase-3ß inhibition promotes lysosome-dependent degradation of c-FLIPL in hepatocellular carcinoma.

Glycogen synthase kinase-3ß (GSK-3ß) is a ubiquitously expressed serine/threonine kinase involved in a variety of functions ranging from the control of glycogen metabolism to transcriptional regulation. We recently demonstrated that GSK-3ß inhibition triggered ASK1-JNK-dependent apoptosis in human hepatocellular carcinoma (HCC) cells. However, the comprehensive picture of downstream GSK-3ß-regulated pathways/functions remains elusive. In this study, we showed that GSK-3ß was aberrantly activated in HCC. Pharmacological inhibition and genetic depletion of GSK-3ß suppressed the growth and induced caspase-dependent apoptosis in HCC cells. In addition, GSK-3ß inhibition-induced apoptosis through downregulation of c-FLIPL in HCC, which was caused by biogenesis of functional lysosomes and subsequently c-FLIPL translocated to lysosome for degradation. This induction of the lysosome-dependent c-FLIPL degradation was associated with nuclear translocation of transcription factor EB (TFEB), a master regulator of lysosomal biogenesis. Moreover, GSK-3ß inhibition-induced TFEB translocation acts through activation of AMPK and subsequently suppression of mTOR activity. Thus our findings reveal a novel mechanism by which inhibition of GSK-3ß promotes lysosome-dependent degradation of c-FLIPL. Our study shows that GSK-3ß may become a promising therapeutic target for HCC.

Synthesis and biological evaluation of new berberine derivatives as cancer immunotherapy agents through targeting IDO1.

To discover small-molecule cancer immunotherapy candidates through targeting Indoleamine 2,3-dioxygenase 1 (IDO1), twenty-five new berberine (BBR) derivatives defined with substituents on position 3 or 9 were synthesized and examined for repression of IFN-γ-induced IDO1 promoter activities. Structure-activity relationship (SAR) indicated that large volume groups at the 9-position might be beneficial for potency. Among them, compounds 2f, 2i, 2n, 2o and 8b exhibited increased activities, with inhibition rate of 71-90% compared with BBR. Their effects on IDO1 expression were further confirmed by protein level as well. Furthermore, compounds 2i and 2n exhibited anticancer activity by enhancing the specific lysis of NK cells to A549 through IDO1, but not cytotoxicity. Preliminary mechanism revealed that both of them inhibited IFN-γ-induced IDO1 expression through activating AMPK and subsequent inhibition of STAT1 phosphorylation. Therefore, compounds 2i and 2n have been selected as IDO1 modulators for small-molecule cancer immunotherapy for next investigation.