Cigarette smoke-induced dysbiosis: comparative analysis of lung and intestinal microbiomes in COPD mice and patients.

BACKGROUND: The impact of cigarette smoke (CS) on lung diseases and the role of microbiome dysbiosis in chronic obstructive pulmonary disease (COPD) have been previously reported; however, the relationships remain unclear. METHODS: Our research examined the effects of 20-week cigarette smoke (CS) exposure on the lung and intestinal microbiomes in C57BL/6JNarl mice, alongside a comparison with COPD patients' intestinal microbiome data from a public dataset. RESULTS: The study found that CS exposure significantly decreased forced vital capacity (FVC), thickened airway walls, and induced emphysema. Increased lung damage was observed along with higher lung keratinocyte chemoattractant (KC) levels by CS exposure. Lung microbiome analysis revealed a rise in Actinobacteriota, while intestinal microbiome showed significant diversity changes, indicating dysbiosis. Principal coordinate analysis highlighted distinct intestinal microbiome compositions between control and CS-exposed groups. In the intestinal microbiome, notable decreases in Patescibacteria, Campilobacterota, Defferibacterota, Actinobacteriota, and Desulfobacterota were observed. We also identified correlations between lung function and dysbiosis in both lung and intestinal microbiomes. Lung interleukins, interferon-É£, KC, and 8-isoprostane levels were linked to lung microbiome dysbiosis. Notably, dysbiosis patterns in CS-exposed mice were similar to those in COPD patients, particularly of Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage 4 patients. This suggests a systemic impact of CS exposure. CONCLUSION: In summary, CS exposure induces significant dysbiosis in lung and intestinal microbiomes, correlating with lung function decline and injury. These results align with changes in COPD patients, underscoring the important role of microbiome in smoke-related lung diseases.

Prediction value of neutrophil and eosinophil count at risk of COPD exacerbation.

INTRODUCTION: Predicting acute exacerbations (AEs) in chronic obstructive pulmonary disease (COPD) is crucial. This study aimed to identify blood biomarkers for predicting COPD exacerbations by inflammatory phenotypes. MATERIALS AND METHODS: We analyzed blood cell counts and clinical outcomes in 340 COPD patients aged 20-90 years. Patients were categorized into eosinophilic inflammation (EOCOPD) and non-eosinophilic inflammation (N-EOCOPD) groups. Blood cell counts, eosinophil-to-lymphocyte ratio (ELR), neutrophil-to-lymphocyte ratio (NLR) and neutrophil-to-eosinophil ratio (NER) were calculated. Linear and logistic regression models assessed relationships between health outcomes and blood cell counts. RESULTS: EOCOPD patients had distinct characteristics compared to N-EOCOPD patients. Increased neutrophil % and decreased lymphocyte % were associated with reduced pulmonary function, worse quality of life and more exacerbations, but they did not show statistical significance after adjusting by age, sex, BMI, smoking status, FEV1% and patient's medication. Subgroup analysis revealed a 1.372-fold increase in the OR of AE for every 1 unit increase in NLR in EOCOPD patients (p < .05). In N-EOCOPD patients, every 1% increase in blood eosinophil decreased the risk of exacerbation by 59.6%. CONCLUSIONS: Our study indicates that distinct white blood cell profiles in COPD patients, with or without eosinophilic inflammation, can help assess the risk of AE in clinical settings.

Role of DCLK1/Hippo pathway in type II alveolar epithelial cells differentiation in acute respiratory distress syndrome.

BACKGROUND: Delay in type II alveolar epithelial cell (AECII) regeneration has been linked to higher mortality in patients with acute respiratory distress syndrome (ARDS). However, the interaction between Doublecortin-like kinase 1 (DCLK1) and the Hippo signaling pathway in ARDS-associated AECII differentiation remains unclear. Therefore, the objective of this study was to understand the role of the DCLK1/Hippo pathway in mediating AECII differentiation in ARDS. MATERIALS AND METHODS: AECII MLE-12 cells were exposed to 0, 0.1, or 1 µg/mL of lipopolysaccharide (LPS) for 6 and 12 h. In the mouse model, C57BL/6JNarl mice were intratracheally (i.t.) injected with 0 (control) or 5 mg/kg LPS and were euthanized for lung collection on days 3 and 7. RESULTS: We found that LPS induced AECII markers of differentiation by reducing surfactant protein C (SPC) and p53 while increasing T1α (podoplanin) and E-cadherin at 12 h. Concurrently, nuclear YAP dynamic regulation and increased TAZ levels were observed in LPS-exposed AECII within 12 h. Inhibition of YAP consistently decreased cell levels of SPC, claudin 4 (CLDN-4), galectin 3 (LGALS-3), and p53 while increasing transepithelial electrical resistance (TEER) at 6 h. Furthermore, DCLK1 expression was reduced in isolated human AECII of ARDS, consistent with the results in LPS-exposed AECII at 6 h and mouse SPC-positive (SPC+) cells after 3-day LPS exposure. We observed that downregulated DCLK1 increased p-YAP/YAP, while DCLK1 overexpression slightly reduced p-YAP/YAP, indicating an association between DCLK1 and Hippo-YAP pathway. CONCLUSIONS: We conclude that DCLK1-mediated Hippo signaling components of YAP/TAZ regulated markers of AECII-to-AECI differentiation in an LPS-induced ARDS model.

Short-term mediating effects of PM2.5 on climate-associated COPD severity.

The impact of short-term exposure to environmental factors such as temperature, relative humidity (RH), and fine particulate matter (PM2.5) on chronic obstructive pulmonary disease (COPD) remains unclear. The objective of this study is to investigate PM2.5 as a mediator in the relationship between short-term variations in RH and temperature and COPD severity. A cross-sectional study was conducted on 930 COPD patients in Taiwan from 2017 to 2022. Lung function, COPD Assessment Test (CAT) score, and modified Medical Research Council (mMRC) dyspnea scale were assessed. The mean and differences in 1-day, 7-day, and 30-day individual-level exposure to ambient RH, temperature, and PM2.5 were estimated. The associations between these factors and clinical outcomes were analyzed using linear regression models and generalized additive mixed models, adjusting for age, sex, smoking, and body mass index. In the total season, increases in RH difference were associated with increases in forced expiratory volume in 1 s (FEV1) / forced vital capacity (FVC), while increases in temperature difference were associated with decreases in FEV1 and FEV1/FVC. Increases in PM2.5 mean were associated with declines in FEV1. In the cold season, increases in temperature mean were associated with decreases in CAT and mMRC scores, while increases in PM2.5 mean were associated with declines in FEV1, FVC, and FEV1/FVC. In the warm season, increases in temperature difference were associated with decreases in FEV1 and FEV1/FVC, while increases in RH difference and PM2.5 mean were associated with decreases in CAT score. PM2.5 fully mediated the associations of temperature mean with FEV1/FVC in the cold season. In conclusion, PM2.5 mediates the effects of temperature and RH on clinical outcomes. Monitoring patients during low RH, extreme temperature, and high PM2.5 levels is crucial. Capsule of findings The significance of this study is that an increase in ambient RH and temperature, as well as PM2.5 exposure, were significantly associated with changes in lung function, and clinical symptoms in these patients. The novelty of this study is that PM2.5 plays a mediating role in the association of RH and temperature with COPD clinical outcomes in the short term.

Sustaining the Activation of EGFR Signal by Inflammatory Cytokine IL17A Prompts Cell Proliferation and EGFR-TKI Resistance in Lung Cancer.

Non-small-cell lung cancer (NSCLC) is a typical inflammation-associated cancer, and lung adenocarcinoma (LUAD) is the most common pathological subtype. Epidermal growth factor (EGF) receptor (EGFR) mutations are the most common driver mutations of LUAD, and they have been identified as important therapeutic targets by EGFR-tyrosine kinase inhibitors (TKIs). The proinflammatory cytokine, interleukin (IL)-17A, and IL-17A-producing cells were reported to be elevated in the tumor microenvironment and peripheral blood of NSCLC patients and to be correlated with tumor progression and poor prognoses. However, the pathophysiological role of IL-17A in NSCLC remains unclear, although some studies suggested its involvement in cancer cell invasion and metastasis. Herein, we observed that expressions of IL-17A and its receptor, IL-17 receptor C (IL-17RC), were elevated in LUAD tissues and were correlated with poor survival in different lung cancer cohorts. In LUAD cells with mutant EGFR, the IL-17A/IL-17RC axis was shown to enhance phosphorylation of EGFR and Met, thereby promoting proliferation and resistance to EGFR-TKIs such as afatinib. In LUAD cells with wild-type (WT) EGFR, we found that the IL-17A/IL-17RC axis enhanced EGF-induced EGFR activation and cell proliferation through causing impairment of EGF-induced EGFR lysosomal degradation. Collectively, our results indicated diverse impacts of the IL-17A/IL-17RC axis on EGFR activation in LUAD cells with WT and mutant EGFR and suggested that developing therapeutic strategies against IL-17A/IL-17RC would be valuable for LUAD treatment.

Norisoboldine exerts antiallergic effects on IgE/ovalbumin-induced allergic asthma and attenuates FcεRI-mediated mast cell activation.

Allergic asthma is an inflammatory lung disorder, and mast cells play crucial roles in the development of this allergic disease. Norisoboldine (NOR), the major isoquinoline alkaloid present in Radix Linderae, has received considerable attention because it has anti-inflammatory effects. Herein, the aim of this study was to explore the antiallergic effects of NOR on allergic asthma in mice and mast cell activation. In a murine model of ovalbumin (OVA)-induced allergic asthma, oral administration at 5 mg/kg body weight (BW) of NOR produced strong reductions in serum OVA-specific immunoglobulin E (IgE) levels, airway hyperresponsiveness, and bronchoalveolar lavage fluid (BALF) eosinophilia, while an increase in CD4+Foxp3+ T cells of the spleen was detected. Histological studies demonstrated that NOR treatment significantly ameliorated the progression of airway inflammation including the recruitment of inflammatory cells and mucus production by decreasing levels of histamine, prostaglandin D2 (PGD2), interleukin (IL)-4, IL-5, IL-6, and IL-13 in BALF. Furthermore, our results revealed that NOR (3 âˆ¼ 30 µM) dose-dependently reduced expression of the high-affinity receptor for IgE (FcεRI) and the production of PGD2 and inflammatory cytokines (IL-4, IL-6, IL-13, and TNF-α), and also decreased degranulation of bone marrow-derived mast cells (BMMCs) activated by IgE/OVA. In addition, a similar suppressive effect on BMMC activation was observed by inhibition of the FcεRI-mediated c-Jun N-terminal kinase (JNK) signaling pathway using SP600125, a selective JNK inhibitor. Collectively, these results suggest that NOR may have therapeutic potential for allergic asthma at least in part through regulating the degranulation and the release of mediators by mast cells.

Importance of surface charge of soot nanoparticles in determining inhalation toxicity in mice.

Physicochemical properties of nanoparticles are important in regulating nanoparticle toxicity; however, the contribution of nanoparticle charge remains unclear. The objective of this study was to investigate the pulmonary effects of inhalation of charged soot nanoparticles. We established a stably charged nanoparticle generation system for whole-body exposure in BALB/c mice, which produced positively charged, negatively charged, and neutral soot nanoparticles in a wide range of concentrations. After a 7-day exposure, pulmonary toxicity was assessed, together with proteomics analysis. The charged soot nanoparticles on average carried 1.17-1.35 electric charges, and the sizes for nanoparticles under different charging conditions were all fixed at 69 ~ 72 nm. We observed that charged soot nanoparticles induced cytotoxic LDH and increased lung permeability, with the release of 8-isoprostane and caspase-3 and systemic IL-6 in mice, especially for positively charged soot nanoparticles. Next, we observed that positive-charged soot nanoparticles upregulated Eif2, Eif4, sirtuin, mammalian target of rapamycin (mTOR), peroxisome proliferator-activated receptors (PPAR), and HIPPO-related signaling pathways in the lungs compared with negatively charged soot nanoparticles. HIF1α, sirt1, E-cadherin, and Yap were increased in mice's lungs by positively charged soot nanoparticle exposure. In conclusion, carbonaceous nanoparticles carrying electric ions, especially positive-charged, are particularly toxic when inhaled and should be of concern in terms of pulmonary health protection.

Reduction of Emphysema Severity by Human Umbilical Cord-Derived Mesenchymal Stem Cells in Mice.

Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality in chronic lung disease patients throughout the world. Mesenchymal stem cells (MSCs) have been shown to regulate immunomodulatory, anti-inflammatory, and regenerative responses. However, the effects of human-umbilical-cord-derived mesenchymal stem cells (hUC-MSCs) on the lung pathophysiology of COPD remain unclear. We aimed to investigate the role of hUC-MSCs in emphysema severity and Yes-associated protein (Yap) phosphorylation (p-Yap) in a porcine-pancreatic-elastase (PPE)-induced emphysema model. We observed that the emphysema percentages (normalized to the total lung volume) measured by chest computed tomography (CT) and exercise oxygen desaturation were significantly reduced by hUC-MSCs at 107 cells/kg body weight (BW) via intravenous administration in emphysematous mice (p < 0.05). Consistently, the emphysema index, as assessed by the mean linear intercept (MLI), significantly decreased with hUC-MSC administration at 3 × 106 and 107 cells/kg BW (p < 0.05). Changes in the lymphocytes, monocytes, and splenic cluster of differentiation 4-positive (CD4+) lymphocytes by PPE were significantly reversed by hUC-MSC administration in emphysematous mice (p < 0.05). An increasing neutrophil/lymphocyte ratio was reduced by hUC-MSCs at 3 × 106 and 107 cells/kg BW (p < 0.05). The higher levels of tumor necrosis factor (TNF)-α, keratinocyte chemoattractant (KC), and lactate dehydrogenase (LDH) in bronchoalveolar lavage fluid (BALF) were significantly decreased by hUC-MSC administration (p < 0.05). A decreasing p-Yap/Yap ratio in type II alveolar epithelial cells (AECII) of mice with PPE-induced emphysema was significantly increased by hUC-MSCs (p < 0.05). In conclusion, the administration of hUC-MSCs improved multiple pathophysiological features of mice with PPE-induced emphysema. The effectiveness of the treatment of pulmonary emphysema with hUC-MSCs provides an essential and significant foundation for future clinical studies of MSCs in COPD patients.

Synergistic Tumor Inhibition via Energy Elimination by Repurposing Penfluridol and 2-Deoxy-D-Glucose in Lung Cancer.

Energy metabolism is the basis for cell growth, and cancer cells in particular, are more energy-dependent cells because of rapid cell proliferation. Previously, we found that penfluridol, an antipsychotic drug, has the ability to trigger cell growth inhibition of lung cancer cells via inducing ATP energy deprivation. The toxic effect of penfluridol is related to energy metabolism, but the underlying mechanisms remain unclear. Herein, we discovered that treatment of A549 and HCC827 lung cancer cells with penfluridol caused a decrease in the total amount of ATP, especially in A549 cells. An Agilent Seahorse ATP real-time rate assay revealed that ATP production rates from mitochondrial respiration and glycolysis were, respectively, decreased and increased after penfluridol treatment. Moreover, the amount and membrane integrity of mitochondria decreased, but glycolysis-related proteins increased after penfluridol treatment. Furthermore, we observed that suppression of glycolysis by reducing glucose supplementation or using 2-deoxy-D-glucose (2DG) synergistically enhanced the inhibitory effect of penfluridol on cancer cell growth and the total amount of mitochondria. A mechanistic study showed that the penfluridol-mediated energy reduction was due to inhibition of critical regulators of mitochondrial biogenesis, the sirtuin 1 (SIRT1)/peroxisome-proliferator-activated receptor co-activator-1α (PGC-1α) axis. Upregulation of the SIRT1/PGC-1α axis reversed the inhibitory effect of penfluridol on mitochondrial biogenesis and cell viability. Clinical lung cancer samples revealed a positive correlation between PGC-1α (PPARGC1A) and SIRT1 expression. In an orthotopic lung cancer mouse model, the anticancer activities of penfluridol, including growth and metastasis inhibition, were also enhanced by combined treatment with 2DG. Our study results strongly support that a combination of repurposing penfluridol and a glycolysis inhibitor would be a good strategy for enhancing the anticancer activities of penfluridol in lung cancer.

How platinum-induced nephrotoxicity occurs? Machine learning prediction in non-small cell lung cancer patients.

BACKGROUND AND OBJECTIVE: Platinum-induced nephrotoxicity is a severe and unexpected adverse drug reaction that could lead to treatment failure in non-small cell lung cancer patients. Better prediction and management of this nephrotoxicity can increase patient survival. Our study aimed to build up and compare the best machine learning models with clinical and genomic features to predict platinum-induced nephrotoxicity in non-small cell lung cancer patients. METHODS: Clinical and genomic data of patients undergoing platinum chemotherapy at Wan Fang Hospital were collected after they were recruited. Twelve models were established by artificial neural network, logistic regression, random forest, and support vector machine with integrated, clinical, and genomic modes. Grid search and genetic algorithm were applied to construct the fine-tuned model with the best combination of predictive hyperparameters and features. Accuracy, precision, recall, F1 score, and area under the receiver operating characteristic curve were calculated to compare the performance of the 12 models. RESULTS: In total, 118 patients were recruited for this study, among which 28 (23.73%) were experiencing nephrotoxicity. Machine learning models with clinical and genomic features achieved better prediction performances than clinical or genomic features alone. Artificial neural network with clinical and genomic features demonstrated the best predictive outcomes among all 12 models. The average accuracy, precision, recall, F1 score and area under the receiver operating characteristic curve of the artificial neural network with integrated mode were 0.923, 0.950, 0.713, 0.808 and 0.900, respectively. CONCLUSIONS: Machine learning models with clinical and genomic features can be a preliminary tool for oncologists to predict platinum-induced nephrotoxicity and provide preventive strategies in advance.

Comparative survival analysis of platinum-based adjuvant chemotherapy for early-stage squamous cell carcinoma and adenocarcinoma of the lung.

BACKGROUND AND PURPOSE: Although cytotoxic platinum-based adjuvant chemotherapy (pACT) has been recommended for patients with completely resected early-stage (ES) non-small-cell lung cancer (ES-NSCLC), therapeutic regimens for NSCLC have evolved in the past two decades. The study was aimed to examine the effectiveness of postoperative pACT for resected ES-NSCLC patients with squamous cell carcinoma (SCC) or adenocarcinoma (ADC) according to real-world data. METHODS AND PATIENTS: Inverse probability treatment weighting (IPTW) was used to adjust baseline characteristics between the group receiving pACT and those not receiving any treatment (observation, OBS) within 3 months after curative surgery. Cox regression models were used to compare overall survival (OS) and treatment failure-free survival (TFS) between the groups. RESULTS: Of 31,208 patients with ES-NSCLC, 4700 undergoing complete tumor resection were eligible, with a mean follow-up period of 4.5 years. The pACT (n = 2347) and OBS (n = 2353) groups were well-balanced after IPTW. The median OS differed between the pACT and OBS groups (77.2 vs. 75.5 months, adjusted hazard ratio [aHR] = 0.87, 95% confidence interval [CI] = 0.79-0.95, p = 0.003), and the 5-year survival rates were 58.2% and 55.3%, respectively (p < 0.001). In the SCC group, pACT was superior to OBS in OS (75.0 vs. 57.4 months, aHR = 0.74, 95% CI = 0.62-0.88, p = 0.001) and TFS (32.7 vs. 21.8 months, aHR = 0.74, 95% CI = 0.63-0.86, p < 0.001). Both OS and TFS did not differ between two groups in those with ADC. CONCLUSION: Real-world data indicated that pACT confers a survival benefit for resected ES-NSCLC patients with SCC but not ADC, which needs to be verified by a large sample of randomized controlled studies.

Acteoside exerts immunomodulatory effects on dendritic cells via aryl hydrocarbon receptor activation and ameliorates Th2-mediated allergic asthma by inducing Foxp3+ regulatory T cells.

Dendritic cells (DCs) are professional antigen-presenting cells that play a key role in directing T-cell responses and are involved in the pathogenesis of allergic asthma. Acteoside, an active phenylethanoid glycoside, is widely distributed in many medicinal plants. Herein, we explored the immunomodulatory effects of acteoside on bone marrow-derived DCs in vitro, and further investigated the immunosuppressive ability of acteoside to manipulate T helper type 2 (Th2)-mediated allergic asthma in mice. Following lipopolysaccharide activation, 50 µM of acteoside significantly reduced the production of proinflammatory mediators, including interleukin (IL)-12 and tumor necrosis factor (TNF)-α, whereas it enhanced secretion of the anti-inflammatory cytokine, IL-10, by DCs. However, these effects of acteoside on DCs were reversed by pretreatment with CH223191, an aryl hydrocarbon receptor (AhR) antagonist. Additionally, coculture of acteoside-treated DCs with CD4+ T cells promoted the generation of forkhead box P3-positive (Foxp3+) regulatory T cells (Tregs) via AhR activation. Using a murine asthma model, our results demonstrated that oral administration of 50 mg/kg of acteoside decreased levels of Th2-type cytokines, such as IL-4, IL-5, and IL-13, whereas the level of IL-10 and the frequency of CD4+Foxp3+ Tregs were augmented. Moreover, acteoside treatment markedly inhibited the elevated serum level of ovalbumin-specific immunoglobulin E, attenuated the development of airway hyperresponsiveness, and reduced inflammatory cell counts in bronchoalveolar lavage fluid. Additionally, histological results reveled that acteoside ameliorated pulmonary inflammation in asthmatic mice. Taken together, these results indicated that acteoside exhibits immunomodulatory effects on DCs and plays an anti-inflammatory role in the treatment of allergic asthma.

Schisandrin B promotes Foxp3+ regulatory T cell expansion by activating heme oxygenase-1 in dendritic cells and exhibits immunomodulatory effects in Th2-mediated allergic asthma.

Allergic asthma is induced by T helper 2 (Th2) responses and allergen-specific immunoglobulin E (IgE). In asthma, regulatory T (Treg) cells play a crucial role in controlling immune homeostasis, and induction of Treg cells is a good strategy to treat Th2-mediated allergic asthma. Schisandrin B (Sch B), the main component isolated from Schisandra chinensis, reportedly possesses various pharmacological properties, but its immunomodulatory mechanism in allergic asthma remains unclear. In the present study, we explored whether Sch B exerts an antiallergic effect through modifying the function of dendritic cells (DCs) to regulate T-cell polarization and further investigated the immunomodulatory effects of Sch B in allergic asthma. Herein, an in vitro study revealed that 20 µM of Sch B-treated bone-marrow-derived DCs exhibited a semi-mature phenotype that secreted low amounts of proinflammatory cytokines including interleukin (IL)-12, IL-1ß, IL-6, and tumor necrosis factor (TNF)-α, and expressed decreased levels of surface molecules of cluster of differentiation 80 (CD80) and CD86. Compared to fully mature DCs, these Sch B-treated DCs displayed a regulatory ability to promote CD4+Foxp3+ Treg cell generation via upregulation of heme oxygenase (HO)-1 expression. Of note, in a murine model of ovalbumin (OVA)-induced asthma, levels of Th2-type cytokines such as IL-4, IL-5, and IL-13, and C-C motif chemokine 11 (CCL11) were dampened, whereas numbers of forkhead box P3 (Foxp3)-positive Treg cells were augmented in Sch B-treated mice. Moreover, administration of 5 mg/kg of Sch B alleviated the cardinal features of Th2-mediated allergic asthma, namely, serum OVA-specific IgE production, the development of airway hyperresponsiveness (AHR), and airway inflammation. Collectively, these findings indicate that the effectiveness of Sch B treatment against Th2-mediated allergic asthma was at least partially due to enhancement of DC induction of Treg cells, and Sch B can possibly be developed as an immunomodulatory adjuvant to treat allergic asthma.

Blocking MMP-12-modulated epithelial-mesenchymal transition by repurposing penfluridol restrains lung adenocarcinoma metastasis via uPA/uPAR/TGF-ß/Akt pathway.

PURPOSE: Metastasis of lung adenocarcinoma (LADC) is a crucial factor determining patient survival. Repurposing of the antipsychotic agent penfluridol has been found to be effective in the inhibition of growth of various cancers. As yet, however, the anti-metastatic effect of penfluridol on LADC has rarely been investigated. Herein, we addressed the therapeutic potential of penfluridol on the invasion/metastasis of LADC cells harboring different epidermal growth factor receptor (EGFR) mutation statuses. METHODS: MTS viability, transwell migration and invasion, and tumor endothelium adhesion assays were employed to determine cytotoxic and anti-metastatic effects of penfluridol on LADC cells. Protease array, Western blot, immunohistochemistry (IHC), immunofluorescence (IF) staining, and expression knockdown by shRNA or exogenous overexpression by DNA plasmid transfection were performed to explore the underlying mechanisms, both in vitro and in vivo. RESULTS: We found that nontoxic concentrations of penfluridol reduced the migration, invasion and adhesion of LADC cells. Protease array screening identified matrix metalloproteinase-12 (MMP-12) as a potential target of penfluridol to modulate the motility and adhesion of LADC cells. In addition, we found that MMP-12 exhibited the most significantly adverse prognostic effect in LADC among 39 cancer types. Mechanistic investigations revealed that penfluridol inhibited the urokinase plasminogen activator (uPA)/uPA receptor/transforming growth factor-ß/Akt axis to downregulate MMP-12 expression and, subsequently, reverse MMP-12-induced epithelial-mesenchymal transition (EMT). Subsequent analysis of clinical LADC samples revealed a positive correlation between MMP12 and mesenchymal-related gene expression levels. A lower survival rate was found in LADC patients with a SNAl1high/MMP12high profile compared to those with a SNAl1low/MMP12low profile. CONCLUSIONS: Our results indicate that MMP-12 may serve as a useful biomarker for predicting LADC progression and as a promising penfluridol target for treating metastatic LADC.

Catalpol exerts antiallergic effects in IgE/ovalbumin-activated mast cells and a murine model of ovalbumin-induced allergic asthma.

Immunoglobulin E (IgE) and mast cells play important roles in the pathogenesis of allergic asthma. Catalpol, an iridoid glycoside, exerts many biological functions including anti-inflammatory activities. Herein, we investigated catalpol to determine both its antiallergic effects on IgE/ovalbumin (OVA)-stimulated mouse bone marrow-derived mast cells and its therapeutic actions in murine allergic asthma. We found that catalpol dramatically suppressed IgE/OVA-induced mast cell degranulation. Meanwhile, 5 ~ 100 µM of catalpol neither affected the expression level of the high-affinity receptor of IgE (FcεRI) by mast cells nor induced mast cell apoptosis. In addition, mRNA expression levels of inflammatory enzymes including cyclooxygenase (COX)-1, COX-2, and 5-lipoxygenase were downregulated. Administration of catalpol also suppressed production of prostaglandin D2 (PGD2), interleukin (IL)-6, and IL-13, while not affecting tumor necrosis factor (TNF)-α production. Further, catalpol pretreatment significantly attenuated the FcεRI-mediated Akt signaling pathway. In mice with IgE/OVA-induced asthma, oral administration of catalpol remarkably suppressed the production of OVA-specific IgE, the development of airway hyperresponsiveness (AHR), and the infiltration of eosinophils and neutrophils into the lungs. Histological studies demonstrated that catalpol substantially inhibited the recruitment of mast cells and increased mucus production in lung tissues. Catalpol-treated mice had significantly lower levels of helper T cell type 2 (Th2) cytokines (IL-4, IL-5, and IL-13), PGD2, eotaxin-1, and C-X-C chemokine ligand-1 (CXCL1) in bronchoalveolar lavage fluid (BALF) than did the allergic group. Collectively, these results indicated that the suppressive effects of catalpol on degranulation and mediator generation by mast cells were beneficial in treating allergic asthma.

Associations of TIMP-3 Genetic Polymorphisms with EGFR Statuses and Cancer Clinicopathologic Development in Lung Adenocarcinoma Patients.

Lung adenocarcinoma (LADC) is a major subtype of lung cancer, particularly among populations of East Asia. The epidermal growth factor receptor (EGFR) is the most frequently mutated oncogene promoting LADC progression and can serve as a therapeutic target in LADC. The tissue inhibitor of metalloproteinases (TIMP)-3 is a major regulator of extracellular matrix turnover via targeting of matrix metalloproteinases (MMPs), and thus, plays a critical role in tumor development and progression. The purpose of this study was to investigate potential associations among TIMP-3 genetic polymorphisms, EGFR statuses, and cancer clinicopathologic development in patients with LADC. In this study, 277 LADC patients with different EGFR statuses were recruited to dissect the allelic discrimination of TIMP-3 -1296 T>C (rs9619311), TIMP3 249T>C (rs9862), and TIMP3 261C>T (rs11547635) polymorphisms using a TaqMan allelic discrimination assay. Our data showed that compared to those LADC patients with wild-type CC homozygotes of TIMP-3 rs9862, patients harboring TT homozygotes of rs9862 were at a higher risk of developing mutant EGFR (adjusted odds ratio (AOR) = 2.530; 95% confidence interval (CI): 1.230-5.205; p = 0.012), particularly the EGFR L858R point mutation (AOR = 2.975; 95% CI: 1.182-7.488; p = 0.021). Moreover, we observed that TIMP-3 TT homozygotes of rs9862 were correlated with the incidence of EGFR mutations in patients with a smoking habit (p = 0.045). Within male patients harboring a mutant EGFR, TIMP-3 rs9862 T (CT+TT) allele carriers were at higher risk of developing an advanced stage (p = 0.025) and lymph node metastasis (p = 0.043). Further analyses of clinical datasets revealed correlations of TIMP-3 expression with a favorable prognosis in patients with LADC. In conclusion, the data suggest that TIMP-3 rs9862 polymorphisms may contribute to identify subgroups of lung cancer patients at high risk for tumor progression, among carriers of LADC-bearing mutant EGFR.

The Curcumin Analogue, EF-24, Triggers p38 MAPK-Mediated Apoptotic Cell Death via Inducing PP2A-Modulated ERK Deactivation in Human Acute Myeloid Leukemia Cells.

Curcumin (CUR) has a range of therapeutic benefits against cancers, but its poor solubility and low bioavailability limit its clinical use. Demethoxycurcumin (DMC) and diphenyl difluoroketone (EF-24) are natural and synthetic curcumin analogues, respectively, with better solubilities and higher anti-carcinogenic activities in various solid tumors than CUR. However, the efficacy of these analogues against non-solid tumors, particularly in acute myeloid leukemia (AML), has not been fully investigated. Herein, we observed that both DMC and EF-24 significantly decrease the proportion of viable AML cells including HL-60, U937, and MV4-11, harboring different NRAS and Fms-like tyrosine kinase 3 (FLT3) statuses, and that EF-24 has a lower half maximal inhibitory concentration (IC50) than DMC. We found that EF-24 treatment induces several features of apoptosis, including an increase in the sub-G1 population, phosphatidylserine (PS) externalization, and significant activation of extrinsic proapoptotic signaling such as caspase-8 and -3 activation. Mechanistically, p38 mitogen-activated protein kinase (MAPK) activation is critical for EF-24-triggered apoptosis via activating protein phosphatase 2A (PP2A) to attenuate extracellular-regulated protein kinase (ERK) activities in HL-60 AML cells. In the clinic, patients with AML expressing high level of PP2A have the most favorable prognoses compared to various solid tumors. Taken together, our results indicate that EF-24 is a potential therapeutic agent for treating AML, especially for cancer types that lose the function of the PP2A tumor suppressor.

Circulating Proteoglycan Endocan Mediates EGFR-Driven Progression of Non-Small Cell Lung Cancer.

Although new generations of EGFR-tyrosine kinase inhibitors (EGFR-TKI) have been developed for the treatment of patients with non-small cell lung cancer (NSCLC) with EGFR-mutant tumors, TKI resistance often returns as a result of additional EGFR mutations. In addition to seeking for next-generation EGFR-TKI, developing novel EGFR-targeting strategies may hold the key to overcome the vicious cycle of TKI resistance. Endocan is known as a receptor tyrosine kinase ligand enhancer in tumorigenesis, but the impact of endocan on EGFR-driven NSCLC progression remains unknown. In this study, higher endocan levels were found in lung tumors compared with cancer-free tissues and correlated with poor prognosis in patients with NSCLC harboring mutant EGFR; circulating endocan levels were also significantly higher in patients with mutant EGFR. Endocan facilitated EGFR signaling via direct binding and enhancing of the EGF-EGFR interaction and supported the growth of tumors driven by mutated EGFR. Activated EGFR in turn upregulated expression of endocan via JAK/STAT3 and ERK/ELK cascades, thus forming a positive regulatory loop of endocan-EGFR signaling. On the basis of the binding region between endocan and EGFR, we designed therapeutic peptides and demonstrated promising therapeutic effects in xenografts harboring EGFR mutations including TKI-resistant T790M. Together, our findings highlight the novel interaction between endocan and EGFR and new opportunities to effectively target endocan-EGFR regulatory axis in patients with TKI-resistant NSCLC. SIGNIFICANCE: Endocan is a novel and critical regulator of EGF/EGFR signaling and serves as an alternative target of EGFR-TKI resistance in NSCLC.

Genetic Variants of lncRNA MALAT1 Exert Diverse Impacts on the Risk and Clinicopathologic Characteristics of Patients with Hepatocellular Carcinoma.

The long noncoding (lnc)RNA, metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), plays a crucial role in the development of hepatocellular carcinoma (HCC). However, potential genetic variants (single nucleotide polymorphisms, SNPs) in MALAT1 that affect the susceptibility and progression of HCC have rarely been explored. Three tagging SNPs, viz., rs3200401 C > T, rs619586 A > G, and rs1194338 C > A, in MALAT1 were genotyped by a TaqMan allelic discrimination assay in 394 HCC patients and 1199 healthy controls. A stratified analysis showed that younger patients (<55 years) with the MALAT1 rs619586 G allele had a decreased risk of HCC under a codominant model (AOR = 0.289, 95% CI: 0.108-0.773, p = 0.013) and dominant model (AOR = 0.286, 95% CI: 0.107-0.765, p = 0.013). Female patients and patients with a smoking habit who carried the CA + AA genotype of rs1194338 had a lower risk of developing vascular invasion (p = 0.049) and a high Child-Pugh grade (B or C) (p = 0.036), respectively. Under the dominant model, smokers with the MALAT1 rs3200401 CT + TT genotype had a higher frequency of hepatitis B virus (HBV) infection (p = 0.034). Moreover, the aspartate aminotransferase was higher in patients with the rs3200401 CT + TT genotype. Furthermore, analyses of clinical datasets revealed that MALAT1 expression level was gradually unregulated during HCC development from normal liver, cirrhotic liver, dysplastic liver to HCC and correlated with poor survival rates in HCC patients, especially in the hepatitis virus-infected population.