Pleural mesothelial cell migration into lung parenchyma by calpain contributes to idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is defined as a specific form of chronic, progressive fibrosing interstitial pneumonia. It is unknown why fibrosis in IPF distributes in the peripheral or named sub-pleural area. Migration of pleural mesothelial cells (PMC) should contribute to sub-pleural fibrosis. Calpain is known to be involved in cell migration, but the role of calpain in PMC migration has not been investigated. In this study, we found that PMCs migrated into lung parenchyma in patients with IPF. Then using Wt1tm1(EGFP/Cre)Wtp /J knock-in mice, we observed PMC migration into lung parenchyma in bleomycin-induced pleural fibrosis models, and calpain inhibitor attenuated pulmonary fibrosis with prevention of PMC migration. In vitro studies revealed that bleomycin and transforming growth factor-ß1 increased calpain activity in PMCs, and activated calpain-mediated focal adhesion (FA) turnover as well as cell migration, cell proliferation, and collagen-I synthesis. Furthermore, we determined that calpain cleaved FA kinase in both C-terminal and N-terminal regions, which mediated FA turnover. Lastly, the data revealed that activated calpain was also involved in phosphorylation of cofilin-1, and p-cofilin-1 induced PMC migration. Taken together, this study provides evidence that calpain mediates PMC migration into lung parenchyma to promote sub-pleural fibrosis in IPF.

VEGF/Src signaling mediated pleural barrier damage and increased permeability contributes to subpleural pulmonary fibrosis.

The distribution of fibrosis in idiopathic pulmonary fibrosis (IPF) is subpleural with basal predominance. Alveolar epithelial cell was considered as the key cell in the initial phase of IPF. However, the idea of activation and damage of alveolar epithelial cells is very difficult to explain why fibrosis distributes in the subpleural area. In this study, human pleural mesothelial cell (PMC) line and primary rat PMC was used as in vitro model. Intraperitoneal injection of bleomycin was used for making a pulmonary fibrosis model. The integrity of cultured monolayer PMCs was determined by transepithelial electric resistance (TEER). Pleural permeability was estimated by measuring paracellular transport of fluorescein isothiocyanate (FITC)-conjugated dextran. Changes in lung tissue of patients with IPF were analyzed by Masson's and immunofluorescence staining. We found bleomycin induced PMCs damage and increased PMCs permeability; increased PMCs permeability aggravated bleomycin-induced subpleural inflammation and pulmonary fibrosis. Moreover, bleomycin was found to activate VEGF/Src signaling which increased PMCs permeability. In vivo, inhibition of VEGF/Src signaling prevented bleomycin-induced subpleural pulmonary fibrosis. At last, activation of VEGF/Src signaling was confirmed in subpleural area in patients with IPF. Taken together, our findings indicate that VEGF/Src signaling mediated pleural barrier damage and increased permeability which contributes to subpleural pulmonary fibrosis.

Bleomycin induced apical-basal polarity loss in alveolar epithelial cell contributes to experimental pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrosing interstitial lung disease with limited therapeutic options and a median survival of 3 years after diagnosis. Dysregulated epithelial regeneration is key event involved in initiating and sustaining IPF. The type II alveolar epithelial cells (AECIIs) play a crucial role for epithelial regeneration and stabilisation of alveoli. Loss of cell apical-basal polarity contributes to fibrosis. AECII has apical-basal polarity, but it is poorly understood whether AECII apical-basal polarity loss is involved in fibrosis. Bleomycin is a traditional inducer of pulmonary fibrosis. Here firstly we observed that bleomycin induced apical-basal polarity loss in cultured AECIIs. Next, cell polarity proteins lethal (2) giant larvae 1 (Lgl1), PAR-3A, aPKC and PAR-6B were investigated. We found bleomycin induced increases of Lgl1 protein and decreases of PAR-3A protein, and bleomycin-induced PAR-3A depression was mediated by increased-Lgl1. Then Lgl1 siRNA was transfected into AECIIs. Lgl1 siRNA prevented apical-basal polarity loss in bleomycin-treated AECIIs. At last, Lgl1-conditional knockout mice were applied in making animal models. Bleomycin induced pulmonary fibrosis, but this was attenuated in Lgl1-conditional knockout mice. Together, these data indicated that bleomycin mediated AECII apical-basal polarity loss which contributed to experimental pulmonary fibrosis. Inhibition of Lgl1 should be a potential therapeutic strategy for the disease.

Crosstalk between pleural mesothelial cell and lung fibroblast contributes to pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a specific form of chronic, progressive and fibrosing interstitial pneumonia of unknown cause. The main feature of IPF is a heterogeneous appearance with areas of sub-pleural fibrosis. However, the mechanism of sub-pleural fibrosis was poorly understood. In this study, our in vivo study revealed that pleural mesothelial cells (PMCs) migrated into lung parenchyma and localized alongside lung fibroblasts in sub-pleural area in mouse pulmonary fibrosis. Our in vitro study displayed that cultured-PMCs-medium induced lung fibroblasts transforming into myofibroblast, cultured-fibroblasts-medium promoted mesothelial-mesenchymal transition of PMCs. Furthermore, these changes in lung fibroblasts and PMCs were prevented by blocking TGF-ß1/Smad2/3 signaling with SB431542. TGF-ß1 neutralized antibody attenuated bleomycin-induced pulmonary fibrosis. Similar to TGF-ß1/Smad2/3 signaling, wnt/ß-catenin signaling was also activated in the process of PMCs crosstalk with lung fibroblasts. Moreover, inhibition of CD147 attenuated cultured-PMCs-medium induced collagen-I synthesis in lung fibroblasts. Blocking CD147 signaling also prevented bleomycin-induced pulmonary fibrosis. Our data indicated that crosstalk between PMC and lung fibroblast contributed to sub-pleural pulmonary fibrosis. TGF-ß1, Wnt/ß-catenin and CD147 signaling was involved in the underling mechanism.

Cigarette smoking aggravates bleomycin-induced experimental pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease that typically leads to respiratory failure and death. The cause of IPF is poorly understood. Although several environmental and occupational factors are considered as risk factors in IPF, cigarette smoking seems to be the most strongly associated risk factor. Here firstly, we treated mice with cigarette (16 mg tar, 1.0 mg nicotine in each cigarette) smoking and tried to explore the role of cigarette smoking in pulmonary fibrosis. Mice were continuously subjected to smoke for about 1 h each day (12 cigarettes per day, 5 days per week) during 40 days. Bleomycin was administrated by intraperitoneal injection at a dose of 40 mg/kg on days 1, 5, 8, 11 and 15. We found bleomycin induced pulmonary fibrosis in mice, and cigarette smoking augmented bleomycin-induced fibrosis reflected by both in fibrotic area and percentages of collagen in the lungs. Then we prepared and employed cigarette smoke extract (CSE) in cell models and found that CSE could induce the activation of p-Smad2/3 and p-Akt, as well as collagen-I synthesis and cell proliferation in lung fibroblasts and pleural mesothelial cells (PMCs). TGF-ß1 signaling mediated CSE-induced PMCs migration. Moreover, in vitro studies revealed that CSE had superimposed effect on bleomycin-induced activation of TGF-ß-Smad2/3 and -Akt signaling. TGF-ß-Smad2/3 and -Akt signaling were further augmented by cigarette smoking in the lung of bleomycin-treated mice. Taken together, these findings represent the first evidence that cigarette smoking aggravated bleomycin-induced pulmonary fibrosis via TGF-ß1 signaling.

Lethal (2) giant larvae regulates pleural mesothelial cell polarity in pleural fibrosis.

Pleural fibrosis is barely reversible and the underlying mechanisms are poorly understood. Pleural mesothelial cells (PMCs) which have apical-basal polarity play a key role in pleural fibrosis. Loss of cell polarity is involved in the development of fibrotic diseases. Partition defective protein (PAR) complex is a key regulator of cell polarity. However, changes of PMC polarity and PAR complex in pleural fibrosis are still unknown. In this study, we observed that PMC polarity was lost in fibrotic pleura. Next we found increased Lethal (2) giant larvae (Lgl) bound with aPKC and PAR-6B competing against PAR-3A in PAR complex, which led to cell polarity loss. Then we demonstrated that Lgl1 siRNA prevented cell polarity loss in PMCs, and Lgl1 conditional knockout (ER-Cre+/-Lgl1flox/flox) attenuated pleural fibrosis in a mouse model. Our data indicated that Lgl1 regulates cell polarity of PMCs, inhibition of Lgl1 and maintenance of cell polarity in PMCs could be a potential therapeutic treatment approach for pleural fibrosis.

Inhibition of angiotensin II and calpain attenuates pleural fibrosis.

Pleural fibrosis is associated with various inflammatory processes such as tuberculous pleurisy and bacterial empyema. There is currently no ideal therapeutic to attenuate pleural fibrosis. Some pro-fibrogenic mediators induce fibrosis through inflammatory processes, suggesting that blockage of these mediators might prevent pleural fibrosis. The MeT-5A human pleural mesothelial cell line (PMC) was used in this study as an in vitro model of fibrosis; and intra-pleural injection of bleomycin with carbon particles was used as an in vivo mouse model of pleural fibrosis. Calpain knockout mice, calpain inhibitor (calpeptin), and angiotensin (Ang) II type 1 receptor (AT1R) antagonist (losartan) were evaluated in prevention of experimental pleural fibrosis. We found that bleomycin and carbon particles induced calpain activation in cultured PMCs. This in vitro response was associated with increased collagen-I synthesis, and was blocked by calpain inhibitor or AT1R antagonist. Calpain genetic or treatment with calpeptin or losartan prevented pleural fibrosis in a mouse model induced by bleomycin and carbon particles. Our findings indicate that Ang II signaling and calpain activation induce collagen-I synthesis and contribute to fibrotic alterations in pleural fibrosis. Inhibition of Ang II and calpain might therefore be a novel strategy in treatment of pleural fibrosis.

miR-18a-5p Inhibits Sub-pleural Pulmonary Fibrosis by Targeting TGF-ß Receptor II.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease that typically leads to respiratory failure and death within 3-5 years of diagnosis. Sub-pleural pulmonary fibrosis is a pathological hallmark of IPF. Bleomycin treatment of mice is a an established pulmonary fibrosis model. We recently showed that bleomycin-induced epithelial-mesenchymal transition (EMT) contributes to pleural mesothelial cell (PMC) migration and sub-pleural pulmonary fibrosis. MicroRNA (miRNA) expression has recently been implicated in the pathogenesis of IPF. However, changes in miRNA expression in PMCs and sub-pleural fibrosis have not been reported. Using cultured PMCs and a pulmonary fibrosis animal model, we found that miR-18a-5p was reduced in PMCs treated with bleomycin and that downregulation of miR-18a-5p contributed to EMT of PMCs. Furthermore, we determined that miR-18a-5p binds to the 3' UTR region of transforming growth factor ß receptor II (TGF-ßRII) mRNA, and this is associated with reduced TGF-ßRII expression and suppression of TGF-ß-Smad2/3 signaling. Overexpression of miR-18a-5p prevented bleomycin-induced EMT of PMC and inhibited bleomycin-induced sub-pleural fibrosis in mice. Taken together, our data indicate that downregulated miR-18a-5p mediates sub-pleural pulmonary fibrosis through upregulation of its target, TGF-ßRII, and that overexpression of miR-18a-5p might therefore provide a novel approach to the treatment of IPF.

Imbalance between subsets of CD8(+) peripheral blood T cells in patients with chronic obstructive pulmonary disease.

Background. CD8(+) T lymphocytes are known to play a critical role in the pathogenesis of chronic obstructive pulmonary disease (COPD). However, systematic analyses of CD8(+) T cell (Cytotoxic T cells, Tc) subsets in COPD patients have yet to be well conducted. Methods. The whole Tc subsets, including Tc1/2/10/17, CD8(+) regulatory T cells (Tregs) and CD8(+) α7(+) T cells, were quantified by flow cytometry in peripheral blood from 24 stable COPD subjects (SCOPD), 14 patients during acute exacerbations (AECOPD), and 14 healthy nonsmokers (HN). Results. Acute exacerbations of COPD were accompanied by elevated levels of circulating CD8(+) T cells. Tc1 cells were increased in both SCOPD and AECOPD patients, whereas the percentage of Tc2 cells was decreased in SCOPD patients but remained normal in AECOPD patients. Tc17 cells were increased only in AECOPD patients, and the percentage of Tc10 cells was reduced in both SCOPD and AECOPD patients. The imbalances of pro/anti-inflammatory Tc subsets observed in COPD may be caused by the lack of Tc10 cells and the impaired anti-inflammatory capacity of CD8(+) Tregs. Conclusions. The imbalances between subsets of CD8(+) peripheral blood T cells contribute to the immune response dysfunction in COPD pathogenesis.

Activation of calpain by renin-angiotensin system in pleural mesothelial cells mediates tuberculous pleural fibrosis.

Pleural fibrosis is defined as an excessive deposition of extracellular matrix (ECM) components that results in destruction of the normal pleural tissue architecture. It can result from diverse inflammatory conditions, especially tuberculous pleurisy. Pleural mesothelial cells (PMCs) play a pivotal role in pleural fibrosis. Calpain is a family of calcium-dependent endopeptidases, which plays an important role in ECM remodeling. However, the role of calpain in pleural fibrosis remains unknown. In the present study, we found that tuberculous pleural effusion (TPE) induced calpain activation in PMCs and that inhibition of calpain prevented TPE-induced collagen-I synthesis and cell proliferation of PMCs. Moreover, our data revealed that the levels of angiotensin (ANG)-converting enzyme (ACE) were significantly higher in pleural fluid of patients with TPE than those with malignant pleural effusion, and ACE-ANG II in TPE resulted in activation of calpain and subsequent triggering of the phosphatidylinositol 3-kinase (PI3K)/Akt/NF-κB signaling pathway in PMCs. Finally, calpain activation in PMCs and collagen depositions were confirmed in pleural biopsy specimens from patients with tuberculous pleurisy. Together, these studies demonstrated that calpain is activated by renin-angiotensin system in pleural fibrosis and mediates TPE-induced collagen-I synthesis and proliferation of PMCs via the PI3K/Akt/NF-κB signaling pathway. Calpain in PMCs might be a novel target for intervention in tuberculous pleural fibrosis.

Cigarette Smoke Disturbs the Survival of CD8+ Tc/Tregs Partially through Muscarinic Receptors-Dependent Mechanisms in Chronic Obstructive Pulmonary Disease.

BACKGROUND: CD8+ T cells (Cytotoxic T cells, Tc) are known to play a critical role in the pathogenesis of smoking related airway inflammation including chronic obstructive pulmonary disease (COPD). However, how cigarette smoke directly impacts systematic CD8+ T cell and regulatory T cell (Treg) subsets, especially by modulating muscarinic acetylcholine receptors (MRs), has yet to be well elucidated. METHODS: Circulating CD8+ Tc/Tregs in healthy nonsmokers (n = 15), healthy smokers (n = 15) and COPD patients (n = 18) were evaluated by flow cytometry after incubating with anti-CD3, anti-CD8, anti-CD25, anti-Foxp3 antibodies. Peripheral blood T cells (PBT cells) from healthy nonsmokers were cultured in the presence of cigarette smoke extract (CSE) alone or combined with MRs agonist/antagonist for 5 days. Proliferation and apoptosis were evaluated by flow cytometry using Ki-67/Annexin-V antibodies to measure the effects of CSE on the survival of CD8+ Tc/Tregs. RESULTS: While COPD patients have elevated circulating percentage of CD8+ T cells, healthy smokers have higher frequency of CD8+ Tregs. Elevated percentages of CD8+ T cells correlated inversely with declined FEV1 in COPD. CSE promoted the proliferation and inhibited the apoptosis of CD8+ T cells, while facilitated both the proliferation and apoptosis of CD8+ Tregs. Notably, the effects of CSE on CD8+ Tc/Tregs can be mostly simulated or attenuated by muscarine and atropine, the MR agonist and antagonist, respectively. However, neither muscarine nor atropine influenced the apoptosis of CD8+ Tregs. CONCLUSION: The results imply that cigarette smoking likely facilitates a proinflammatory state in smokers, which is partially mediated by MR dysfunction. The MR antagonist may be a beneficial drug candidate for cigarette smoke-induced chronic airway inflammation.

Crosstalk between calpain activation and TGF-ß1 augments collagen-I synthesis in pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease of unknown cause that typically leads to respiratory failure and death within 3-5years of diagnosis. TGF-ß1 is considered a major profibrotic factor. However, TGF-ß1 is necessary but not sufficient to the pathogenesis of fibrotic lesion of the lungs. Recent observations have revealed that calpain, a calcium dependent protease, plays a pivotal role in tissue remodeling and fibrosis. However, the mechanism of calpain mediating pulmonary fibrosis is not understood. Calpain conditional knockout (ER-Cre(+/-)capns1(flox/flox)) mice and primary human lung fibroblasts (HLFs) were used here to investigate the relationship between calpain and TGF-ß1. Calpain knockout mice were protected from fibrotic effects of bleomycin. Bleomycin induced increases in TGF-ß1 via calpain activation in HLFs. Moreover, TGF-ß1 also activated calpain. This crosstalk between calpain activation and TGF-ß1 triggered the downstream signaling pathway including TGF-ß1 Smad2/3 and non-Smad (Akt) pathways, as well as collagen-I synthesis. Taken together, our data indicate that the crosstalk between calpain activation and TGF-ß1 augments collagen-I synthesis in HLFs and in pulmonary fibrosis. Intervention in the crosstalk between calpain activation and TGF-ß1 is a novel potential strategy to prevent pulmonary fibrosis.

Bleomycin induced epithelial-mesenchymal transition (EMT) in pleural mesothelial cells.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease characterized by the development of subpleural foci of myofibroblasts that contribute to the exuberant fibrosis. Recent studies revealed that pleural mesothelial cells (PMCs) undergo epithelial-mesenchymal transition (EMT) and play a pivotal role in IPF. In animal model, bleomycin induces pulmonary fibrosis exhibiting subpleural fibrosis similar to what is seen in human IPF. It is not known yet whether bleomycin induces EMT in PMCs. In the present study, PMCs were cultured and treated with bleomycin. The protein levels of collagen-I, mesenchymal phenotypic markers (vimentin and α-smooth muscle actin), and epithelial phenotypic markers (cytokeratin-8 and E-cadherin) were measured by Western blot. PMC migration was evaluated using wound-healing assay of culture PMCs in vitro, and in vivo by monitoring the localization of PMC marker, calretinin, in the lung sections of bleomycin-induced lung fibrosis. The results showed that bleomycin induced increases in collagen-I synthesis in PMC. Bleomycin induced significant increases in mesenchymal phenotypic markers and decreases in epithelial phenotypic markers in PMC, and promoted PMC migration in vitro and in vivo. Moreover, TGF-ß1-Smad2/3 signaling pathway involved in the EMT of PMC was demonstrated. Taken together, our results indicate that bleomycin induces characteristic changes of EMT in PMC and the latter contributes to subpleural fibrosis.

Cigarette smoking promotes inflammation in patients with COPD by affecting the polarization and survival of Th/Tregs through up-regulation of muscarinic receptor 3 and 5 expression.

BACKGROUND: CD4+ T cells in the lung are involved in the pathogenesis of chronic obstructive pulmonary disease (COPD), although CD4+ T cell subsets and the direct effect of smoking on these cells, especially the expression of MRs, have not been comprehensively examined. METHODS: First, circulating CD4+ T cell subsets in healthy nonsmokers, patients with SCOPD and patients with AECOPD were evaluated by flow cytometry. Then, differentiation experiments were carried out using RT-PCR, and Ki-67/Annexin V antibodies were used to measure proliferation and apoptosis. We also explored the impact of CSE on the differentiation and survival of CD4+Th/Tregs and examined the expression of MRs in healthy nonsmokers and patients with SCOPD. RESULTS: We found the percentages of circulating Th1 and Th17 cells were increased in patients with AECOPD, while the percentage of Th2 cells was decreased in patients with SCOPD. The percentages of Th10 cells were decreased in both patients with SCOPD and patients with AECOPD, while the percentages of Tregs were increased. In addition, the percentages of CD4+α-7+ T cells were decreased in patients with SCOPD and patients with AECOPD. However, only the decrease observed in patients with AECOPD was significant. In vitro studies also revealed MR expression affected the polarization of T cells, with different CD4+ T cell subtypes acquiring different MR expression profiles. The addition of CSE facilitated CD4+ T cell polarization towards pro-inflammatory subsets (Th1 and Th17) and affected the survival of CD4+ T cells and Treg cells by up-regulating the expression of MR3 and 5, resulting in an imbalance of CD4+ T cell subsets. CONCLUSIONS: Our findings suggest an imbalance of circulating CD4+ T cell subsets is involved in COPD pathogenesis in smokers. Cigarette smoking may contribute to this imbalance by affecting the polarization and survival of Th/Tregs through the up-regulation of MR3 and MR5.

Treg/IL-17 ratio and Treg differentiation in patients with COPD.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by chronic pulmonary and systematic inflammation. An abnormal adaptive immune response leads to an imbalance between pro- and anti-inflammatory processes. T-helper (Th), T-cytotoxic (Tc) and T-regulatory (Treg) cells may play important roles in immune and inflammatory responses. This study was conducted to clarify the changes and imbalance of cytokines and T lymphocyte subsets in patients with COPD, especially during acute exacerbations (AECOPD). METHODS: Twenty-three patients with stable COPD (SCOPD) and 21 patients with AECOPD were enrolled in the present study. In addition, 20 age-, sex- and weight-matched non-smoking healthy volunteers were included as controls. The serum levels of selected cytokines (TGF-ß, IL-10, TNF-α, IL-17 and IL-9) were measured by enzyme-linked immunosorbent assay (ELISA) kits. Furthermore, the T lymphocyte subsets collected from peripheral blood samples were evaluated by flow cytometry after staining with anti-CD3-APC, anti-CD4-PerCP, anti-CD8- PerCP, anti-CD25-FITC and anti-FoxP3-PE monoclonal antibodies. Importantly, to remove the confounding effects of inflammatory factors, the authors introduced a concept of "inflammation adjustment" and corrected each measured value using representative inflammatory markers, such as TNF-α and IL-17. RESULTS: Unlike the other cytokines, serum TGF-ß levels were considerably higher in patients with AECOPD relative to the control group regardless of adjustment. There were no significant differences in the percentages of either CD4+ or CD8+ T cells among the three groups. Although Tregs were relatively upregulated during acute exacerbations, their capacities of generation and differentiation were far from sufficient. Finally, the authors noted that the ratios of Treg/IL-17 were similar among groups. CONCLUSIONS: These observations suggest that in patients with COPD, especially during acute exacerbations, both pro-inflammatory and anti-inflammatory reactions are strengthened, with the pro-inflammatory reactions dominating. Although the Treg/IL-17 ratios were normal, the regulatory T cells were still insufficient to suppress the accompanying increases in inflammation. All of these changes suggest a complicated mechanism of pro- and anti-inflammatory imbalance which needs to be further investigated.

Diagnostic and prognostic value of plasma adrenomedullin in COPD exacerbation.

BACKGROUND: Adrenomedullin (ADM) is a regulatory peptide with many biological actions, but little is known about its role in patients with COPD exacerbation. The purpose of this study was to evaluate the diagnostic and prognostic value of plasma ADM levels on hospital admission in patients with COPD exacerbation. METHODS: Consecutive subjects admitted to the hospital for COPD exacerbation were included and were followed up for 1 y; in addition, subjects with stable COPD from an out-patient clinic and healthy volunteers were recruited as controls. RESULTS: Compared with healthy subjects (145 pg/mL [interquartile range {IQR} 103-290 pg/mL]), plasma ADM levels were significantly higher in subjects with COPD exacerbation (270 pg/mL [IQR 170-510 pg/mL], P = .001) and in subjects with stable COPD (400 pg/mL [IQR 210-525 pg/mL], P < .001). In subjects with COPD exacerbation, ADM levels were significantly elevated during exacerbation (560 pg/mL [IQR 495-630 pg/mL]) compared with the recovery phase (470 pg/mL [IQR 393-553 pg/mL], P = .01) and the stable phase (200 pg/mL [IQR 143-308 pg/mL], P < .001). In receiver operating characteristic analysis, in subjects with COPD exacerbation, ADM had high diagnostic accuracy in differentiating between exacerbation and the stable phase (area under the curve 0.97, 95% CI 0.93-1.02, P < .001). In Cox regression analysis, plasma ADM was not independently associated with 1-y survival (P = .97), but it could accurately predicted the need for ICU care (hazard ratio 1.37, 95% CI 1.09-1.72, P = .008). CONCLUSIONS: Plasma ADM is a valuable biomarker to confirm COPD exacerbation; furthermore, plasma ADM independently predicts the need of ICU care, although it is not associated with long-term mortality in patients with COPD exacerbation.

Reactive oxygen species and x-ray disrupted spontaneous [Ca²âº]I oscillation in alveolar macrophages.

Radiation leads to a rapid burst of reactive oxygen species (ROS), which is considered to be one of the major causes of radiation-induced injury. ROS have previously been shown to induce changes in cytosolic Ca²âº ([Ca²âº]i) including [Ca²âº]i oscillation. However, the role of radiation in [Ca²âº]i oscillation is poorly understood. The purpose of this study was to identify the effect of ROS and X ray on [Ca²âº]i oscillation, as well as their role in radiation-induced lung injury. Alveolar macrophages were cultured in the absence and presence of different doses of hydrogen peroxide (H2O2) or exposed to X-ray irradiation with or without pretreatment of diphenyleneiodonium chloride (DPI, an inhibitor of NADPH oxidases) or tetrandrine (TET, a calcium entry blocker) and cytosolic Ca²âº concentration was detected by fluorescent Ca²âº indicator Fura-2. Rat radiation lung injury was induced in vivo by using 40 Gy X ray and DPI or TET was used to prevent radiation-induced lung injury. The results showed that there was spontaneous [Ca²âº]i oscillation in alveolar macrophages under normal conditions, and treatment of H2O2 (100-500 µM) or 2 Gy X ray inhibited the spontaneous [Ca²âº]i oscillation and induced [Ca²âº]i rise. TET abolished H2O2 or X ray induced [Ca²âº]i rise in alveolar macrophages, and attenuated X ray- induced rat alveolitis in vivo. DPI prevented X-ray-induced inhibition of [Ca²âº]i oscillation in alveolar macrophages and prevented X-ray-induced rat alveolitis. Taken together, the data suggest that the disruption of [Ca²âº]i oscillation and induction of [Ca²âº]i rise through ROS is involved in the mechanism of radiation-induced lung injury.

Interleukin 22-producing CD4+ T cells in malignant pleural effusion.

Th22 cells have been reported to be involved in human cancers. However, differentiation and immune regulation of Th22 cells in malignant pleural effusion (MPE) remain unknown. We noted that Th22 cell numbers were increased in MPE, and that IL-22 substantially promoted the proliferation and migratory activity of A549 cells. Moreover, IL-22 could strongly facilitate intercellular adhesion of A549 cells to pleural mesothelial cell monolayers. Our data revealed that the increase in Th22 cells in MPE was due to pleural cytokines and chemokines, and that Th22 exerted an important immune regulation on cancer cells in human pleural malignant environment.

CD39+ regulatory T cells suppress generation and differentiation of Th17 cells in human malignant pleural effusion via a LAP-dependent mechanism.

BACKGROUND: Both regulatory T cells (Tregs) and T helper IL-17-producing cells (Th17 cells) have been found to be involved in human malignancies, however, the possible implication of Tregs in regulating generation and differentiation of Th17 cells in malignant pleural effusion remains to be elucidated. METHODS: The numbers of both CD39(+)Tregs and Th17 cells in malignant pleural effusion and peripheral blood from patients with lung cancer were determined by flow cytometry. The regulation and mechanism of Tregs on generation and differentiation of Th17 cells were explored. RESULTS: Both CD39(+)Tregs and Th17 cells were increased in malignant pleural effusion when compared with blood, and the numbers of CD39(+)Tregs were correlated negatively with those of Th17 cells. It was also noted that high levels of IL-1ß, IL-6, and TGF-ß1 could be observed in malignant pleural effusion when compared the corresponding serum, and that pleural CD39(+)Tregs could express latency-associated peptide on their surface. When naïve CD4(+) T cells were cocultured with CD39(+)Tregs, Th17 cell numbers decreased as CD39(+)Treg numbers increased, addition of the anti-latency-associated peptide mAb to the coculture reverted the inhibitory effect exerted by CD39(+)Tregs. CONCLUSIONS: Therefore, the above results indicate that CD39(+)Tregs inhibit generation and differentiation of Th17 cells via a latency-associated peptide-dependent mechanism.

Diagnostic accuracy of T-cell interferon-γ release assays in tuberculous pleurisy: a meta-analysis.

BACKGROUND AND OBJECTIVE: The diagnosis of tuberculous pleurisy by analysis of pleural fluid using standard diagnostic tools is difficult. Recently, T-cell interferon-γ release assays (IGRA) have been introduced for the diagnosis of tuberculous pleurisy. The aim of the present meta-analysis was to establish the overall diagnostic accuracy of IGRA on both pleural fluid and peripheral blood, for diagnosing tuberculous pleurisy. METHODS: A systematic review was performed of English language publications. Sensitivity, specificity and other measures of the accuracy of IGRA for the diagnosis tuberculous pleurisy using both pleural fluid and blood were pooled using a random-effects model or a fixed-effects model. Receiver operating characteristic curves were used to summarize overall test performance. RESULTS: Seven out of eight studies met the inclusion criteria. The summary estimates of sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, positive predictive value, negative predictive value and diagnostic odds ratio were, for pleural fluid: 0.75, 0.82, 3.49, 0.24, 0.85, 0.70 and 19.04, respectively; and for blood: 0.80, 0.72, 2.86, 0.28, 0.78, 0.74 and 11.06, respectively. CONCLUSIONS: As almost 20% of non-tuberculosis patients would be erroneously treated for tuberculosis and 25% of patients with tuberculous pleurisy would be missed, pleural fluid IGRA are not useful for the clinical diagnosis of tuberculous pleurisy.