Cigarette smoke alters IL-33 expression and release in airway epithelial cells

AbstractAirway epithelium is a regulator of innate immune responses to a variety of insults including cigarette smoke. Cigarette smoke alters the expression and the activation of Toll Like Receptor 4 (TLR4), an innate immunity receptor. IL-33, an alarmin, increases innate immunity Th2 responses. The aims of this study were to explore whether mini-bronchoalveolar lavage (mini-BAL) or sera from smokers have altered concentrations of IL-33 and whether cigarette smoke extracts (CSE) alter both intracellular expression (mRNA and protein) and release of IL-33 in bronchial epithelial cells. The role of TLR4 in the expression of IL-33 was also explored.Mini-BALs, but not sera, from smokers show red…

Notch-1 decreased expression contributes to leptin receptor downregulation in nasal epithelium from allergic turbinates

Abstract BACKGROUND: Allergic rhinitis is characterized by a remodeling of nasal epithelium. Since the Notch and TGF-β signaling pathways are known to be involved in cell differentiation and remodeling processes and leptin adipokine has already been identified as a marker for homeostasis in human bronchial and nasal epithelial cells of asthmatics, roles played by these pathways have been investigated for chronic allergic rhinitis. METHODS: The leptin/leptin receptor expression has been investigated in a study with 40 biopsies from allergic (AR, n = 18) and non-allergic (C, n = 22) inferior turbinates, using immunohistochemistry, immunofluorescence staining and RT-PCR. In addition, extracts …

Effect of indomethacin on the kinetics of tumour necrosis factor alpha release and tumour necrosis factor alpha gene expression by human blood monocytes

Summary In this investigation we have examined the effects of indomethacin, an inhibitor of the cyclooxygenase pathway of arachidonic acid, upon the kinetics of the release of tumour necrosis factor alpha (TNF) and of the expression of TNF gene by lipopolysaccharide (LPS)-stimulated human blood monocytes (BM). Following stimulation of BM with LPS, TNF was released within 2 h, reached peak values at 8 h and declined at subsequent time-points (24 and 48 h). Indomethacin (10−5 m ) slightly stimulated the production of TNF at 2, 4, and 8 h and prevented the decline of TNF observed at 24 and 48 h. This effect was related to the persistence of TNF synthesis, as demonstrated by kinetics evaluation…

Relationship among Body Composition, Adipocytokines, and Irisin on Exercise Capacity and Quality of Life in COPD: A Pilot Study

Adipose tissue is an endocrine organ that interferes with the severity of chronic obstructive pulmonary disease (COPD). Although inflammatory markers, body composition, and nutritional status have a significant impact on pulmonary function, the real contribution of adipocytokines and myokines in COPD is still controversial. We aimed to evaluate the role played by the body composition, leptin, adiponectin, haptoglobin, and irisin on the functional exercise capacity, respiratory function, and quality of life (QoL) in COPD. In 25 COPD (20% GOLD-1; 60% GOLD-2; 20% GOLD-3) patients and 26 matched control subjects, we find that leptin, total adiponectin and haptoglobin are significantly increased…

Leptin and TGF-β1 Downregulate PREP1 Expression in Human Adipose-Derived Mesenchymal Stem Cells and Mature Adipocytes

International audience; Adipose tissue is widely recognized as an extremely active endocrine organ producing adipokines as leptin that bridge metabolism and the immune system. Pre-B-cell leukemia homeobox (Pbx)-regulating protein-1 (PREP1) is a ubiquitous homeodomain transcription factor involved in the adipogenic differentiation and insulin-sensitivity processes. Leptin, as pleiotropic adipokine, and TGF-β, known to be expressed by primary pre-adipocytes [adipose-derived stem cells (ASCs)] and mature differentiated adipocytes, modulate inflammatory responses. We aimed to assess for the first time if leptin and TGF-β interfere with PREP1 expression in both ASCs and mature differentiated adi…

Hypoallergenic fragment of Par j 2 increases functional expression of Toll-like receptors in atopic children.

Background: Parietaria judaica (Par j) is one of the main causes of allergy in the Mediterranean countries. The activation of Toll-like receptor 4 (TLR4) by lipopolysaccharide (LPS) inhibits nasal inflammation of atopic children. Objective:  To examine, in vivo and in vitro, the effect of recombinant Par j 2 (rPar j 2) and of its fragments (1–55 and 52–102) on atopic children. Methods:  We used skin prick test for in vivo evaluations. We assessed, in vitro, in peripheral blood mononuclear cells (PBMC), the effect of rPar j 2 and of the two fragments on neutrophil chemotaxis, on CD45RO, on TLR2 and TLR4 expression, on LPS binding and on interferon (IFN)-γ release, by a microchemotaxis chambe…

Antitumour activity of mononuclear phagocytes: role of tumour necrosis factor alpha.

Tumour necrosis factor alpha (TNF) is a cytokine produced by mononuclear phagocytes (MP) originally discovered for its cytotoxic activity on tumour cell targets. It was subsequently demonstrated that, in addition to its oncolytic potential, TNF exerts a wide variety of activities on the host defensive system against malignancies. This article briefly reviews the current concepts on the role of TNF in the antitumour activity of MP.

Cigarette smoke increases BLT2 receptor functions in bronchial epithelial cells: in vitro and ex vivo evidence

Summary Leukotriene B4 (LTB4) is a neutrophil chemotactic molecule with important involvement in the inflammatory responses of chronic obstructive pulmonary disease (COPD). Airway epithelium is emerging as a regulator of innate immune responses to a variety of insults including cigarette smoke, the major risk factor for COPD. In this study we have explored whether cigarette smoke extracts (CSE) or soluble mediators present in distal lung fluid samples (mini-bronchoalveolar lavages) from smokers alter the expression of the LTB4 receptor 2 (BLT2) and peroxisome proliferator-activated receptor-α (PPAR-α) in bronchial epithelial cells. We also evaluated the effects of CSE on the expression of i…

Cigarette smoke promotes inflammasome‐independent activation of caspase‐1 and ‐4 leading to gasdermin D cleavage in human macrophages

Mechanisms and consequences of gasdermin D (GSDMD) activation in cigarette smoke (CS)-associated inflammation and lung disease are unknown. GSDMD is a downstream effector of caspase-1, -8, and -4. Upon cleavage, GSDMD generates pores into cell membranes. Different degrees of GSDMD activation are associated with a range of physiological outputs ranging from cell hyperactivation to pyroptosis. We have previously reported that in human monocyte-derived macrophages CS extract (CSE) inhibits the NLRP3 inflammasome and shifts the response to lipopolysaccharide (LPS) towards the TLR4-TRIF axis leading to activation of caspase-8, which, in turn, activates caspase-1. In the present work, we investig…

Biomarkers of Oxidative Stress in the Upper and Lower Airways of Atopic Children.

Development of a nanostructured sensor for monitoring oxidative stress in living cells

Oxidative burden is elevated in the lung of COPD patients and is associated with aging and chronic inflammation. When overcoming physiological levels, reactive oxygen species (ROS) cause cell damage and sustain inflammation. Both lung epithelium and alveolar macrophages contribute to ROS generation. Currently, ROS generation is measured using fluorescent probes and colorimetric/fluorimetric assays. We present an amperometric nanostructured sensor for real-time detection of hydrogen peroxide (H2O2) released by living cells. The H2O2 sensing performance was evaluated through the current vs time response of platinum rod at a working potential of −0.45 V vs saturated calomel electrode acting as…

Leptin and leptin receptor expression in asthma.

Background The adipokine leptin is a potential new mediator for bronchial epithelial homeostasis. Asthma is a chronic inflammatory disease characterized by airway remodeling that might affect disease chronicity and severity. TGF-β is a tissue growth factor the dysregulation of which is associated with airway remodeling. Objective We sought to determine whether a bronchial epithelial dysfunction of the leptin/leptin receptor pathway contributes to asthma pathogenesis and severity. Methods We investigated in vitro the presence of leptin/leptin receptor on human bronchial epithelial cells. Then we studied the effect of TGF-β and fluticasone propionate on leptin receptor expression. Finally, th…

Budesonide increases TLR4 and TLR2 expression in Treg lymphocytes of allergic asthmatics

Abstract Background Reduced innate immunity responses as well as reduced T regulatory activities characterise bronchial asthma. Objectives In this study the effect of budesonide on the expression of TLR4 and TLR2 in T regulatory lymphocyte sub-population was assessed. Methods TLR4 and TLR2 expression in total peripheral blood mononuclear cells (PBMC), in CD4+/CD25+ and in CD4+/CD25− was evaluated, by flow cytometric analysis, in mild intermittent asthmatics (n = 14) and in controls (n = 11). The in vitro effects of budesonide in modulating: TLR4 and TLR2 expression in controls and in asthmatics; IL-10 expression and cytokine release (IL-6 and TNF-α selected by a multiplex assay) in asthmati…

Lymphocyte apoptosis in children with central nervous system tuberculosis: a case control study

Abstract Background Studies of the apoptosis mechanisms involved in the pathogenesis of tuberculosis have suggested that Mycobacterium tuberculosis can actively interfere with the apoptosis of infected cells. In vivo studies have been performed in adult populations but have not focused on this process in children. In the present study, we analyzed spontaneous T lymphocyte (PBT) apoptosis in the peripheral blood of children with central nervous system tuberculosis (CNS TB), before and after chemotherapy, and compared the results with healthy controls. Methods A case-control study was conducted from January 2002 to June 2009. It included 18 children with CNS TB and 17 healthy controls. Sponta…

Mononuclear Cells in Exudative Malignant Pleural Effusions

The aims of this study were to develop a methodology for the isolation of highly enriched mononuclear phagocyte populations from exudative malignant pleural effusions (EMPE) and to characterize the phenotype and functional properties of these cells. Pleural effusion mononuclear cells (PEMC) were isolated by Ficoll centrifugation of EMPE and transudative pleural effusions and allowed to adhere to plastic for 1 h to obtain a pleural effusion mononuclear adherent cell (PEMAC) fraction. Only 66.0±4.2 percent of PEMAC ingested latex particles, indicating that a significant proportion of PEMAC were not phagocytic cells. Latex-positive PEMAC had the morphologic appearance of macrophages and staine…

TLR4 Up-regulation and Reduced Foxp3 Expression in Mechanically Ventilated Smokers with Obstructive Chronic Bronchitis

Background: Chronic bronchitis (CB) is a risk factor in chronic obstructive pulmonary disease (COPD) for accelerated lung function decline and increased mortality. The lung and systemic inflammatory and immunological profile of COPD patients with CB which acutely experience respiratory failure upon a disease exacerbation is unknown. Methods: In this study, we explored the expression of Foxp3 by western blot analysis, TLR4 by immunocytochemistry and the concentrations of IP-10 and IL-8 by ELISA in the mini-bronchoalveolar lavages (mini-BAL) and in the peripheral blood of patients with respiratory failure requiring intubation and mechanical ventilation. The recruited subjects were separated i…

Role of prostaglandin E2 in the invasiveness, growth and protection of cancer cells in malignant pleuritis.

The recurrence of pleural effusions is a common event in a variety of neoplastic diseases. The objective of this study was to identify the mechanisms promoting the homing and growth of cancer cells within the pleural space. A cancer cell line recovered from malignant pleural fluids (lung adenocarcinoma cell line) that constitutively expresses cyclooxygenase 2 (COX-2) and all types of prostaglandin receptors was studied. It was first demonstrated using a matrigel system, that malignant pleural fluids increase the invasiveness of adenocarcinoma cells more than congestive heart failure (CHF) pleural fluids. Moreover, exposure to exudative malignant, but not to CHF pleural fluids, increased the…

Multiple in vitro and in vivo regulatory effects of budesonide in CD4+ T lymphocyte subpopulations of allergic asthmatics.

Abstract BACKGROUND: Increased activation and increased survival of T lymphocytes characterise bronchial asthma. OBJECTIVES: In this study the effect of budesonide on T cell survival, on inducible co-stimulator T cells (ICOS), on Foxp3 and on IL-10 molecules in T lymphocyte sub-populations was assessed. METHODS: Cell survival (by annexin V binding) and ICOS in total lymphocytes, in CD4+/CD25+ and in CD4+/CD25- and Foxp3 and IL-10 in CD4+/CD25+ and in CD4+/CD25-cells was evaluated, by cytofluorimetric analysis, in mild intermittent asthmatics (n = 19) and in controls (n = 15). Allergen induced T lymphocyte proliferation and the in vivo effects of budesonide in mild persistent asthmatics (n =…

Salmeterol Xinafoate (SX) loaded into mucoadhesive solid lipid microparticles for COPD treatment

Chronic obstructive pulmonary disease (COPD) is one of the main health problems worldwide. It is characterised by chronic inflammation in the lungs that leads to progressive, chronic, largely irreversible airflow obstruction. The use of long-acting β agonists remain today the frontline treatment for COPD with the aim of minimizing side effects and enhancing therapeutic usefulness. To this purpose, in this paper, mucoadhesive solid lipid microparticles (SLMs) containing a long-acting β-2 agonist, Salmeterol Xinafoate (SX) were prepared, characterised (size, z-potential, aerodynamic diameter, turbidimetric evaluations, drug loading and entrapping efficiency) and tested in a model of bronchial…

Lack of Dystrophin Affects Bronchial Epithelium inmdxMice

Mild exercise training may positively affect the course of Duchenne Muscular Dystrophy (DMD). Training causes mild bronchial epithelial injury in both humans and mice, but no study assessed the effects of exercise in mdx mice, a well known model of DMD. The airway epithelium was examined in mdx (C57BL/10ScSn-Dmdmdx) mice, and in wild type (WT, C57BL/10ScSc) mice either under sedentary conditions (mdx-SD, WT-SD) or during mild exercise training (mdx-EX, WT-EX). At baseline, and after 30 and 45 days of training (5 d/wk for 6 weeks), epithelial morphology and markers of regeneration, apoptosis, and cellular stress were assessed. The number of goblet cells in bronchial epithelium was much lower…

Fluticasone furoate maintains epithelial homeostasis via leptin/leptin receptor pathway in nasal cells

Leptin is involved in the lung epithelial homeostasis. Its role in the nasal tract is largely unknown. Allergic rhinitis (AR) is induced by the allergen exposure leading to consequential structural abnormalities in the nasal epithelium. Topical corticosteroids are recommended as first-line therapy in AR. Parietaria pollen is one of the most important allergenic sources in the southern Europe. In vitro, in human nasal epithelial cell line RPMI 2650, we aimed to determine whether allergen stimulation acts on leptin/leptin receptor pathway and how fluticasone furoate (FF) influences this pathway. The effects of the major allergen recombinant Par j 1 (rPar j 1), of FF, of leptin, and of TGF-b1 …

Electrochemical Quantification of H2O2 Released by Airway Cells Growing in Different Culture Media

Quantification of oxidative stress is a challenging task that can help in monitoring chronic inflammatory respiratory airway diseases. Different studies can be found in the literature regarding the development of electrochemical sensors for H2O2 in cell culture medium to quantify oxidative stress. However, there are very limited data regarding the impact of the cell culture medium on the electrochemical quantification of H2O2. In this work, we studied the effect of different media (RPMI, MEM, DMEM, Ham’s F12 and BEGM/DMEM) on the electrochemical quantification of H2O2. The used electrode is based on reduced graphene oxide (rGO) and gold nanoparticles (AuNPs) and was obtained by co-electrode…

Electrochemical sensor based on rGO/Au nanoparticles for monitoring H2O2 released by human macrophages

Abstract Increased oxidative burden contributes to the pathogenesis of most inflammatory diseases and is associated with aging and chronic inflammation. Macrophages contribute to the generation of reactive oxygen species (ROS) within inflamed tissues. Currently, ROS generation is measured using fluorescent probes and colorimetric/fluorimetric biochemical assays. Hydrogen peroxide (H2O2) diffuses through the cell membrane and can be monitored in the extracellular space. Herein, we present a sensor for H2O2 detection released by cells in culture supernatants. H2O2 sensing performance was evaluated using chronoamperometric detection. A sensitivity of 0.0641 μA μM−1 cm−2 with a limit of detecti…

Impairment of leptin/leptin receptor pathway in nasal epithelium from allergic turbinates

BACKGROUND: Allergic rhinitis (AR) is characterized by a remodeling of nasal epithelium. Leptin adipokine has been already identified as a marker of homeostasis in human bronchial epithelial of asthmatics. TGF-β is a multi-functional cytokine and conflicting findings exist regarding its role in the remodeling responses of the upper airways in allergic rhinitis. OBJECTIVE: We sought to investigate ex-vivo the expression of leptin/leptin receptor pathway and TGF-β in human nasal epithelium. METHODS: 41 biopsies of inferior turbinates obtained from allergic patients with AR (A, n = 20) and from healthy control subjects (C, n = 21) were analyzed for leptin/leptin receptor and for TGF-β 1, 2, 3 …

Cilomilast counteracts the effects of cigarette smoke in airway epithelial cells.

Abstract Cigarette smoke extracts (CSE) alter TLR4 expression and activation in bronchial epithelial cells. Cilomilast, a phosphodiesterase-4 inhibitor, inhibits cigarette smoke-induced neutrophilia. This study was aimed to explore whether cilomilast, in a human bronchial epithelial cell line (16-HBE), counteracted CSE effects. In particular, TLR4 expression, IP-10 and IL-8 release, lymphocyte and neutrophil chemotactic activity and ERK and IkBa phosphorylation in CSE and LPS-stimulated 16-HBE were assessed. CSE increased TLR4 expression, reduced IP-10 release and lymphocyte chemotactic activity and increased IL-8 release and neutrophil chemotactic activity. Cilomilast reduced TLR4 expressi…

Advances in asthma pathophysiology: stepping forward from the Maurizio Vignola experience

Maurizio Vignola was a superb and innovative researcher, who wrote seminal papers on the biology of airway epithelium in asthma. Inflammation and remodelling were the main topics of his research, mostly conducted in biopsy specimens from patients with asthma of variable severity, encompassing the entire spectrum of the disease from mild to severe asthma. His observations contributed to define the biology of asthma as we know it today, and opened the way to the personalised treatment of asthma. His group has successfully continued to investigate the biology and clinical aspects of bronchial asthma, with major interest in the clinical use of biomarkers to monitor disease activity, and in the …

Cellular and Molecular Signatures of Oxidative Stress in Bronchial Epithelial Cell Models Injured by Cigarette Smoke Extract

Exposure of the airways epithelium to environmental insults, including cigarette smoke, results in increased oxidative stress due to unbalance between oxidants and antioxidants in favor of oxidants. Oxidative stress is a feature of inflammation and promotes the progression of chronic lung diseases, including Chronic Obstructive Pulmonary Disease (COPD). Increased oxidative stress leads to exhaustion of antioxidant defenses, alterations in autophagy/mitophagy and cell survival regulatory mechanisms, thus promoting cell senescence. All these events are amplified by the increase of inflammation driven by oxidative stress. Several models of bronchial epithelial cells are used to study the molec…

Notch‐1 signaling activation sustains overexpression of interleukin 33 in the epithelium of nasal polyps

Abstract BACKGROUND: Alterations in the nasal epithelial barrier homeostasis and increased interleukin 33 (IL-33) expression contribute to the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). AIMS: As Notch-1 signaling is crucial in repair processes of mucosa, the current study assessed Notch-1/Jagged-1 signaling and IL-33 in the epithelium of nasal polyps biopsies from allergic (A-CRSwNP; n = 9) and not allergic (NA-CRSwNP; n = 9) subjects by immunohistochemistry. We also assessed, in a model of nasal epithelial cells, the effects of stimulation of Notch-1 with Jagged-1 on the expression of IL-33 (by flow cytometry, immunofluorescence, and immunocytochemistry), Jagged-1 (…

Oxidative stress and innate immunity responses in cigarette smoke stimulated nasal epithelial cells

Cigarette smoke extracts (CSE) may play a significant role in diseases of the upper airway including chronic rhinosinusitis. Even short term exposure of cigarette smoke has adverse effects on mitochondrial functions and redox homeostasis in tissues which may progress to further complications associated with chronic smoking. Cigarette smoke alters toll-like receptor 4 (TLR4) expression and activation in bronchial epithelial cells. Carbocysteine is an anti-oxidant and mucolytic agent. The effects of carbocysteine on CSE induced oxidative stress and on associated innate immune and inflammatory responses in nasal epithelial cells are largely unknown. The present study was aimed to assess in CSE…

Resolvin D1 and miR‐146a are independent distinctive parameters in children with moderate and severe asthma

To date, a large number of mediators and biomarkers have been detected in childhood asthma, with a potential role in monitoring the disease course. Lung function evaluation through spirometry, forced oscillation technique and multiple breath washout proved its role in asthma monitoring, especially for investigating small airway impairment in children with more severe symptoms. Recently, novel specialized pro-resolving mediators, such as resolvins and lipoxins, have been recognized as crucial in promoting the resolution of lung inflammation. Increasing evidence suggests that miRNAs are crucially entailed in asthma. Partial least squares discriminant analysis (plsDA) is a modern technique for…

Electrochemical sensor for evaluating oxidative stress in airway epithelial cells

Cigarette smoke exposure induces oxidative stress within the airways. Increased oxidative burden contributes to the pathogenesis of chronic lung disorders and is associated with aging and chronic inflammation. Airway epithelial cells highly contribute to Reactive Oxygen Species (ROS) generation within injured and inflamed lung tissues. Among ROS, hydrogen peroxide (H2O2) can be monitored in the extracellular space. Herein, we present an amperometric/voltammetric sensor based on gold nanoparticles and graphene oxide able to detect H2O2 with good sensitivity and selectivity. Using this sensor, H2O2 release was measured in conditioned medium from primary bronchial epithelial cells (PBEC), bron…

15-Lipoxygenase expression and 15(S)-hydroxyeicoisatetraenoic acid release and reincorporation in induced sputum of asthmatic subjects

Recent evidence shows that 15(S)-hydroxy-eicoisatetraenoic acid (15[S]-HETE) can be released and rapidly reincorporated into cellular lipids. These mechanisms exert several immunoregulatory functions that may be relevant in airway inflammation.Our purpose was to evaluate the levels of both soluble and cell-associated 15(S)-HETE and to examine 15-lipoxygenase (15-LO) messenger RNA (mRNA) expression in sputum samples obtained from 10 control and 18 asthmatic subjects.Levels of 15(S)-HETE were measured by reverse-phase HPLC separation followed by RIA in supernatants and in cell membrane-extracted phospholipids after acid hydrolysis. 15-LO mRNA was evaluated by primed in situ hybridization (PRI…

Noninvasive methods for the detection of upper and lower airway inflammation in atopic children

Background Exhaled nitric oxide (FE NO ) and exhaled breath condensate (EBC) are noninvasive methods to assess inflammation. Objective To investigate the role of the FE NO and of the EBC pH and IL-5 levels in atopic children. Methods We evaluated oral and nasal FE NO and the pH and IL-5 of oral and nasal EBC in children with atopic dermatitis (AD; n=18), allergic rhinitis (AR; n=18), intermittent asthma (n = 21), moderate persistent asthma (n = 18), and healthy controls (HCs; n=16). Results Oral FE NO was significantly increased in asthma, whereas the nasal values were increased in AR and asthma in comparison with HCs. The pH of oral EBC was lower in AD and asthma than in AR and HCs, wherea…

Carbocysteine reverses the effects of cigarette smoke and improves the effects of beclomethasone on the histone deacetylases in bronchial epithelial cells

Cigarette smoke exposure, increasing oxidative stress, may negatively affect histone deacetylase expression/activity. Histone deacetylase expression/activity and in particular HDAC2, HDAC3, and SIRT-1 may control inflammation, cell senescence and responses to corticosteroids. The effects of carbocysteine and of beclomethasone on the histone deacetylase expression/activity in human bronchial epithelial cells stimulated with cigarette smoke extracts (CSE) are largely unknown. This study was aimed to explore whether carbocysteine and beclomethasone, in a bronchial epithelial cell line (16-HBE) exposed to CSE, were able to modulate the expression/activity of HDAC2, HDAC3, and of SIRT-1. Methods…

Apigenin affects leptin/leptin receptor pathway and induces cell apoptosis in lung adenocarcinoma cell line

Abstract Background Apigenin, a common edible plant flavonoid, is a well characterised antioxidant. The adipokine leptin exerts proliferative and anti-apoptotic activities in a variety of cell types. In cancer cells, apigenin may induce a pro-apoptotic pathway whereas leptin has an anti-apoptotic role. The purpose of the study is to investigate the role of apigenin and of leptin/leptin receptor pathway on proliferation and on apoptosis in lung adenocarcinoma. Methods Immunocytochemistry, flow cytometry and RT-q-RT PCR, were used to investigate the expression and modulation of leptin receptors on the lung adenocarcinoma cell line A549 in presence or absence of apigenin and of leptin, alone o…

Innate Immunity Responses Are Differently Altered in Central and Distal Airways of COPD Patients.

Dual anti-oxidant and anti-inflammatory actions of the electrophilic cyclooxygenase-2-derived 17-oxo-DHA in lipopolysaccharide- and cigarette smoke-induced inflammation.

Abstract Background 17-Oxo-DHA is an endogenous electrophilic derivative of the omega-3 fatty acid docosahexaenoic acid (DHA) which is generated in activated macrophages by the action of cyclooxygenase-2. Methods The ability of 17-oxo-DHA to control inflammation and oxidative stress was tested in human macrophages (THP-1) and bronchial epithelial cell line (16HBE) stimulated with cigarette smoke extract (CSE) and lipopolysaccharide (LPS). All data were further confirmed using primary bronchial epithelial cells, alveolar macrophages and peripheral blood mononuclear cells. Results 17-Oxo-DHA was a strong inducer of the anti-oxidant response promoting Nrf2 nuclear accumulation, leading to the …

Polyaspartamide-based nanoparticles loaded with fluticasone propionate and the in vitro evaluation towards cigarette smoke effects

This paper describes the evaluation of polymeric nanoparticles (NPs) as a potential carrier for lung administration of fluticasone propionate (FP). The chosen polymeric material to produce NPs was a copolymer based on α,β-poly(N-2-hydroxyethyl)-d,l-aspartamide (PHEA) whose backbone was derivatised with different molecules, such as poly(lactic acid) (PLA) and polyethylenglycol (PEG). The chosen method to produce NPs from PHEA-PLA-PEG2000 was the method based on high-pressure homogenization and subsequent solvent evaporation by adding Pluronic F68 during the process and trehalose before lyophilisation. Obtained colloidal FP-loaded NPs showed a slightly negative surface charge and nanometric d…

Effects in cigarette smoke stimulated bronchial epithelial cells of a corticosteroid entrapped into nanostructured lipid carriers

Background Nanomedicine studies have showed a great potential for drug delivery into the lung. In this manuscript nanostructured lipid carriers (NLC) containing Fluticasone propionate (FP) were prepared and their biocompatibility and effects in a human bronchial epithelial cell line (16-HBE) stimulated with cigarette smoke extracts (CSE) were tested. Results Biocompatibility studies showed that the NLC did not induce cell necrosis or apoptosis. Moreover, it was confirmed that CSE increased intracellular ROS production and TLR4 expression in bronchial epithelial cells and that FP-loaded NLC were more effective than free drug in modulating these processes. Finally, the nanoparticles increased…

Carbocysteine regulates innate immune responses and senescence processes in cigarette smoke stimulated bronchial epithelial cells

Cigarette smoke represents the major risk factor for chronic obstructive pulmonary disease (COPD). Cigarette smoke extracts (CSE) alter TLR4 expression and activation in bronchial epithelial cells. Carbocysteine, an anti-oxidant and mucolytic agent, is effective in reducing the severity and the rate of exacerbations in COPD patients. The effects of carbocysteine on TLR4 expression and on the TLR4 activation downstream events are largely unknown. This study was aimed to explore whether carbocysteine, in a human bronchial epithelial cell line (16-HBE), counteracted some pro-inflammatory CSE-mediated effects. In particular, TLR4 expression, LPS binding, p21 (a senescence marker), IL-8 mRNA and…

Increased expression of IL-19 in the epithelium of patients with chronic rhinosinusitis and nasal polyps.

Background : Chronic rhinosinusitis (CRS) is an inflammation of the nose and of the paranasal sinuses. The involvement of the respiratory epithelium in the mechanisms of CRS is poorly understood. Aims : Among proteins expressed by nasal epithelial cells in CRS, IL-19 may have key functions. We here aimed to determine the expression and regulation of IL-19. Methods : Nasal biopsies from normal subjects (n = 12), subjects with CRS but without nasal polyps (NP) (CRSsNP, n = 12) and with CRS with NP (CRSwNP, n = 15) were collected. Human Asthma Gene Array and real-time PCR were used to evaluate gene expression, western blot analysis and immunohistochemistry for protein expression. Results for I…

Mucoadhesive solid lipid microparticles for controlled release of a corticosteroid in the chronic obstructive pulmonary disease treatment.

Therapeutic efficacy of inhaled drugs is limited by rapid clearance from the site of action due to absorption into systemic circulation or metabolic degradation by alveolar macrophages. Drug delivery systems offer new solutions to clinical problems especially in the treatment of pulmonary diseases. In particular, Solid Lipid Microparticles (SLM) in the range of 3-5 µm are suggested as systems for delivery of therapeutics to the lung as, because of their size, they are able to deposit into secondary bronchi, avoiding systemic absorption typical of alveolar regions. Here, we describe two novel different SLMs prepared with chitosan and alginate for sustained release of fluticasone propionate (…

Reduced apoptosis of CD8+ T-Lymphocytes in the airways of smokers with mild/moderate COPD

SummaryChronic obstructive pulmonary disease (COPD) is characterised by chronic inflammation in airways and lung parenchyma. CD8+ T-lymphocytes, crucial effector and regulatory cells in inflammation, are increased in the central and peripheral airways in COPD. The aim of this study was to assess the role of apoptosis in the accumulation of CD8+ T-lymphocytes within the airway wall in COPD. We examined the submucosa of transverse sections of central and peripheral airways from post-operative tissues from non-smokers (n = 16), smokers with normal lung function (n = 16), smokers with mild/moderate COPD (n = 16), and smokers with severe/very severe COPD (n = 9). TUNEL and immunohistochemistry t…

Comparative cytoprotective effects of carbocysteine and fluticasone propionate in cigarette smoke extract-stimulated bronchial epithelial cells

Cigarette smoke extracts (CSE) induce oxidative stress, an important feature in chronic obstructive pulmonary disease (COPD), and oxidative stress contributes to the poor clinical efficacy of corticosteroids in COPD patients. Carbocysteine, an antioxidant and mucolytic agent, is effec- tive in reducing the severity and the rate of exacerbations in COPD patients. The effects of carbocysteine on CSE-induced oxidative stress in bronchial epithelial cells as well as the comparison of these antioxidant effects of carbocysteine with those of fluticasone propionate are unknown. The present study was aimed to assess the effects of carbocysteine (10−4 M) in cell survival and intracellular reactive o…

Wearable sensor for real-time monitoring of oxidative stress

Ceftaroline modulates the innate immune and host defense responses of immunocompetent cells exposed to cigarette smoke.

Abstract Background Cigarette smoke, the principal risk factor for chronic obstructive pulmonary disease (COPD), negatively influences the effectiveness of the immune system’s response to a pathogen. The antibiotic ceftaroline exerts immune-modulatory effects in bronchial epithelial cells exposed to cigarette smoke. Aims and methods The present study aims to assess the effects of ceftaroline on TLR2 and TLR4 expression, LPS binding and TNF-α and human beta defensin (HBD2) release in an undifferentiated and PMA-differentiated human monocyte cell line (THP-1) exposed or not to cigarette smoke extracts (CSE). TLR2, TLR4, and LPS binding were assessed by flow cytometry, TNF-α and HBD2 release w…

Wearable sensor for real-time monitoring of hydrogen peroxide in simulated breath

Exposure to cigarette smoke extract and lipopolysaccharide modifies cytoskeleton organization in bronchial epithelial cells

The integrity of the respiratory epithelium is crucial for airway homeostasis. Tobacco smoke exposure and recurrent infections of the airways play a crucial role in the progression and in the decline of the respiratory function in chronic obstructive pulmonary disease (COPD). The aim of this study was to detect differentially expressed proteins in a bronchial epithelial cell line (16-HBE) stimulated with cigarette smoke extract (CSE) and lipopolysaccharide (LPS), a constituent of gram-negative bacteria, alone and/or in combination, by using two-dimensional electrophoresis (2DE) analysis coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Western blot a…

Electrochemical Synthesis of Zinc Oxide Nanostructures on Flexible Substrate and Application as an Electrochemical Immunoglobulin-G Immunosensor

Immunoglobulin G (IgG), a type of antibody, represents approximately 75% of serum antibodies in humans, and is the most common type of antibody found in blood circulation. Consequently, the development of simple, fast and reliable systems for IgG detection, which can be achieved using electrochemical sandwich-type immunosensors, is of considerable interest. In this study we have developed an immunosensor for human (H)-IgG using an inexpensive and very simple fabrication method based on ZnO nanorods (NRs) obtained through the electrodeposition of ZnO. The ZnO NRs were treated by electrodepositing a layer of reduced graphene oxide (rGO) to ensure an easy immobilization of the antibodies. On I…

15(S)-HETE modulates LTB(4) production and neutrophil chemotaxis in chronic bronchitis.

We evaluated the levels of 15(S)-hydroxyeicosatetraenoic acid [15(S)-HETE] and the expression of 15-lipoxygenase (15-LO) mRNA in induced sputum obtained from 10 control and 15 chronic bronchitis subjects. 15(S)-HETE was evaluated by reverse phase high-performance liquid chromatography separation followed by specific RIA. 15-LO mRNA expression was determined by primed in situ labeling. The levels of both soluble and cell-associated 15(S)-HETE resulted significantly higher in chronic bronchitis than in control subjects. The percentage of cells expressing 15-LO mRNA was significantly higher in chronic bronchitis than in control subjects ( P < 0.01). Double staining for specific cell type ma…

Gemcitabine sensitizes lung cancer cells to Fas/FasL system-mediated killing

Gemcitabine is a chemotherapy agent commonly used in the treatment of non-small cell lung cancer (NSCLC) that has been demonstrated to induce apoptosis in NSCLC cells by increasing functionally active Fas expression. The aim of this study was to evaluate the Fas/Fas ligand (FasL) system involvement in gemcitabine-induced lung cancer cell killing. NSCLC H292 cells were cultured in the presence or absence of gemcitabine. FasL mRNA and protein were evaluated by real-time PCR, and by Western blot and flow cytometry, respectively. Apoptosis of FasL-expressing cells was evaluated by flow cytometry, and caspase-8 and caspase-3 activation by Western blot and a colorimetric assay. Cytotoxicity of ly…

Chlorinative stress in age-related diseases: A literature review

Abstract Aging is an agglomerate of biological long-lasting processes that result being inevitable. Main actors in this scenario are both long-term inflammation and oxidative stress. It has been proved that oxidative stress induce alteration in proteins and this fact itself is critically important in the pathophysiological mechanisms leading to diseases typical of aging. Among reactive species, chlorine ones such as hypochlorous acid (HOCl) are cytotoxic oxidants produced by activated neutrophils during chronic inflammation processes. HOCl can also cause damages by reacting with biological molecules. HOCl is generated by myeloperoxidase (MPO) and augmented serum levels of MPO have been desc…

Cigarette smoke increases Toll-like receptor 4 and modifies lipopolysaccharide-mediated responses in airway epithelial cells.

Airway epithelium is emerging as a regulator of innate immune responses to a variety of insults including cigarette smoke. The main goal of this study was to explore the effects of cigarette smoke extracts (CSE) on Toll-like receptor (TLR) expression and activation in a human bronchial epithelial cell line (16-HBE). The CSE increased the expression of TLR4 and the lipopolysaccharide (LPS) binding, the nuclear factor-kappaB (NF-kappaB) activation, the release of interleukin-8 (IL-8) and the chemotactic activity toward neutrophils. It did not induce TLR2 expression or extracellular signal-regulated signal kinase 1/2 (ERK1/2) activation. The LPS increased the expression of TLR4 and induced bot…

TLR4 upregulation underpins airway neutrophilia in smokers with chronic obstructive pulmonary disease and acute respiratory failure

Activation of Toll-like receptors (TLR) seems to be involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Upon TLR activation the release of defensins, including human beta defensin 2 (HBD-2), may occur. In this study, we explored the innate responses in patients with respiratory failure, with and without COPD, requiring intubation and mechanical ventilation. Mini-bronchoalveolar lavage (mini-BAL) samples were collected from nonsmoker subjects without COPD (n = 10), smokers without COPD (n = 6), and smokers with COPD (n = 15). TLR4, TLR2, and HBD-2 expression was evaluated by immunocytochemistry; interleukin (IL)-8, IP-10, and HBD-2 concentrations were evaluated by e…

Altered CD94/NKG2A and perforin expression reduce the cytotoxic activity in malignant pleural effusions.

CD94/NKG2A is an inhibitory receptor expressed by NK cells and cytotoxic lymphocytes and, upon activation by HLA-E, downregulates the cytolytic activities of these cells thus representing a tumour immune escape mechanism. This study was aimed at assessing whether cytotoxic lymphocytes (CD8+) and NK cells from malignant pleural effusions have a deregulated expression of CD94/NKG2A. The expression of membrane CD94/NKG2A and perforin was evaluated by flow-cytometry in CD8+ and NK cells from pleural effusions and autologous peripheral blood of cancer (n=19) and congestive heart failure (CHF) (n=11) patients. Intracellular CD94/NKG2A expression was evaluated by flow-cytometry in pleural effusion…

Synergistic effects of fluticasone propionate and salmeterol on in vitro T-cell activation and apoptosis in asthma

Background In asthma T cells are characterized by an increased activation state and by reduced apoptosis. Objective Because the clinical efficacy of inhaled corticosteroids combined with long-acting β 2 -agonists has been widely demonstrated in asthma, we studied, in vitro , the effect of fluticasone propionate (FP) and salmeterol alone and in combination on the activation and apoptosis of peripheral blood T cells (PBTs), on the expression of phosphorylated nuclear factor κB inhibitor (IκBα), and on the nuclear translocation of glucocorticoid receptor (GR) in PBTs from asthmatic subjects. Methods Apoptosis was evaluated on the basis of annexin V binding, whereas the expression of caspases 8…

Carbocysteine counteracts the effects of cigarette smoke on cell growth and on the SIRT1/FoxO3 axis in bronchial epithelial cells

Abstract Background Cigarette smoke may accelerate cellular senescence by increasing oxidative stress. Altered proliferation and altered expression of anti-aging factors, including SIRT1 and FoxO3, characterise cellular senescence. The effects of carbocysteine on the SIRT1/FoxO3 axis and on downstream molecular mechanisms in human bronchial epithelial cells exposed to cigarette smoke are largely unknown. Aims Aim of this study was to explore whether carbocysteine modulated SIRT1/FoxO3 axis, and downstream molecular mechanisms associated to cellular senescence, in a bronchial epithelial cell line (16-HBE) exposed to cigarette smoke. Methods 16HBE cells were stimulated with/without cigarette …

Pleural Mesothelial Cells Express Both BLT2 and PPARα and Mount an Integrated Response to Pleural Leukotriene B4

Abstract Leukotriene B4 (LTB4) plays a crucial role in the recruitment of neutrophils into the pleural space. We identified for the first time the mechanisms by which LTB4 interacts with mesothelial cells and recruits neutrophils in the pleural compartment. Primary pleural mesothelial cells express both the proinflammatory receptor for LTB4 BLT2, and the anti-inflammatory receptor for LTB4, PPARα. Parapneumonic pleural effusions highly increase BLT2 expression and, via BLT2 activation, increase the adhesion between mesothelial cells and neutrophils and the expression of ICAM-1 on mesothelial cells. The block of PPARα further increases both cell adhesion and ICAM-1 expression. BLT2 activatio…