0000000000124120
AUTHOR
Celia Sanz
Phosphodiesterase-4 inhibition improves corticosteroid insensitivity in pulmonary endothelial cells under oxidative stress.
Several clinical studies have shown that smoking in asthmatics and chronic obstructive pulmonary disease patients is closely associated with corticosteroid refractoriness. In this work, we have analyzed glucocorticoid insensitivity in human pulmonary artery endothelial cells (HPAECs) under cigarette smoke extract (CSE) exposure as well as the possible additive effects of the combination therapy with a phosphodiesterase (PDE)-4 inhibitor. Interleukin (IL)-8 was measured in cell supernatants by ELISA. Histone deacetylase (HDAC), histone acetylase (HAT), and intracellular cAMP levels were measured by colorimetric assays and enzyme immunoassay, respectively. PDE4 isotypes and glucocorticoid rec…
Roflumilast improves corticosteroid resistance COPD bronchial epithelial cells stimulated with toll like receptor 3 agonist
Chronic obstructive pulmonary disease (COPD) is characterised by chronic pulmonary inflammation punctuated by periods of viral exacerbations. Recent evidence suggests that the combination of roflumilast with corticosteroids may improve the compromised anti-inflammatory properties of corticosteroids in COPD. We analyzed differential and combination anti-inflammatory effects of dexamethasone and roflumilast N-oxide in human bronchial epithelial cells (HBECs) stimulated with viral toll like receptor (TLR) agonists. Lung tissue and HBECs were isolated from healthy (n = 15), smokers (n = 12) and smokers with COPD (15). TLR3 expression was measured in lung tissue and in HBECs. IL-8 secretion was …
Non-neuronal cholinergic system contributes to corticosteroid resistance in chronic obstructive pulmonary disease patients
Background: Inhaled corticosteroid (ICS) with long-acting beta-2 agonists is a well-documented combination therapy for chronic obstructive pulmonary disease (COPD) based on its additive anti-inflammatory properties. By contrast, the recommendation of ICS in combination with long-acting muscarinic antagonist (LAMA) is not evidence-based. In this study, neutrophils obtained from COPD patients were used to compare the anti-inflammatory effects of aclidinium bromide (a long-acting muscarinic antagonist) with corticosteroids and their potential additive effect. Methods: Human sputum and blood neutrophils were isolated from healthy individuals ( n = 37), patients with stable COPD ( n = 52) and th…
Effects of dantrolene on the responses to methylxanthines in the isolated guinea-pig trachea.
The effect of dantrolene on the responses to methylxanthines (caffeine and theophylline) and a beta-adrenoceptor agonist (salbutamol) was studied in isolated guinea-pig trachea. Caffeine and theophylline (1 microM-10 mM) and salbutamol (1 nM-10 microM) produced concentration-dependent relaxation of spontaneous and stimulated (acetylcholine 1 mM) tone. Responses to high concentrations (10 mM) of caffeine and theophylline added to tracheal strips either unstimulated (spontaneous tone) or indomethacin (2.8 microM)-treated were reversed to contractions in the presence of dantrolene (3 microM-0.3 mM). This effect was not observed for salbutamol or when relaxant responses to the agonists were gen…
The effects of phorbol 12,13-diacetate on responses of guinea-pig isolated trachea to methylxanthines, isoprenaline and ryanodine
1. Using guinea-pig isolated trachea, we have studied how phorbol 12,13-diacetate (PDA) modulates mechanical responses of the tissue to methylxanthines, isoprenaline and ryanodine. 2. Caffeine (10 microM-5 mM), theophylline (10 microM-5 mM) and isoprenaline (1 nM-1 microM), each inhibited the spontaneous tone of the trachea. Pretreatment with PDA (0.1-10 microM) converted relaxant responses to high concentrations of the methylxanthines into contractions. PDA produced no equivalent effect against isoprenaline. Pretreatment with verapamil (1 or 10 microM), nifedipine (0.1 microM) or incubation with Ca(2+)-free, EGTA (0.1 mM)-containing physiological salt solution (PSS) suppressed the contract…
DIFFERENT ABILITY OF TRIFLUOPERAZINE TO INHIBIT AGONIST-INDUCED CONTRACTION OF LUNG PARENCHYMA STRIPS FROM CONTROL AND SENSITIZED GUINEA-PIGS
Abstract There is increasing interest in the therapeutic potential of calcium antagonists in asthma. Among them the use of calmodulin antagonists deserves consideration. In the present work the effect of trifluoperazine on contractions generated by different mechanisms (CaCl2, KCl, acetylcholine, histamine and 5-hydroxytryptamine) in lung parenchyma strip isolated from control and actively sensitized guinea-pigs has been studied. Trifluoperazine produced both in unsensitized and sensitized lung strips, a concentration-dependent, right, downward displacement of the concentration-response curves to the agonists used, although the sensitization procedure resulted in a potentiation in the abili…
Evaluation of Mucociliary Clearance by Three Dimension Micro-CT-SPECT in Guinea Pig: Role of Bitter Taste Agonists
Different image techniques have been used to analyze mucociliary clearance (MCC) in humans, but current small animal MCC analysis using in vivo imaging has not been well defined. Bitter taste receptor (T2R) agonists increase ciliary beat frequency (CBF) and cause bronchodilation but their effects in vivo are not well understood. This work analyzes in vivo nasal and bronchial MCC in guinea pig animals using three dimension (3D) microCT-SPECT images and evaluates the effect of T2R agonists. Intranasal macroaggreggates of albumin-Technetium 99 metastable (MAA-Tc99m) and lung nebulized Tc99m albumin nanocolloids were used to analyze the effect of T2R agonists on nasal and bronchial MCC respecti…
Simvastatin Increases the Ability of Roflumilast N-oxide to Inhibit Cigarette Smoke-Induced Epithelial to Mesenchymal Transition in Well-differentiated Human Bronchial Epithelial Cells in vitro.
Cigarette smoking contributes to epithelial-mesenchymal transition (EMT) in COPD small bronchi as part of the lung remodeling process. We recently observed that roflumilast N-oxide (RNO), the active metabolite of the PDE4 inhibitor roflumilast, prevents cigarette smoke-induced EMT in differentiated human bronchial epithelial cells. Further, statins were shown to protect renal and alveolar epithelial cells from EMT. To analyze how RNO and simvastatin (SIM) interact on CSE-induced EMT in well-differentiated human bronchial epithelial cells (WD-HBEC) from small bronchi in vitro. Methods: WD-HBEC were stimulated with CSE (2.5%). The mesenchymal markers vimentin, collagen type I and α-SMA, the e…
Non-specific Hyperreactivity to Pharmacological Stimuli in Tracheal and Lung Parenchymal Strips of Actively Sensitized Guinea-pigs
Abstract The responsiveness of tracheal and lung parenchymal strips isolated from actively sensitized guinea-pigs to CaCl2 (in K+-depolarized tissue), KCl, acetylcholine and histamine was compared with that of strips from unsensitized animals. The concentration-response curves to the mentioned agonists exhibited, in the sensitized group, a left upward displacement (greater maximal effect, lesser effective concentration 50% and a steeper slope) compared with those obtained in the unsensitized group. These results indicate the existence of a non-specific increase in responsiveness in the airway smooth muscle from sensitized animals.
Bafetinib inhibits functional responses of human eosinophils in vitro
Eosinophils play a prominent role in the process of allergic inflammation. Non-receptor associated Lyn tyrosine kinases generate key initial signals in eosinophils. Bafetinib, a specific Abl/Lyn tyrosine kinase inhibitor has shown a potent antiproliferative activity in leukemic cells, but its effects on eosinophils have not been reported. Therefore, we studied the effects of bafetinib on functional and mechanistic responses of isolated human eosinophils. Bafetinib was more potent than non-specific tyrosin kinase comparators genistein and tyrphostin inhibiting superoxide anion triggered by N-formyl-Met-Leu-Phe (fMLF; 100 nM) (−log IC50=7.25±0.04 M; 6.1±0.04 M; and 6.55±0.03 M, respectively).…
Role of MUC1 in idiopathic pulmonary fibrosis
Background: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and irreversible form of fibrotic intersticial lung disease, characterized by uncontrolled fibroblast proliferative processes and alveolar type II epithelial dysfunction. MUC1 is considered as oncogenic molecule by altering signaling pathways involved in cellular proliferation and cell death. Objective: To analyze the implication of MUC1 in IPF Methods: Lung tissue from 14 healthy and 14 IPF patients was obtained. The expression of MUC1 cytoplasmic tail (CT) and its phosphorylation forms in T-1224 and Y-1229 residues were analyzed by western blot and immunohistochemistry. The effect of MUC1 on TGFβ1-Iinduced epithelia…
Responses to histamine and selective H2-receptor agonists in lung parenchymal strips from normal and sensitized guinea-pigs.
Histamine produces concentration-dependent contractions of lung parenchyma strips obtained from normal and sensitized guinea-pigs. The responsiveness of the sensitized lung strips to histamine was significantly increased compared to normal tissues. Clemizole (0.1 microM) was equally effective as an H1-antagonist in normal (dose-ratio 9.12) and sensitized (dose-ratio 9.77) tissues. The concentration-response curves to histamine were displaced to the left by cimetidine (0.1 microM to 0.1 mM) with similar dose-ratios in normal and sensitized tissues. Cimetidine enhanced maximal responses to histamine only in normal lung strips. The effects of submaximal equieffective concentrations of histamin…
MUC1-CT mediates corticosteroid responses in COPD
Background: Corticosteroid resistance is an acquired condition in chronic obstructive pulmonary disease (COPD) patients and a challenge to develop new anti-inflammatory therapies. In previous reports we showed that cytoplasmic tail of the membrain tethered mucin 1 (MUC1-CT) interacts with glucocorticoid receptor (GR) mediating corticosteroid anti-inflammatory efficacy. Objectives: To analyze the role of MUC1-CT as a key marker of corticosteroid efficacy in COPD Methods: The expression of MUC1-CT and the anti-inflammatory role of dexamethasone were evaluated in neutrophils and bronchial epithelial cells from healthy and COPD patients. Anti-inflammatory effects of dexamethasone and glucocorti…
Effect of dantrolene sodium in isolated guinea-pig trachea.
The effect of dantrolene sodium (3 microM-0.3 mM) on the spontaneous tone and responses to various contractile agonists was studied in isolated guinea-pig trachea. Dantrolene produced a concentration-related inhibition of the spontaneous tracheal tone, reaching a value of 94.8 +/- 4.8% of the relaxation induced by caffeine 10 mM. Removal of the epithelium did not affect the dantrolene-induced relaxation. Dantrolene did not alter the concentration-response curve for KCl and produced only small displacements of the concentration-response curves for CaCl2, acetylcholine and histamine, without affecting their maximal effects. Dantrolene dose relatedly inhibited the contraction induced by caffei…
COOLING-INDUCED CONTRACTION OF TRACHEA ISOLATED FROM NORMAL AND SENSITIZED GUINEA-PIGS
Fast (−7°C/min) cooling of guinea-pig isolated trachea produced a rapidly developing, transient contraction followed by relaxation. Cooling-induced contraction was dependent on temperature (30, 20 or 10°C) and responses in trachea obtained from actively sensitized guinea pigs were significantly greater (20 and 10°-C) than those observed in normal trachea. Cooling to 20°C was selected for subsequent experiments. Pre-treatment with sufficient concentrations of atropine, clemastine, cromoglycate, indomethacin, or nordihydroguaiaretic acid did not depress contraction to cooling in either normal or sensitized trachea. This indicates a direct effect of cooling. The contraction. produced by coolin…
Additional file 1: of Non-neuronal cholinergic system contributes to corticosteroid resistance in chronic obstructive pulmonary disease patients
Supplementary Table S1. Maximal percentage of inhibition of IL-8, MMP-9, CCL-5, GM-CSF and IL-1β release from neutrophils of healthy subjects and COPD patients. Neutrophils were incubated with aclidinium (Acl; 0.1nM-1 μM), fluticasone (Flut; 0.1nM-1 μM), formoterol (Form; 0.01nM-100nM) or salmeterol (Salm; 0.1nM-1 μM) in response to LPS (1 μg/ml) or cigarette smoke extract (CSE 5 %). The levels of different cytokines in the cell supernatant were determined and the maximal percent of inhibitions were calculated. Values are mean ± SD of 3 independent experiments run in triplicate. *p
Effects of SCA40 on human isolated bronchus and human polymorphonuclear leukocytes: comparison with rolipram, SKF94120 and levcromakalim
1. SCA40 (0.1 nM-0.1 mM) produced concentration-dependent suppression of the spontaneous tone of human isolated bronchus (-log EC50 = 6.85 +/- 0.09; n = 10) and reached a maximal relaxation similar to that of theophylline (3 mM). The potency (-log EC50 values) of SCA40 compared to other relaxants was rolipram (7.44 +/- 0.12; n = 9) > SCA40 > or = levcromakalim (6.49 +/- 0.04; n = 6) > SKF94120 (5.87 +/- 0.10; n = 9). 2. When tested against the activity of the isoenzymes of cyclic nucleotide phosphodiesterase (PDE) isolated from human bronchus, SCA40 proved highly potent against PDE III (-log IC50 = 6.47 +/- 0.16; n = 4). It was markedly less potent against PDE IV (4.82 +/- 0.18; n = 4) and …
Additional file 2: of Non-neuronal cholinergic system contributes to corticosteroid resistance in chronic obstructive pulmonary disease patients
Supplementary Figure E1. Effects of formoterol and salmeterol on pro-inflammatory markers. Concentration-dependent inhibition of lipopolysaccharide (LPS)-induced cytokines or MMP-9 release by formoterol (Form) and salmeterol (Salm) from peripheral blood neutrophils of healthy subjects and COPD patients. Neutrophils were preincubated with Salm (0.1nM-1 μM) or form (0.01nM-100nM) for 1 h followed by cell stimulation with LPS (1 μg/ml) for 6 h. Results are expressed as means ± SD of n = 3 (3 cell healthy and 3 cell COPD populations run in triplicate) independent experiments. Two-way ANOVA was followed by the post hoc Bonferroni test. *p