Release of IL-1ß and IL-18 in human primary bronchial epithelial cells exposed to cigarette smoke is independent of NLRP3.

Cigarette smoke (CS) is a major risk factor for chronic lung diseases and promotes activation of pattern recognition receptors in the bronchial epithelium. NOD-like receptor family, pyrin domain-containing 3 (NLRP3) is a pattern recognition receptor whose activation leads to caspase-1 cleavage, maturation/release of IL-1ß and IL-18, and eventually pyroptosis. Whether the NLRP3 inflammasome participates in CS-induced inflammation in bronchial epithelial cells is still unclear. Herein, we evaluated the involvement of NLRP3 in CS-induced inflammatory responses in human primary bronchial epithelial cells. To this purpose, human primary bronchial epithelial cells were stimulated with CS extracts (CSE) and lytic cell death, caspase activation (-1, -8, -3/7), cytokine release (IL-1ß, IL-18, and IL-8), NLRP3, pro-IL-1ß/pro-IL-18 mRNA, and protein expression were measured. The impact of inhibitors of NLRP3 (MCC950), caspases, and the effect of the antioxidant N-acetyl cysteine were evaluated. We found that CSE increased pro-IL-1ß expression and induced activation of caspase-1 and release of IL-1ß and IL-18. These events were independent of NLRP3 and we found that NLRP3 was not expressed. N-acetyl cysteine reverted CSE-induced caspase-1 activation. Overall, our findings support that the bronchial epithelium may play a central role in the release of IL-1 family cytokines independently of NLRP3 in the lungs of smokers.

Stem cells, Notch-1 signaling and Oxidative stress: a hellish trio in cancer development and progression within the airways. Is there a role for natural compounds?

Notch-1 signaling plays a crucial role in stem cell maintenance and in repair mechanisms in various mucosal surfaces, including airway mucosa. Persistent injury can induce an aberrant activation of Notch-1 signaling in stem cells leading to an increased risk of cancer initiation and progression. Chronic inflammatory respiratory disorders, including Chronic Obstructive Pulmonary Disease (COPD) is associated to both over-activation of Notch-1 signaling and increased lung cancer risk. Increased oxidative stress, also due to cigarette smoke, can further contribute to promote cancer initiation and progression by amplifying inflammatory responses, by activating the Notch-1 signaling and by blocking regulatory mechanisms that inhibit the growth capacity of stem cells. This review offers a comprehensive overview on the effects of aberrant Notch-1 signaling activation in stem cells and of increased oxidative stress in the lung cancer. The putative role of natural compounds with anti-oxidant properties is also described.

Role of airway epithelium in viral respiratory infections: Can carbocysteine prevent or mitigate them?

Alterations in airway epithelial homeostasis increase viral respiratory infections risk. Viral infections frequently are associated with chronic obstructive pulmonary disease (COPD) exacerbations, events that dramatically promote disease progression. Mechanism promoting the main respiratory viruses entry and virus-evocated innate and adaptive immune responses have now been elucidated, and an oxidative stress central role in these pathogenic processes has been recognized. Presence of reactive oxygen species in macrophages and other cells allows them to eliminate virus, but its excess alters the balance between innate and adaptive immune responses and proteases/anti-proteases and leads to uncontrolled inflammation, tissue damage, and hypercoagulability. Different upper and lower airway cell types also play a role in viral entry and infection. Carbocysteine is a muco-active drug with anti-oxidant and anti-inflammatory properties used for the management of several chronic respiratory diseases. Although the use of anti-oxidants has been proposed as an effective strategy in COPD exacerbations management, the molecular mechanisms that explain carbocysteine efficacy have not yet been fully clarified. The present review describes the most relevant features of the common respiratory virus pathophysiology with a focus on epithelial cells and oxidative stress role and reports data supporting a putative role of carbocysteine in viral respiratory infections.

Formoterol Exerts Anti-Cancer Effects Modulating Oxidative Stress and Epithelial-Mesenchymal Transition Processes in Cigarette Smoke Extract Exposed Lung Adenocarcinoma Cells.

Lung cancer frequently affects patients with Chronic Obstructive Pulmonary Disease (COPD). Cigarette smoke (CS) fosters cancer progression by increasing oxidative stress and by modulating epithelial-mesenchymal transition (EMT) processes in cancer cells. Formoterol (FO), a long-acting ß2-agonist widely used for the treatment of COPD, exerts antioxidant activities. This study explored in a lung adenocarcinoma cell line (A549) whether FO counteracted the effects of cigarette smoke extract (CSE) relative to oxidative stress, inflammation, EMT processes, and cell migration and proliferation. A549 was stimulated with CSE and FO, ROS were evaluated by flow-cytometry and by nanostructured electrochemical sensor, EMT markers were evaluated by flow-cytometry and Real-Time PCR, IL-8 was evaluated by ELISA, cell migration was assessed by scratch and phalloidin test, and cell proliferation was assessed by clonogenic assay. CSE significantly increased the production of ROS, IL-8 release, cell migration and proliferation, and SNAIL1 expression but significantly decreased E-cadherin expression. FO reverted all these phenomena in CSE-stimulated A549 cells. The present study provides intriguing evidence that FO may exert anti-cancer effects by reverting oxidative stress, inflammation, and EMT markers induced by CS. These findings must be validated in future clinical studies to support FO as a valuable add-on treatment for lung cancer management.

Cigarette smoke inhibits the NLRP3 inflammasome and leads to caspase-1 activation via the TLR4-TRIF-caspase-8 axis in human macrophages.

The NLRP3 inflammasome is formed by the sensor NLRP3, the adaptor ASC, and pro-caspase-1. Assembly and activation of the inflammasome trigger caspase-1-dependent cleavage of pro-IL-1ß and pro-IL-18 into their secreted forms. Cigarette smoke is a risk factor for chronic inflammatory diseases and is associated with macrophage dysfunction. The impact of cigarette smoke on NLRP3-dependent responses in macrophages is largely unknown. Herein, we investigated the effects of cigarette smoke extract (CSE) on the NLRP3 inflammasome in human monocyte-derived macrophages (MDMs) and THP-1 cells stimulated with lipopolysaccharide (LPS) and LPS plus the NLRP3 inflammasome activator ATP. We found that CSE inhibited the release of IL-1ß and IL-18 as well as the expression of NLRP3 acting mainly at the transcriptional level. Interestingly, we found that CSE increased the caspase-1 activity via an NLRP3-independent and TLR4-TRIF-caspase-8-dependent pathway. Activation of caspase-1 by CSE led to a reduction of the basal glycolytic flux and impaired glycolytic burst in response to LPS. Overall, our findings unveil novel pathways leading to immune-metabolic alterations in human macrophages exposed to cigarette smoke. These mechanisms may contribute to macrophage dysfunction and increased risk of infection in smokers.

MiR-21 upregulation increases IL-8 expression and tumorigenesis program in airway epithelial cells exposed to cigarette smoke.

BACKGROUND: Cigarette smoke exposure, increasing Toll-like receptor 4 (TLR4) and reactive oxygen species (ROS), promotes inflammatory responses in airway epithelial cells. Chronic inflammation, microRNA (miRNA), and oxidative stress are associated with cancer development. AIMS: The present study was aimed to explore whether cigarette smoke exposure, altering miR-21 expression, promoted inflammatory responses and tumorigenesis processes in airway epithelial cells. METHODS: Airway normal and cancer epithelial cells (16HBE and A549) were exposed to cigarette smoke extracts (CSE) or with/without agomiR-21, and then it was assessed: a) miR-21 expression; b) signal transducer and activator of transcription 3 (STAT3) nuclear protein expression and ERK1/2 activation; c) IL-8 gene expression and protein release. An antagonist of TLR4 (CLI-095) and the antioxidant flavonoid, apigenin, were also included to evaluate miR-21 expression in CSE exposed cells. RESULTS: It was demonstrated that: a) A549 cells constitutively expressed higher levels of miR-21 and IL-8; b) CSE increased STAT3 nuclear expression in 16HBE; c) in both cell lines, CSE and agomiR-21 increased: miR-21 expression; ERK1/2 activation and IL-8 gene expression and protein release; d) TLR4 inhibition counteracted the effects of CSE on miR-21 in A549; e) apigenin reduced miR-21 and IL-8 gene expression in both cell lines. CONCLUSIONS: Data herein provided identified for the first time new mechanisms supporting the crucial role of cigarette smoke-induced miR-21 expression in the amplification of inflammatory responses and in tumorigenesis processes within the airways.

SIRT1/FoxO3 axis alteration leads to aberrant immune responses in bronchial epithelial cells.

Inflammation and ageing are intertwined in chronic obstructive pulmonary disease (COPD). The histone deacetylase SIRT1 and the related activation of FoxO3 protect from ageing and regulate inflammation. The role of SIRT1/FoxO3 in COPD is largely unknown. This study evaluated whether cigarette smoke, by modulating the SIRT1/FoxO3 axis, affects airway epithelial pro-inflammatory responses. Human bronchial epithelial cells (16HBE) and primary bronchial epithelial cells (PBECs) from COPD patients and controls were treated with/without cigarette smoke extract (CSE), Sirtinol or FoxO3 siRNA. SIRT1, FoxO3 and NF-κB nuclear accumulation, SIRT1 deacetylase activity, IL-8 and CCL20 expression/release and the release of 12 cytokines, neutrophil and lymphocyte chemotaxis were assessed. In PBECs, the constitutive FoxO3 expression was lower in patients with COPD than in controls. Furthermore, CSE reduced FoxO3 expression only in PBECs from controls. In 16HBE, CSE decreased SIRT1 activity and nuclear expression, enhanced NF-κB binding to the IL-8 gene promoter thus increasing IL-8 expression, decreased CCL20 expression, increased the neutrophil chemotaxis and decreased lymphocyte chemotaxis. Similarly, SIRT1 inhibition reduced FoxO3 expression and increased nuclear NF-κB. FoxO3 siRNA treatment increased IL-8 and decreased CCL20 expression in 16HBE. In conclusion, CSE impairs the function of SIRT1/FoxO3 axis in bronchial epithelium, dysregulating NF-κB activity and inducing pro-inflammatory responses.

Effects of Carbocysteine and Beclomethasone on Histone Acetylation/Deacetylation Processes in Cigarette Smoke Exposed Bronchial Epithelial Cells.

Histone deacetylase expression/activity may control inflammation, cell senescence, and responses to corticosteroids. Cigarette smoke exposure, increasing oxidative stress, may negatively affect deacetylase expression/activity. The effects of cigarette smoke extracts (CSE), carbocysteine, and beclomethasone dipropionate on chromatin remodeling processes in human bronchial epithelial cells are largely unknown. The present study was aimed to assess the effects of cigarette smoke, carbocysteine, and beclomethasone dipropionate on histone deacetylase 3 (HDAC3) expression/activity, N-CoR (nuclear receptor corepressor) expression, histone acetyltransferases (HAT) (p300/CBP) expression, p-CREB and IL-1 m-RNA expression, neutrophil chemotaxis. Increased p-CREB expression was observed in the bronchial epithelium of smokers. CSE increased p-CREB expression and decreased HDAC3 expression and activity and N-CoR m-RNA and protein expression. At the same time, CSE increased the expression of the HAT, p300/CBP. All these events increased acetylation processes within the cells and were associated to increased IL-1 m-RNA expression and neutrophil chemotaxis. The incubation of CSE exposed cells with carbocysteine and beclomethasone counteracted the effects of cigarette smoke on HDAC3 and N-CoR but not on p300/CBP. The increased deacetylation processes due to carbocysteine and beclomethasone dipropionate incubation is associated to reduced p-CREB, IL-1 m-RNA expression, neutrophil chemotaxis. These findings suggest a new role of combination therapy with carbocysteine and beclomethasone dipropionate in restoring deacetylation processes compromised by cigarette smoke exposure. J. Cell. Physiol. 232: 2851-2859, 2017. © 2016 Wiley Periodicals, Inc.

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 analysis was applied to confirm the expression of significantly modulated proteins. Flow cytometry and immunofluorescence were used to assess F-actin polimerization by phalloidin method. Fourteen proteins, with significant (p < 0.05) changes in intensity, were identified at various experimental points: 6 were up-regulated and 8 were down-regulated. As expected, bioinformatic analysis revealed that most of these proteins are involved in anti-oxidant and immune responses and in cytoskeleton stability. Western blot analysis confirmed that: Proteasome activator complex subunit 2 (PSME2), Peroxiredoxin-6 (PRDX6), Annexin A5 (ANXA5) and Heat shock protein beta-1 (HSPB1) were reduced and Coactosin-like protein (COTL-1) was increased by co-exposure of CSE and LPS. Furthermore, LPS and CSE increased actin polimerization. In conclusion, although further validation studies are needed, our findings suggest that, CSE and LPS could contribute to the progressive deterioration of lung function, altering the expression of proteins involved in metabolic processes and cytoskeleton rearrangement in bronchial epithelial cells.

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

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 expression of heme oxygenase 1 and more than doubling glutathione levels. This resulted in suppression of CSE-induced ROS generation in macrophages. In macrophages, 17-oxo-DHA potently suppressed TNFα release in response to LPS, CSE and IL-1ß acting at transcriptional level via a mechanism independent of Nrf2. Externally supplemented 17-oxo-DHA displayed the same effects in the presence of the Cox-inhibitor indomethacin. The non-electrophilic 17-oxo-DHA precursor DHA did not show any biological actions, indicating that the electrophilic moiety was required for this compound to become bioactive. CONCLUSIONS: 17-Oxo-DHA promotes cytoprotective actions both in immune and structural cells. In immune cells, 17-oxo-DHA is effective in contrasting CSE- and LPS-induced oxidative damage and inflammation acting via multiple independent pathways. GENERAL SIGNIFICANCE: Herein we provide insights on how the novel endogenous electrophilic DHA-derivative 17-oxo-DHA promotes anti-oxidant and anti-inflammatory actions. Data herein reported indicate that 17-oxo-DHA is an attractive lead compound for the development of new treatments for cigarette smoke-related airway inflammatory disorders.

MD2 expression is reduced in large airways of smokers and COPD smokers.

Toll-like receptor 4 (TLR4) signaling requires a number of accessory proteins to initiate a signal. MD-2 is one of the accessory proteins with a relevant role in lipopolysaccharide responses. Although cigarette smoke increases TLR4 expression, TLR4 signaling is altered in smokers and in smokers COPD patients. The main aims of this study were to explore whether MD2 is altered in large and small airways of COPD and of smokers without COPD. The expression of MD2 ex vivo was assessed by immunohistochemistry in surgical specimens from current smokers COPD (s-COPD; n = 14), smokers without COPD (S; n = 7), and from non-smoker non-COPD subjects (C; n = 11. The in vitro effects of cigarette smoke extracts on the MD2 expression in a human bronchial epithelial cell line (16-HBE) were also assessed by flow cytometry. MD2 is reduced in the epithelium and in the submucosa in large airways but not in the epithelium and in the submucosa in small airways of smokers and of s-COPD. The expression of MD2 in the submucosa of the large airways is significantly higher in comparison to the submucosa of the small airways in all the studied groups. In vitro, cigarette smoke is able to increase TLR4 but it reduces MD2 in a dose-dependent manner in bronchial epithelial cells. Cigarette smoke may alter innate immune responses reducing the expression of the MD2, a molecule with an important role in TLR4 signaling.

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 GSH levels improving cell protection against oxidative stress. CONCLUSIONS: The present study shows that NLC may be considered a promising strategy to improve corticosteroid mediated effects in cellular models associated to corticosteroid resistance. The NLC containing FP can be considered good systems for dosage forms useful for increasing the effectiveness of fluticasone decreasing its side effects.

Gold nanowires-based sensor for quantification of H2O2 released by human airway epithelial cells.

Hydrogen peroxide (H2O2) is a biomarker relevant for oxidative stress monitoring. Most chronic airway diseases are characterized by increased oxidative stress. To date, the main methods for the detection of this analyte are expensive and time-consuming laboratory techniques such as fluorometric and colorimetric assays. There is a growing interest in the development of electrochemical sensors for H2O2 detection due to their low cost, ease of use, sensitivity and rapid response. In this work, an electrochemical sensor based on gold nanowire arrays has been developed. Thanks to the catalytic activity of gold against hydrogen peroxide reduction and the high surface area of nanowires, this sensor allows the quantification of this analyte in a fast, efficient and selective way. The sensor was obtained by template electrodeposition and consists of gold nanowires about 5 µm high and with an average diameter of about 200 nm. The high active surface area of this electrode, about 7 times larger than a planar gold electrode, ensured a high sensitivity of the sensor (0.98 µA µM-1cm-2). The sensor allows the quantification of hydrogen peroxide in the range from 10 µM to 10 mM with a limit of detection of 3.2 µM. The sensor has excellent properties in terms of reproducibility, repeatability and selectivity. The sensor was validated by quantifying the hydrogen peroxide released by human airways A549 cells exposed or not to the pro-oxidant compound rotenone. The obtained results were validated by comparing them with those obtained by flow cytometry after staining the cells with the fluorescent superoxide-sensitive Mitosox Red probe giving a very good concordance.

Tyndallized bacteria prime bronchial epithelial cells to mount an effective innate immune response against infections.

Airway epithelium represents a physical barrier against toxic substances and pathogens but also presents pattern recognition receptors on the epithelial cells that detect pathogens leading to molecule release and sending signals that activate both the innate and adaptive immune responses. Thus, impaired airway epithelial function and poor integrity may increase the recurrence of infections. Probiotic use in respiratory diseases as adjuvant of traditional therapy is increasingly widespread. There is growing interest in the use of non-viable heat-killed bacteria, such as tyndallized bacteria (TB), due to safety concerns and to their immunomodulatory properties. This study explores in vitro the effects of a TB blend on the immune activation of airway epithelium. 16HBE bronchial epithelial cells were exposed to different concentrations of TB. Cell viability, TB internalization, TLR2 expression, IL-6, IL-8 and TGF-ßl expression/release, E-cadherin expression and wound healing were assessed. We found that TB were tolerated, internalized, increased TLR2, E-cadherin expression, IL-6 release and wound healing but decreased both IL-8 and TGF-ßl release. In conclusion, TB activate TLR2 pathway without inducing a relevant pro-inflammatory response and improve barrier function, leading to the concept that TB preserve epithelial homeostasis and could be used as strategy to prevent and to manage respiratory infection, exacerbations included.

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

Leukotriene B(4) (LTB(4)) 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 LTB(4) 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 intercellular adhesion molecule 1 (ICAM-1) and on the binding of signal transducer and activator of transcription 1 (STAT-1) to ICAM-1 promoter as well as the adhesiveness of neutrophils to bronchial epithelial cells. CSE and mini-bronchoalveolar lavages from smokers increased BLT2 and ICAM-1 expression as well as the adhesiveness of neutrophils to bronchial epithelial cells and decreased PPAR-α expression. CSE induced the activation of STAT-1 and its binding to ICAM-1 promoter. These findings suggest that, in bronchial epithelial cells, CSE promote a prevalent induction of pro-inflammatory BLT2 receptors and activate mechanisms leading to increased neutrophil adhesion, a mechanism that contributes to airway neutrophilia and to tissue damage.

Effects of condensates from volcanic fumaroles and cigarette smoke extracts on airway epithelial cells.

The impact of volcanic airborne products on airway epithelium homeostasis is largely unknown. This study assessed the effects of volcanic Fumarole Condensates (FC) alone or combined with Cigarette Smoke Extracts (CSE) on airway epithelial cells (16HBE and A549). Chemical composition of FC was analyzed by gas chromatography and HPLC. Cells were exposed to FC and IL-33 and IL-8 were assessed. The effects of FC and CSE on cell injury were evaluated assessing cell metabolism/cell viability, mitochondrial stress, cell apoptosis/cell necrosis, and cell proliferation. FC contained: water vapor (70-97%), CO2 (3-30%), acid gases (H2S, SO2, HCl, HF) around 1%. FC increased the intracellular IL-33 but differently modulated IL-33 and IL-8 gene expression and IL-8 release in the tested cell lines. FC without/with CSE: (a) increased cell metabolism/cell viability in 16HBE, while decreased it in A549; (b) increased mitochondrial stress in both cell types. FC with CSE increased cell necrosis in A549 in comparison to CSE alone. CSE reduced cell proliferation in 16HB,E while increased it in A549 and FC counteracted these effects in both cell types. Overall, FC induce a pro-inflammatory profile associated to a metabolic reprogramming without a relevant toxicity also in presence of CSE in airway epithelial cells.

Tyndallized Bacteria Preferentially Induce Human Macrophage M1 Polarization: An Effect Useful to Balance Allergic Immune Responses and to Control Infections.

Macrophage polarization is a dynamic process through which macrophages acquire specific features whose extremes are represented by M1 and M2 polarization. Interleukin (IL)-6, IL-1ß, IL-12 and IL-8 belong to M1 macrophages while transforming growth factor-beta (TGF-ß belongs to M2 cytokines. M2 polarization prevalence is observed in allergic diseases. Tyndallization is a thermal process able to inactivate microorganisms and to allow their use for chronic respiratory disease treatment via immune response modulation. The present study explores the effects of a blend of tyndallized bacteria (TB) on macrophage polarization. THP-1-derived macrophages were exposed to different concentrations of TB (106, 5 × 106, 107, 5 × 107, 108 CFU/mL) and then cell viability and TB phagocytosis, and IL-8, IL-1ß, IL-6, IL-12 and TGF-ß1 gene expression and release were assessed. TB were tolerated, phagocyted and able to increase IL-8, IL-1ß and IL-6 gene expression and release IL-12 gene expression, as well as decrease TGF-ß1 gene expression and release. The effects on IL-8, IL-6 and TGF-ß1 release were confirmed in human monocyte-derived macrophages (hMDMs) exposed to TB. In conclusion, TB promote M1 polarization, and this mechanism might have valuable potential in controlling allergic diseases and infections, possibly preventing disease exacerbations.

Carbocysteine Modifies Circulating miR-21, IL-8, sRAGE, and fAGEs Levels in Mild Acute Exacerbated COPD Patients: A Pilot Study.

Patients with Chronic Obstructive Pulmonary Disease (COPD) periodically experience acute exacerbation (AECOPD). Carbocysteine represents a valid add on therapy in COPD by exerting antioxidant and anti-inflammatory activities. The in vivo effects of carbocysteine on inflammatory markers are not yet fully understood. The aims of this study were to assess: (i) miR-21, IL-8, soluble Receptor for Advanced Glycation End Products (sRAGE), and fluorescent Advanced Glycation End Products (fAGEs) in control subjects (n = 9), stable (n = 9), and AECOPD patients (n = 24); and (ii) whether carbocysteine modifies these markers and the functional parameters in mild AECOPD patients. Mild AECOPD patients received or not carbocysteine along with background inhalation therapy for 20 days. At the onset and at the end of the observation period, the following parameters were evaluated: FEV1, FEF25-75%, CAT questionnaire; miR-21 by Real Time PCR; IL-8 and sRAGE by ELISA; and fAGEs by spectro-fluorescence method. COPD patients showed higher levels of miR-21, IL-8, fAGEs and lower levels of sRAGE compared to that of controls. miR-21 inversely correlated with FEV1. IL-8 and fAGEs were significantly different in stable and exacerbated COPD patients. Carbocysteine improved symptoms, FEV1 and FEF25-75%, increased sRAGE, and reduced miR-21, IL-8, and fAGEs in mild AECOPD patients. The present study provides compelling evidence that carbocysteine may help to manage mild AECOPD by downregulating some parameters of systemic inflammation.