Chronic Cigarette Smoke Exposure Research Articles

Long-term persistent exposure to cigarette smoke induces AhR driven corneal endothelial dysfunction in mice

Epidemiological studies show cigarette smoking enhances corneal endothelial dysfunction, but mechanisms remain unclear. Our study reveals that prolonged smoke exposure activates the aryl hydrocarbon receptor (AhR), increasing CYP1B1 expression and accelerating senescence and fibrosis in corneal endothelium, potentially reflecting adaptive responses to maintain corneal resilience. Although these molecular modifications indicate early endothelial dysfunction, no pathological changes were observed. The findings indicate that while chronic cigarette smoke exposure triggers initial molecular alterations and endothelial dysfunction, the progression to Fuchs endothelial corneal dystrophy likely requires additional environmental or genetic factors beyond smoke exposure alone.

Chronic cigarette smoke exposure masks pathological features of Helicobacter pylori infection while promoting tumor initiation.

Gastric cancer is the fifth most common cancer and the fifth leading cause of cancer deaths worldwide. Chronic infection by the bacterium Helicobacter pylori is the most prominent gastric cancer risk factor, but only 1-3% of infected individuals will develop gastric cancer. Cigarette smoking is another independent gastric cancer risk factor, and H. pylori- infected smokers are at a 2-11-fold increased risk of gastric cancer development, but the direct impacts of cigarette smoke on H. pylori pathogenesis remain unknown. In this study, male C57BL/6 mice were infected with H. pylori and began smoking within one week of infection. The mice were exposed to cigarette smoke (CS) five days/week for 8 weeks. CS exposure had no notable impact on gross gastric morphology or inflammatory status compared to filtered-air (FA) exposed controls in mock-infected mice. However, CS exposure significantly blunted H. pylori- induced gastric inflammatory responses, reducing gastric atrophy and pyloric metaplasia development. Despite blunting these classic pathological features of H. pylori infection, CS exposures increased DNA damage within the gastric epithelial cells and accelerated H. pylori- induced dysplasia onset in the INS-GAS gastric cancer model. These data suggest that cigarette smoking may clinically silence classic clinical symptoms of H. pylori infection but enhance the accumulation of mutations and accelerate gastric cancer initiation.

Cortex Mori Radicis[Morus Alba L. (Moraceae)] Extract Alleviates Senescence via PI3K/Akt Signaling in COPD Fibroblasts

BackgroundChronic obstructive pulmonary disease (COPD) is marked by prolonged exposure to cigarette smoke, which accelerates senescence in lung fibroblasts and contributes to lung fibrosis. Cortex Mori Radicis [Morus alba L. (Moraceae)], a traditional Chinese medicinal herb known for its antitussive properties, has emerged as a potential therapeutic agent for COPD. This study aims to elucidate the immunological mechanisms by which Cortex Mori Radicis mitigates COPD progression, utilizing a mouse model and the MRC-5 cell line. Methods and ResultsCOPD mouse models were established through chronic cigarette smoke (CS) exposure, followed by isolation of lung fibroblasts. Senescence markers and inflammatory mediators were assessed in both the isolated cells and the mice. Lung fibroblasts and bleomycin-treated MRC-5 cells exhibited elevated expression of senescence markers, including senescence-associated beta-galactosidase activity, p16INK4A, p21, p38 MAPK, and p53, along with increased levels of senescence-associated secretory phenotype (SASP) mRNA, such as IL-6 and IL-8. Treatment with Cortex Mori Radicis significantly attenuated the protein levels of these senescence markers and reduced SASP mRNA expression. Furthermore, integration of transcriptomic data from lung tissues and primary fibroblasts, combined with network pharmacology analysis, indicated that Cortex Mori Radicis inhibits fibroblast senescence via the PI3K/Akt pathway, thereby ameliorating lung pathology in COPD mice. ConclusionThrough the application of transcriptomics and network analysis, this study identifies that Cortex Mori Radicis suppresses cigarette smoke-induced senescence in pulmonary tissues and bleomycin-exposed MRC-5 cells by targeting the PI3K/Akt signaling pathway. These findings underscore the therapeutic potential of Cortex Mori Radicis as a novel intervention for COPD.

Endothelial knockdown of the tumor suppressor, WWOX, increases inflammation in ventilator-induced lung injury.

Chronic cigarette smoke exposure decreases lung expression of WWOX which is known to protect the endothelial barrier during infectious models of acute respiratory distress syndrome (ARDS). Proteomic analysis of WWOX-silenced endothelial cells (ECs) was done using tandem mass tag mass spectrometry (TMT-MS). WWOX-silenced ECs as well as those isolated from endothelial cell Wwox knockout (EC Wwox KO) mice were subjected to cyclic stretch (18% elongation, 0.5 Hz, 4 h). Cellular lysates and media supernatant were harvested for assays of cellular signaling, protein expression, and cytokine release. These were repeated with dual silencing of WWOX and zyxin. Control and EC Wwox KO mice were subjected to high tidal volume ventilation. Bronchoalveolar lavage fluid and mouse lung tissue were harvested for cellular signaling, cytokine secretion, and histological assays. TMT-MS revealed upregulation of zyxin expression during WWOX knockdown which predicted a heightened inflammatory response to mechanical stretch. WWOX-silenced ECs and ECs isolated from EC Wwox mice displayed significantly increased cyclic stretch-mediated secretion of various cytokines (IL-6, KC/IL-8, IL-1β, and MCP-1) relative to controls. This was associated with increased ERK and JNK phosphorylation but decreased p38 mitogen-activated kinases (MAPK) phosphorylation. EC Wwox KO mice subjected to VILI sustained a greater degree of injury than corresponding controls. Silencing of zyxin during WWOX knockdown abrogated stretch-induced increases in IL-8 secretion but not in IL-6. Loss of WWOX function in ECs is associated with a heightened inflammatory response during mechanical stretch that is associated with increased MAPK phosphorylation and appears, in part, to be dependent on the upregulation of zyxin.NEW & NOTEWORTHY Prior tobacco smoke exposure is associated with an increased risk of acute respiratory distress syndrome (ARDS) during critical illness. Our laboratory is investigating one of the gene expression changes that occurs in the lung following smoke exposure: WWOX downregulation. Here we describe changes in protein expression associated with WWOX knockdown and its influence on ventilator-induced ARDS in a mouse model.

Abstract 17706: Role of p62/SQSTM1 on Acute Ischemia-Induced Arrhythmias After Chronic Smoke in Mice

Introduction: Cigarette smoke presents a high-risk factor for coronary heart disease development. Cigarette smoke is often linked with increased susceptibility to cardiac ischemia and arrhythmias. p62/SQSTM1 is a stress-sensing protein responsible for proteasomal degradation that maintains redox homeostasis during cardiac tissue injury. However, its exact mechanism for arrhythmia development is not clear. Hypothesis: Here we tested the biological role of p62/SQSTM1 mice on cardiac ischemia injury-induced arrhythmogenesis after chronic cigarette smoke exposure. Methods: Mice (global p62/SQSTM1 deletion mice; p62/SQSTM1 -/- , 4 months old age) were exposed to cigarette smoke (9.4 mg tar/0.726 mg nicotine) for 16 weeks. Afterward, we utilized ischemia-reperfusion (IR) injury in intact hearts to test the risk of arrhythmias. Results: The smoke-exposed p62/SQSTM1 -/- isolated hearts (8 months old age) significantly reduced the incidence of ventricular arrhythmias compared to hearts obtained from p62/SQSTM1 +/+ mice post-IR during the first 15 minutes reperfusion (% of mice with ventricular arrhythmias, p62/SQSTM1 +/+ ; 7/10 hearts, p62/SQSTM1 -/- ; 2/8 hearts*, * <0.05). Abnormal Ca 2+ regulation in cardiomyocytes is triggering diastolic delayed afterdepolarizations and often leads to ventricular tachycardia. Therefore, we tested cellular Ca 2+ regulation with acute cigarette smoke extract (CSE) exposures in cardiomyocytes. p62/SQSTM1 -/- cardiomyocytes for 30 minutes exposed to CSE significantly alleviated CSE-induced spontaneous Ca 2+ waves (% of cardiomyocytes with spontaneous Ca 2+ waves, p62/SQSTM1 +/+ ; 6/13 cells, p62SQSTM1 -/- ; 1/15 cells*, * <0.05). Conclusions: Overall, our results suggest that the deletion of p62/SQSTM1 protein has a protective effect on acute cardiac ischemia-induced arrhythmogenesis after chronic cigarette smoke in mice.

Cigarette smoke-induced injury induces distinct sex-specific transcriptional signatures in mice tracheal epithelial cells.

The airway epithelial barrier is crucial for defending against respiratory insults and diseases. Disruption of epithelial integrity contributes to respiratory diseases, and sex-specific differences in susceptibility and severity have been observed. However, sex-specific differences in the context of respiratory diseases are often overlooked, especially in murine models. In this study, we investigated the in vitro transcriptomics of male and female murine tracheal epithelial cells (mTECs) in response to chronic cigarette smoke (CS) exposure using an International Organization for Standardization (ISO) puff regimen. Our findings reveal sex-specific differences in the baseline characteristics of airway epithelial cells. Female mTECs demonstrated stronger barrier function and higher ciliary function compared with males. The barrier function was disrupted in both males and females following chronic CS, but the difference was more significant in females due to their higher baseline. Female mice exhibited transcriptional signatures suggesting dedifferentiation with increased basal cells and markers of cellular senescence. Pathway analysis indicated potential protective roles of planar cell polarity (PCP) in preventing dedifferentiation in male mice exposed to CS. We also observed sex-specific differences in the DNA damage response and antioxidant levels, suggesting distinct mechanisms underlying cellular stress. Understanding these sex-specific mechanisms could facilitate the development of targeted therapeutic strategies for lung diseases associated with environmental insults. Recognizing sex-based differences in disease susceptibility and treatment response can lead to personalized care and improved outcomes. Clinical trials should consider sex as a biological variable to develop effective interventions that address the unique differences between men and women in respiratory diseases.NEW & NOTEWORTHY The study underscores the importance of considering sex-specific differences in the airway epithelium in respiratory diseases such as COPD. Differences in gene expression between males and females at baseline and in response to chronic injury in the airway epithelium could have implications on disease susceptibility, both in COPD and other respiratory diseases. Therefore, understanding these differences is crucial for developing targeted therapies to treat respiratory diseases based on a sex-specific manner.

Caveolin-1-derived peptide attenuates cigarette smoke-induced airway and alveolar epithelial injury.

Chronic obstructive pulmonary disease (COPD) is a debilitating lung disease with no effective treatment that can reduce mortality or slow the disease progression. COPD is the third leading cause of global death and is characterized by airflow limitations due to chronic bronchitis and alveolar damage/emphysema. Chronic cigarette smoke (CS) exposure damages airway and alveolar epithelium and remains a major risk factor for the pathogenesis of COPD. We found that the expression of caveolin-1, a tumor suppressor protein; p53; and plasminogen activator inhibitor-1 (PAI-1), one of the downstream targets of p53, was markedly increased in airway epithelial cells (AECs) as well as in type II alveolar epithelial (AT2) cells from the lungs of patients with COPD or wild-type mice with CS-induced lung injury (CS-LI). Moreover, p53- and PAI-1-deficient mice resisted CS-LI. Furthermore, treatment of AECs, AT2 cells, or lung tissue slices from patients with COPD or mice with CS-LI with a seven amino acid caveolin-1 scaffolding domain peptide (CSP7) reduced mucus hypersecretion in AECs and improved AT2 cell viability. Notably, induction of PAI-1 expression via increased caveolin-1 and p53 contributed to mucous cell metaplasia and mucus hypersecretion in AECs, and reduced AT2 viability, due to increased senescence and apoptosis, which was abrogated by CSP7. In addition, treatment of wild-type mice having CS-LI with CSP7 by intraperitoneal injection or nebulization via airways attenuated mucus hypersecretion, alveolar injury, and significantly improved lung function. This study validates the potential therapeutic role of CSP7 for treating CS-LI and COPD. NEW & NOTEWORTHY Chronic cigarette smoke (CS) exposure remains a major risk factor for the pathogenesis of COPD, a debilitating disease with no effective treatment. Increased caveolin-1 mediated induction of p53 and downstream plasminogen activator inhibitor-1 (PAI-1) expression contributes to CS-induced airway mucus hypersecretion and alveolar wall damage. This is reversed by caveolin-1 scaffolding domain peptide (CSP7) in preclinical models, suggesting the therapeutic potential of CSP7 for treating CS-induced lung injury (CS-LI) and COPD.

Evaluating Novel Protein Phosphatase 2A Activators as Therapeutics for Emphysema.

The activity of PP2A (protein phosphatase 2A), a serine-threonine phosphatase, is reduced by chronic cigarette smoke (SM) exposure and α-1 antitrypsin (AAT) deficiency, and chemical activation of PP2A reduces the loss of lung function in SM-exposed mice. However, the previously studied PP2A-activator tricyclic sulfonamide compound DBK-1154 has low stability to oxidative metabolism, resulting in fast clearance and low systemic exposure. Here we compare the utility of a new more stable PP2A activator, ATUX-792, versus DBK-1154 for the treatment of SM-induced emphysema. ATUX-792 was also tested in human bronchial epithelial cells and a mouse model of AAT deficiency, Serpina1a-e-knockout mice. Human bronchial epithelial cells were treated with ATUX-792 or DBK-1154, and cell viability, PP2A activity, and MAP (mitogen-activated protein) kinase phosphorylation status were examined. Wild-type mice received vehicle, DBK-1154, or ATUX-792 orally in the last 2 months of 4 months of SM exposure, and 8-month-old Serpina1a-e-knockout mice received ATUX-792 daily for 4 months. Forced oscillation and expiratory measurements and histology analysis were performed. Treatment with ATUX-792 or DBK-1154 resulted in PP2A activation, reduced MAP kinase phosphorylation, immune cell infiltration, reduced airspace enlargements, and preserved lung function. Using protein arrays and multiplex assays, PP2A activation was observed to reduce AAT-deficient and SM-induced release of CXCL5, CCL17, and CXCL16 into the airways, which coincided with reduced neutrophil lung infiltration. Our study indicates that suppression of the PP2A activity in two models of emphysema could be restored by next-generation PP2A activators to impact lung function.

Agarwood Oil Nanoemulsion Attenuates Cigarette Smoke-Induced Inflammation and Oxidative Stress Markers in BCi-NS1.1 Airway Epithelial Cells

Chronic obstructive pulmonary disease (COPD) is an irreversible inflammatory respiratory disease characterized by frequent exacerbations and symptoms such as cough and wheezing that lead to irreversible airway damage and hyperresponsiveness. The primary risk factor for COPD is chronic cigarette smoke exposure, which promotes oxidative stress and a general pro-inflammatory condition by stimulating pro-oxidant and pro-inflammatory pathways and, simultaneously, inactivating anti-inflammatory and antioxidant detoxification pathways. These events cause progressive damage resulting in impaired cell function and disease progression. Treatments available for COPD are generally aimed at reducing the symptoms of exacerbation. Failure to regulate oxidative stress and inflammation results in lung damage. In the quest for innovative treatment strategies, phytochemicals, and complex plant extracts such as agarwood essential oil are promising sources of molecules with antioxidant and anti-inflammatory activity. However, their clinical use is limited by issues such as low solubility and poor pharmacokinetic properties. These can be overcome by encapsulating the therapeutic molecules using advanced drug delivery systems such as polymeric nanosystems and nanoemulsions. In this study, agarwood oil nanoemulsion (agarwood-NE) was formulated and tested for its antioxidant and anti-inflammatory potential in cigarette smoke extract (CSE)-treated BCi-NS1.1 airway basal epithelial cells. The findings suggest successful counteractivity of agarwood-NE against CSE-mediated pro-inflammatory effects by reducing the expression of the pro-inflammatory cytokines IL-1α, IL-1β, IL-8, and GDF-15. In addition, agarwood-NE induced the expression of the anti-inflammatory mediators IL-10, IL-18BP, TFF3, GH, VDBP, relaxin-2, IFN-γ, and PDGF. Furthermore, agarwood-NE also induced the expression of antioxidant genes such as GCLC and GSTP1, simultaneously activating the PI3K pro-survival signalling pathway. This study provides proof of the dual anti-inflammatory and antioxidant activity of agarwood-NE, highlighting its enormous potential for COPD treatment.

Mesenchymal stem cells attenuate the proinflammatory cytokine pattern in a guinea pig model of chronic cigarette smoke exposure.

Cigarette smoke often induces pulmonary and systemic inflammation. In animal models, mesenchymal stem cells (MSC) tend to ameliorate these effects. We aimed to explore the local and systemic expression of cytokines in guinea pigs chronically exposed to cigarette smoke, and their modifications by MSC. Concentrations of IL-1β, IL-6, IL-8, IL-12, TNF-α, INF-ɣ, TSG-6, MMP-9, TIMP-1, and/or TIMP-2 in serum and bronchoalveolar lavage (BALF) from animals exposed to tobacco smoke (20 cigarettes/day, 5days/week for 10weeks) were determined, and mRNA expression of some of them was measured in lung tissue. Intratracheal instillation of allogeneic bone marrow MSC (5x106 cells in 1ml) was done at week 2. After cigarette smoke, IL-6 and IFN-γ increased in serum and BALF, while IL-1β and IL-12 decreased in serum, and TSG-6 and TIMP-2 increased in BALF. IL-1β had a paradoxical increase in BALF. MSC had an almost null effect in unexposed animals. The intratracheal administration of MSC in guinea pigs exposed to cigarette smoke was associated with a statistically significant decrease of IL-12 and TSG-6 in serum, as well as a decrease of IL-1β and IFN-γ and an increase in TIMP-1 in BALF. Concerning mRNA expression in lung tissue, cigarette smoke did not modify the relative amount of the studied transcripts, but even so, MSC decreased the IL-12 mRNA and increased the TIMP-1 mRNA. A single intratracheal instillation of MSC reduces the pulmonary and systemic proinflammatory pattern induced by chronic exposure to cigarette smoke in guinea pigs. Not applicable.

Evaluation of chronic cigarette smoke exposure in human bronchial epithelial cultures.

Cigarette smoke (CS) exposure induces both cytotoxicity and inflammation, and often causes COPD, a growing cause of morbidity and mortality. CS also inhibits the CFTR Cl- channel, leading to airway surface liquid dehydration, which is predicated to impair clearance of inhaled pathogens and toxicants. Numerous in vitro studies have been performed that utilize acute (≤24 h) CS exposures. However, CS exposure is typically chronic. We evaluated the feasibility of using British-American Tobacco (BAT)-designed CS exposure chambers for chronically exposing human bronchial epithelial cultures (HBECs) to CS. HBECs are polarized and contain mucosal and serosal sides. In vivo, inhaled CS interacts with mucosal membranes, and BAT chambers are designed to direct CS to HBEC mucosal surfaces while keeping CS away from serosal surfaces via a perfusion system. We found that serosal perfusion was absolutely required to maintain HBEC viability over time following chronic CS exposure. Indeed, with this system, we found that CS increased inflammation and mucin levels, while decreasing CFTR function. Without this serosal perfusion, CS was extremely toxic within 24 h. We therefore propose that 5- and 10-day CS exposures with serosal perfusion are suitable for measuring chronic CS exposure and can be used for monitoring new and emerging tobacco products.

Inhibition of neutrophil elastase prevents cigarette smoke exposure-induced formation of neutrophil extracellular traps and improves lung function in a mouse model of chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is an important public health challenge worldwide, and is usually caused by significant exposure to noxious agents, particularly cigarette smoke. Recent studies have revealed that excessive production of neutrophil extracellular traps (NETs) in the airways is associated with disease severity in COPD patients. NETs are extracellular neutrophil-derived structures composed of chromatin fibers decorated with histones and granule proteases including neutrophil elastase (NE). However, the effective prevention of NET formation in COPD remains elusive. Here, we demonstrated that treatment with GW311616A, a potent and selective inhibitor of NE, prevented cigarette smoke extract (CSE)-induced NET formation in human neutrophils by blocking NE nuclear translocation and subsequent chromatin decondensation. Inhibition of NE also abrogated CSE-induced ROS production and migration impairment of neutrophils. Administration of GW311616A in vivo substantially reduced pulmonary generation of NETs while attenuating the key pathological changes in COPD, including airway leukocyte infiltration, mucus-secreting goblet cell hyperplasia, and emphysema-like alveolar destruction in a mouse model of COPD induced by chronic cigarette smoke exposure. Mice treated with GW311616A also showed significant attenuation of neutrophil numbers and percentages and the levels of neutrophil chemotactic factors (LTB4, KC, and CXCL5) and proinflammatory cytokines (IL-1β, and TNF-α) in bronchoalveolar lavage fluid compared to mice treated with cigarette smoke exposure only. Furthermore, GW311616A treatment considerably improved lung function in the COPD mouse model, including preventing the decline of FEV100/FVC and delta PEF as well as inhibiting the increase in FRC, TLC, and FRC/TLC. Overall, our study suggests that NE plays a critical role in cigarette smoke-induced NET formation by neutrophils and that inhibition of NE is a promising strategy to suppress NET-mediated pathophysiological changes in COPD.

RIPK1 kinase-dependent inflammation and cell death contribute to the pathogenesis of COPD.

Receptor-interacting protein kinase 1 (RIPK1) is a key mediator of regulated cell death (including apoptosis and necroptosis) and inflammation, both drivers of COPD pathogenesis. We aimed to define the contribution of RIPK1 kinase-dependent cell death and inflammation in the pathogenesis of COPD. We assessed RIPK1 expression in single-cell RNA sequencing (RNA-seq) data from human and mouse lungs, and validated RIPK1 levels in lung tissue of COPD patients via immunohistochemistry. Next, we assessed the consequences of genetic and pharmacological inhibition of RIPK1 kinase activity in experimental COPD, using Ripk1 S25D/S25D kinase-deficient mice and the RIPK1 kinase inhibitor GSK'547. RIPK1 expression increased in alveolar type 1 (AT1), AT2, ciliated and neuroendocrine cells in human COPD. RIPK1 protein levels were significantly increased in airway epithelium of COPD patients compared with never-smokers and smokers without airflow limitation. In mice, exposure to cigarette smoke (CS) increased Ripk1 expression similarly in AT2 cells, and further in alveolar macrophages and T-cells. Genetic and/or pharmacological inhibition of RIPK1 kinase activity significantly attenuated airway inflammation upon acute and subacute CS exposure, as well as airway remodelling, emphysema, and apoptotic and necroptotic cell death upon chronic CS exposure. Similarly, pharmacological RIPK1 kinase inhibition significantly attenuated elastase-induced emphysema and lung function decline. Finally, RNA-seq on lung tissue of CS-exposed mice revealed downregulation of cell death and inflammatory pathways upon pharmacological RIPK1 kinase inhibition. RIPK1 kinase inhibition is protective in experimental models of COPD and may represent a novel promising therapeutic approach.

Respiratory mechanics following chronic cigarette smoke exposure in the Apoe[Formula: see text] mouse model.

Even though cigarette smoking (CS) has been on the decline over the past 50 years, it is still the leading cause of preventable premature death in the United States. Preclinical models have investigated the cardiopulmonary effects of CS exposure (CSE), but the structure-function relationship in the respiratory system has not yet been fully explored. To evaluate these relationships, we exposed female apolipoprotein E-deficient (Apoe[Formula: see text]) mice to mainstream CS ([Formula: see text]) for 5 days/week over 24 weeks with room air as a control (AE, [Formula: see text]). To contextualize the impact of CSE, we also assessed the natural aging effects over 24 weeks of air exposure (baseline, [Formula: see text]). Functional assessments were performed on a small animal mechanical ventilator (flexiVent, SCIREQ), where pressure-volume curves and impedance data at four levels of positive end-expiratory pressure ([Formula: see text]) and with increasing doses of methacholine were collected. Constant phase model parameters ([Formula: see text]: Newtonian resistance, H: coefficient of tissue elastance, and G: coefficient of tissue resistance) were calculated from the impedance data. Perfusion fixed-left lung tissue was utilized for quantification of parenchyma airspace size and tissue thickness, airway wall thickness, and measurements of elastin, cytoplasm + nucleus, fibrin, and collagen content for the parenchyma and airways. Aging caused the lung to become more compliant, with an upward-leftward shift of the pressure-volume curve and a reduction in all constant phase model parameters. This was supported by larger parenchyma airspace sizes, with a reduction in cell cytoplasm + nucleus area. Airway walls became thinner, even though low-density collagen content increased. In contrast, CSE caused a downward-rightward shift of the pressure-volume curve along with an increase in H, G, and hysteresivity ([Formula: see text]). Organ stiffening was accompanied by enhanced airway hyper-responsiveness following methacholine challenge. Structurally, parenchyma airspaces enlarged, as indicated by an increase in equivalent airspace diameter ([Formula: see text]), and the septum thickened with significant deposition of low-density collagen along with an influx of cells. Airway walls thickened due to deposition of both high and low-density collagen, infiltration of cells, and epithelial cell elongation. In all, our data suggest that CSE in female Apoe[Formula: see text] mice reduces respiratory functionality and causes morphological alterations in both central and peripheral airways that results in lung stiffening, compared to AE controls.

Hypoxia and Cigarette Smoke - A Combined Threat to the Brain: Role of Toll-like Receptor 4 in Neuroinflammatory Changes

The mammalian brain is composed of 20% of oxygen. The depletion in the level of oxygen may lead to hypoxia, resulting in neurological changes. Recent findings of Toll-like receptors (TLR) such as TLR-2, 3, 4 and 8 in the mammalian nervous system showedneuroinflammation in the brain. The activation of TLR4 by both exogenous and endogenous ligands leads to various inflammatory diseases likeatherosclerosis, Crohn’s disease, ulcerative colitis, rheumatoid arthritis and prostate cancer. This study confirms the involvement of NF-κB in TLR4-axis-induced neuroinflammationin the brain. Cigarette Smoke contains many toxic chemicals such as carcinogenic compounds which areinjurious to health, causing death worldwide. The role of TLR4 in activating the Brain during Cigarette Smoke exposure is not yet understood. Hence this study reviewed the use of the cell culture in vitro model as well as in vivo mouse model of TLR4-/- mouse exposed to hypobaric hypoxia to test hypothesis. Methodology: This review explored articles fromthe past 15 years to study the action of toll-like receptor 4 in neuroinflammatory changes. Those articles which lacked full texts were excluded from the review. A total of 61 articles were enrolled in this review article. Results: The recent findings of TLR in Neurons, showed the involvement of external stimuli such as Cigarette smoke causing various neurological disorders and brain inflammation. The role of TLR4 in activating the brain during acute and chronic cigarette smoke exposure is not yet investigated and also the neuroimmune mechanisms of cigarette smoke are not yet clearly understood. Conclusion: Smoking-related neurological disorders requirea proper understanding of the mechanisms of such damage for therapeutic development. Hence research should be carried out to study the TLR4 mechanisms and pathways in various inflammatory diseases.

Glycyl-L-histidyl-L-lysine-Cu2+ attenuates cigarette smoke-induced pulmonary emphysema and inflammation by reducing oxidative stress pathway.

Background: Chronic obstructive pulmonary disease (COPD) is a common respiratory disorder manifested as chronic airway inflammation and persistent airflow limitation with the essential mechanism as inflammatory response and oxidative stress induced by toxic exposures such as cigarette smoke (CS). Glycyl-L-histidyl-L-lysine (GHK) is a nontoxic tripeptide involved in the process of healing and regeneration as a natural product. With the combination of Cu(II), glycyl-L-histidyl-L-lysine-Cu2+ (GHK-Cu) improves antioxidative and anti-inflammatory bioavailability, and they might offer potential therapeutic properties for COPD. Thus, the present study aimed to identify the potential effects of GHK-Cu on emphysema induced by cigarette smoke. Methods: In the in vivo experiment, C57BL/6J mice were exposed to CS for 12 weeks to induce pulmonary emphysema. GHK-Cu was injected intraperitoneally at doses of 0.2, 2 and 20 μg/g/day in 100 µl of saline on alternative days from the 1st day after CS exposure. The effects of GHK-Cu on the morphology of CS-induced emphysema, the inflammatory response and oxidative stress were evaluated. The antioxidative effect of GHK-Cu on human alveolar epithelial A549 cells was assessed in vitro. Results: GHK-Cu treatment attenuated the CS-induced emphysematous changes and partially reversed the matrix metalloprotein -9 (MMP-9)/tissue inhibitor of metalloproteinases-1 (TIMP-1) imbalance in the lung tissue. GHK-Cu reduced the inflammation and oxidation by decreasing the expression of inflammatory cytokines (IL-1β and TNF-α) in the bronchoalveolar lavage and the enzymatic activity of MPO and MDA in the lung homogenate while restoring the T-AOC and GSH content. Furthermore, administration of GHK-Cu reversed the increase in NF-κB expression induced by CS and increased the Nrf2 level, as an antioxidant defense component, in mice with chronic CS exposure. In CSE-exposed human alveolar epithelial A549 cells, GHK-Cu also inhibited oxidative stress by suppressing MDA levels and restoring T-AOC and GSH levels, which were modulated by upregulating Nrf2 expression. Conclusion: GHK-Cu treatment attenuated CS-induced emphysema by anti-inflammation by downregulating NF-κB and antioxidation via upregulation of the Nrf2/Keap1 in lung tissues.

Airway and parenchymal transcriptomics in a novel model of asthma and COPD overlap

Asthma and chronic obstructive pulmonary disease (COPD) are common chronic respiratory diseases, and some patients have overlapping disease features, termed asthma-COPD overlap (ACO). Patients characterized with ACO have increased disease severity; however, the mechanisms driving this have not been widely studied. This study sought to characterize the phenotypic and transcriptomic features of experimental ACO in mice induced by chronic house dust mite antigen and cigarette smoke exposure. Female BALB/c mice were chronically exposed to house dust mite antigen for 11 weeks to induce experimental asthma, cigarette smoke for 8 weeks to induce experimental COPD, or both concurrently to induce experimental ACO. Lung inflammation, structural changes, and lung function were assessed. RNA-sequencing was performed on separated airway and parenchyma lung tissues to assess transcriptional changes. Validation of a novel upstream driver SPI1 in experimental ACO was assessed using the pharmacological SPI1 inhibitor, DB2313. Experimental ACO recapitulated features of both asthma and COPD, with mixed pulmonary eosinophilic/neutrophilic inflammation, small airway collagen deposition, and increased airway hyperresponsiveness. Transcriptomic analysis identified common and distinct dysregulated gene clusters in airway and parenchyma samples in experimental asthma, COPD, and ACO. Upstream driver analysis revealed increased expression of the transcription factor Spi1. Pharmacological inhibition of SPI1 using DB2313, reduced airway remodeling and airway hyperresponsiveness in experimental ACO. A new experimental model of ACO featuring chronic dual exposures to house dust mite and cigarette smoke mimics key disease features observed in patients with ACO and revealed novel disease mechanisms, including upregulation of SPI1, that are amenable to therapy.

Cigarette Smoke Exposure Induces Neurocognitive Impairments and Neuropathological Changes in the Hippocampus

Background and ObjectiveNeurocognitive dysfunction is present in up to ∼61% of people with chronic obstructive pulmonary disease (COPD), with symptoms including learning and memory deficiencies, negatively impacting the quality of life of these individuals. As the mechanisms responsible for neurocognitive deficits in COPD remain unknown, we explored whether chronic cigarette smoke (CS) exposure causes neurocognitive dysfunction in mice and whether this is associated with neuroinflammation and an altered neuropathology.MethodsMale BALB/c mice were exposed to room air (sham) or CS (9 cigarettes/day, 5 days/week) for 24 weeks. After 23 weeks, mice underwent neurocognitive tests to assess working and spatial memory retention. At 24 weeks, mice were culled and lungs were collected and assessed for hallmark features of COPD. Serum was assessed for systemic inflammation and the hippocampus was collected for neuroinflammatory and structural analysis.ResultsChronic CS exposure impaired lung function as well as driving pulmonary inflammation, emphysema, and systemic inflammation. CS exposure impaired working memory retention, which was associated with a suppression in hippocampal microglial number, however, these microglia displayed a more activated morphology. CS-exposed mice showed changes in astrocyte density as well as a reduction in synaptophysin and dendritic spines in the hippocampus.ConclusionWe have developed an experimental model of COPD in mice that recapitulates the hallmark features of the human disease. The altered microglial/astrocytic profiles and alterations in the neuropathology within the hippocampus may explain the neurocognitive dysfunction observed during COPD.

Cellular Senescence as Fibrogenic Mechanism in Smoking Lung

Age is a risk factor for many diseases including idiopathic pulmonary fibrosis (IPF) by affecting wound healing through abnormal fibrogenesis. Common cellular and molecular traits associated with aging can be found in senescent cells, which may occur throughout the lifespan playing physiological roles during normal development or limiting the proliferation of damaged and aged cells. In this study we investigated in smoking mice whether the senescence of lung cells, triggered by the oxidative stress response, is associated with molecular pathways that can influence lung fibrogenesis and contribute to the remodeling of the fibrotic tissue characterizing some phenotypes of COPD in humans. Among the complex mixture of hundreds senescence secreted factors that include proinflammatory cytokines, chemokines, growth factors, and proteases, we selected important factors involved in differentiation of myofibroblasts, their dedifferentiation and apoptosis, as well as in their metabolic function.The expression of these factors and the amount of matrix deposition was analyzed after chronic cigarette smoke (CS) exposure of C57 Bl/6 and DBA/2 mice sensitive to oxidative, in different lung compartmentscharacterized by a fibrotic remodeling using histochemistry or immunohistochemistry.Unlike C57 Bl/6, DBA/2 mice at 4 months of exposure show marked positivity for 8‐OHdG, a marker of oxidative damage, in subpleural areas, in central parts of the lung parenchyma, on airways and on cells of the fibro‐muscular layer.In these areas, a marked positivity is observed for fibrogenic cytokines such as TGF‐β, PDGF‐b and CTGF, and for other factors such as the p16ink4A senescence marker, and proliferation markers PCNA and Ki67. In DBA/2 mice, these compartments are characterized by a progressive fibrous remodeling at various experimental time points from 5 months of CS exposure onwards as demonstrated by Masson’s trichrome staining.Fibroproliferative foci are present in sub‐pleural areas, in peripheral areas of airways, and in central parts of the lung parenchyma. They are characterized by increasing number of α‐SMA positive myofibroblasts, which express the senescence marker p16ink4A and the transcription factor MyoD that plays a crucial role in myoblast proliferation and differentiation. MyoD induces cell cycle arrest, promotes cellular terminal differentiation of fibroblasts into myofibroblasts, and in senescent myofibroblasts it has been reported to determine resistance to apoptosis and opposes their dedifferentiation.All these factors contribute to the development in smoking DBA/2 mice of lung lesions, which are like those seen in patients with “Combined Fibrosis/Emphysema Syndrome.”Despite consistent levels of TGF‐β, PDGF‐b and CTGF, C57 Bl/6 mice develop after chronic CS exposure fibrotic remodeling at lower extent. Of interest, a very low expression of factors indicative of senescence (p16ink4A and MyoD) and cell proliferation (PCNA and Ki67) is observed.Our results suggest that apoptosis, senescence, and proliferation induced at different rate by fibrotic cytokines and senescence associated secretory factors play important role in early or late appearance of fibrotic remodeling of lung and airways exposed to CS.

Ebselen abolishes vascular dysfunction in influenza A virus-induced exacerbations of cigarette smoke-induced lung inflammation in mice.

People with chronic obstructive pulmonary disease (COPD) are susceptible to respiratory infections which exacerbate pulmonary and/or cardiovascular complications, increasing their likelihood of death. The mechanisms driving these complications remain unknown but increased oxidative stress has been implicated. Here we investigated whether influenza A virus (IAV) infection, following chronic cigarette smoke (CS) exposure, worsens vascular function and if so, whether the antioxidant ebselen alleviates this vascular dysfunction. Male BALB/c mice were exposed to either room air or CS for 8 weeks followed by inoculation with IAV (Mem71, 1 × 104.5 pfu). Mice were treated with ebselen (10 mg/kg) or vehicle (5% w/v CM-cellulose in water) daily. Mice were culled 3- and 10-days post-infection, and their lungs lavaged to assess inflammation. The thoracic aorta was excised to investigate endothelial and smooth muscle dilator responses, expression of key vasodilatory and oxidative stress modulators, infiltrating immune cells and vascular remodelling. CS increased lung inflammation and caused significant vascular endothelial dysfunction, which was worsened by IAV infection. CS-driven increases in vascular oxidative stress, aortic wall remodelling and suppression of endothelial nitric oxide synthase (eNOS) were not affected by IAV infection. CS and IAV infection significantly enhanced T cell recruitment into the aortic wall. Ebselen abolished the exaggerated lung inflammation, vascular dysfunction and increased T cell infiltration in CS and IAV-infected mice. Our findings showed that ebselen treatment abolished vascular dysfunction in IAV-induced exacerbations of CS-induced lung inflammation indicating it may have potential for the treatment of cardiovascular comorbidities seen in acute exacerbations of COPD (AECOPD).