Long-term Cigarette Smoke Exposure Research Articles

Long-Term Cigarette Smoke Exposure and Changes in MiRNA Expression and Proteome in Non-Small-Cell Lung Cancer.

Chronic exposure to cigarette smoke markedly increases the risk for lung cancer. Regulation of gene expression at the post-transcriptional level by miRNAs influences a variety of cancer-related interactomes. Yet, relatively little is known on the effects of long-term cigarette smoke exposure on miRNA expression and gene regulation. NCI-H292 (H292) is a cell line sensitive to cigarette smoke with mucoepidermoid characteristics in culture. We report, in this study, original observations on long-term (12 months) cigarette smoke effects in the H292 cell line, using microarray-based miRNA expression profiling, and stable isotopic labeling with amino acids in cell culture-based quantitative proteomic analysis. We identified 112 upregulated and 147 downregulated miRNAs (by twofold) in cigarette smoke-treated H292 cells. The liquid chromatography-tandem mass spectrometry analysis identified 3,959 proteins, of which, 303 proteins were overexpressed and 112 proteins downregulated (by twofold). We observed 39 miRNA target pairs (proven targets) that were differentially expressed in response to chronic cigarette smoke exposure. Gene ontology analysis of the target proteins revealed enrichment of proteins in biological processes driving metabolism, cell communication, and nucleic acid metabolism. Pathway analysis revealed the enrichment of phagosome maturation, antigen presentation pathway, nuclear factor erythroid 2-related factor 2-mediated oxidative stress response, and cholesterol biosynthesis pathways in cigarette smoke-exposed cells. In conclusion, this report makes an important contribution to knowledge on molecular changes in a lung cell line in response to long term cigarette smoke exposure. The findings might inform future strategies for drug target, biomarker and diagnostics innovation in lung cancer, and clinical oncology. These observations also call for further research on the extent to which continuing or stopping cigarette smoking in patients diagnosed with lung cancer translates into molecular and clinical outcomes.

Long-term cigarette smoke exposure inhibits histone deacetylase 2 expression and enhances the nuclear factor-κB activation in skeletal muscle of mice.

Long-term cigarette smoke induces lung inflammatory injury and chronic obstructive pulmonary disease (COPD), associated with skeletal muscle inflammation. This study aimed at investigating how cigarette smoke promotes skeletal muscle inflammation and its molecular pathogenesis. Mice were exposed to air or cigarette smoke for 12 or 24 weeks, and C2C12 cells were stimulated with cigarette smoke extract (CSE). The mass and function, myotube formation, inflammatory cytokine production, histone deacetylase 2 (HDAC2) and nuclear factor-κB (NF-κB) p65 expression were detected in the gastrocnemius muscles of mice and C2C12 cells. In comparison with the control mice, cigarette smoke significantly damaged the lung and reduced the gastrocnemius muscle mass and body weights in mice. Cigarette smoke significantly down-regulated myosin heavy chain (MHC)-IIβ and HDAC2 expression, but enhanced NF-κBp65, keratinocyte chemoattractant (KC) and tumor necrosis factor (TNF)-α expression in the gastrocnemius muscles. CSE stimulation significantly inhibited the myotube formation, MyoD and HDAC2 expression, but enhanced NF-κBp65 expression, KC and TNF-α production in C2C12 cells, which were enhanced by HDAC2 knockdown and abrogated by a NF-κB inhibitor. CSE significantly inhibited the interaction of HDAC2 with NF-κBp65, and increased the levels of acetyl-NF-κBp65 in C2C12 cells. These data indicated that cigarette smoke inhibited HDAC2 expression and its interaction with NF-κBp65 to stimulate inflammation, contributing to the pathogenesis of COPD-related skeletal muscle atrophy in mice.

In Vivo Effects of Long-Term Cigarette Smoke Exposure on Mammary Tissue in Mice

Cigarette smoking is the leading cause of preventable death worldwide and has been linked to the development and progression of cancer. Many cohort studies have described the link between patients with breast cancer and those with long-term smoking history. Despite the claim of correlation, the mechanism by which cigarette smoke alters normal breast epithelial cells and stroma and contributes to tumor cell growth remains undefined. To investigate whether cigarette smoke promotes ductal epithelial cell hyperplasia by stimulating stromal endothelial cell proliferation, we exposed mice to cigarette smoke for 6 months. We observed epithelial proliferation, increased fibrosis, increased vascularity, and mast cell infiltration. This is the first study to look at the invivo changes in the breast after long-term cigarette smoke exposure and provides a novel insight to understanding how cigarette smoke contributes to early changes that may contribute to tumor formation and progression. In conclusion, this study suggests that cigarette smoke modulates key stromal-epithelial interactions to support increased angiogenesis, desmoplasia, and abnormal ductal epithelial cell growth.

Bone turnover and bone structure in clinical animal models of COPD.

The features of chronic obstructive pulmonary disease(COPD)are chronic inflammatory and emphysema changes in the lungs by long-term cigarette smoke(CS)exposure. Osteoporosis is an important systemic comorbidity of COPD. Severe emphysema and low body mass index(BMI)are independent risk factors for low bone mineral density in COPD patients. However the pathophysiologic mechanisms underlying emphysema and osteoporosis have not been fully elucidated. An established mouse model of CS-induced emphysema with decrease body weight and bone mineral density is important for investigating the cause of osteoporosis in COPD patients.

Regulation of Retinoic Acid Receptor Beta by Interleukin-15 in the Lung during Cigarette Smoking and Influenza Virus Infection.

Virus-induced exacerbations often lead to further impairment of lung function in chronic obstructive pulmonary disease. IL-15 is critical in antiviral immune responses. Retinoic acid (RA) signaling plays an important role in tissue maintenance and repair, particularly in the lung. We studied RA signaling and its relation to IL-15 in the lung during cigarette smoke (CS) exposure and influenza virus infection. In vivo studies show that RA signaling is diminished by long-term CS exposure or influenza virus infection alone, which is further attenuated during infection after CS exposure. RA receptor β (RARβ) is specifically decreased in the lung of IL-15 transgenic (overexpression; IL-15Tg) mice, and a greater reduction in RARβ is found in these mice compared with wild-type (WT) mice after infection. RARβ is increased in IL-15 knockout (IL-15KO) mice compared with WT mice after infection, and the additive effect of CS and virus on RARβ down-regulation is diminished in IL-15KO mice. IL-15 receptor α (IL-15Rα) is increased and RARβ is significantly decreased in lung interstitial macrophages from IL-15Tg mice compared with WT mice. In vitro studies show that IL-15 down-regulates RARβ in macrophages via IL-15Rα signaling during influenza virus infection. These studies suggest that RA signaling is significantly diminished in the lung by CS exposure and influenza virus infection. IL-15 specifically down-regulates RARβ expression, and RARβ may play a protective role in lung injury caused by CS exposure and viral infections.

Oxidative DNA damage is involved in cigarette smoke-induced lung injury in rats.

Reactive oxygen species (ROS) induced by exogenous toxicants are suggested to be involved in carcinogenesis by oxidative modification of DNA. 8-Hydroxyl-2-deoxyguanosine (8-OHdG) has been considered as a reliable biomarker for oxidative DNA damage both in vivo and in vitro studies. But the effect of smoking on oxidative damage has not yet been fully elucidated. Wistar rats were exposed to cigarette smoke at concentrations of 20 and 60% for 30min, twice/day for 45weeks. Then the histopathology of lung tissues, levels of ROS, 8-OHdG, and total antioxidant (T-AOC), expression of DNA repair enzymes, e.g. 8-oxyguaine DNA glycosylase (OGG1), and MutThomolog 1 (Oxidized Purine Nucleoside Triphosphatase, MTH1) were determined in urine, peripheral blood lymphocytes, and lung tissue. The results showed that long-term cigarette smoke exposure can cause obvious damages of lung tissue in rats. In addition, a significant and cigarette smoke concentration-dependent increase in ROS and 8-OHdG were observed compared with the non-exposed control rats. In contrast, the expression of OGG1 and MTH1, and T-AOC levels were obviously decreased after long-term exposure to cigarette smoke. These findings indicate that long-term exposure to cigarette smoker increases ROS levels, decreases total antioxidant capacity, and interferes DNA repair capacity that eventually induces oxidative DNA damage, which appears to play an important role in cigarette smoke-induced lung injury in rats, and determination of 8-OHdG levels might be a useful method for monitoring oxidative damage in cigarette smokers.

Tc17 cells are associated with cigarette smoke-induced lung inflammation and emphysema.

Some types of T lymphocytes, especially cytotoxic T-cells (Tc1) and T-helper (Th17) cells, play a pivotal role in cigarette smoke-induced lung diseases. However, whether Tc17 cells are involved remains largely unknown. We investigated Tc17 involvement using a cigarette smoke-exposure model. Groups of mice were exposed to cigarette smoke or filtered air. At weeks 2, 8, 12 and 24, mice were sacrificed to observe histological changes by HE stain and/or immunohistochemical staining. The frequency of T cell subsets in the lung and spleen were detected by flow cytometry. In addition, the expression levels of T cell-related factors were measured by real-time polymerase chain reaction or enzyme-linked immunosorbent assay. Cigarette smoke caused substantial inflammatory cell infiltration and led to emphysema. Cigarette smoke exposure promoted the expression of interferon-gamma (IFN)-γ and interleukin (IL)-17A at the messenger ribonucleic acid and protein levels. In addition to Tc1 and Th17 cells, pulmonary and splenic Tc17 cells increased, which was accompanied by the upregulation of cytokines IL-6, transforming growth factor beta (TGF)-β) and transcriptional factors Stat3 and RAR-related orphan receptor gamma. Compared with untreated mice, γH2AX-positive cells were more frequently observed in mice exposed to cigarette smoke. Long-term cigarette smoke exposure induced Tc17 cell expansion both locally and distally, which was associated with emphysema and deoxyribonucleic acid damage. As an important source of IL-17A, this T cell subset may be a potential target for chronic obstructive pulmonary disease therapy.

Cigarette smoke increases cardiomyocyte ceramide accumulation and inhibits mitochondrial respiration.

BackgroundCigarette smoking is a common and lethal worldwide habit, with considerable mortality stemming from its deleterious effects on heart function. While current theories posit altered blood lipids and fibrinogen metabolism as likely mediators, none have explored the role of the sphingolipid ceramide in exacerbating heart function with smoke exposure. Ceramide production is a consequence of cigarette smoke in the lung, and considering ceramide’s harmful effects on mitochondrial function, we sought to elucidate the role of ceramide in mediating smoke-induced altered heart mitochondrial respiration.MethodsLung cells (A549) were exposed to cigarette smoke extract (CSE) and heart cells (H9C2) were exposed to the lung-cell conditioned medium. Adult male mice were exposed sidestream cigarette smoke for 8 wk with dietary intervention and ceramide inhibition. Ceramides and heart cell or myocardial mitochondrial respiration were determined.ResultsLung cell cultures revealed a robust response to cigarette smoke extract in both production and secretion of ceramides. Heart cells incubated with lung-cell conditioned medium revealed a pronounced inhibition of myocardial mitochondrial respiration, though this effect was mitigated with ceramide inhibition via myriocin. In vivo, heart ceramides increased roughly 600% in adult mice with long-term sidestream cigarette smoke exposure. This resulted in a significant ceramide-dependent reduction in left myocardial mitochondrial respiration, as heart mitochondria from the mice exposed to both smoke and myriocin injections respired normally.ConclusionsThese results suggest ceramide to be an important mediator of altered myocardial mitochondrial function with cigarette smoke exposure. Thus, anti-ceramide therapies might be considered in the future to protect heart mitochondrial function with smoke exposure.

Impact of inflammation, emphysema, and smoking cessation on V/Q in mouse models of lung obstruction

BackgroundChronic obstructive pulmonary disease (COPD) is known to greatly affect ventilation (V) and perfusion (Q) of the lung through pathologies such as inflammation and emphysema. However, there is little direct evidence regarding how these pathologies contribute to the V/Q mismatch observed in COPD and models thereof. Also, little is known regarding how smoking cessation affects V/Q relationships after inflammation and airspace enlargement have become established. To this end, we have quantified V/Q on a per-voxel basis using single photon emission computed tomography (SPECT) in mouse models of COPD and lung obstruction.MethodsThree distinct murine models were used to investigate the impact of different pathologies on V/Q, as measured by SPECT. Lipopolysaccharide (LPS) was used to produce neutrophilic inflammation, porcine pancreatic elastase (PPE) was used to produce emphysema, and long-term cigarette smoke (CS) exposure and cessation were used to investigate the combination of these pathologies.ResultsCS exposure resulted in an increase in mononuclear cells and neutrophils, an increase in airspace enlargement, and an increase in V/Q mismatching. The inflammation produced by LPS was more robust and predominantly neutrophilic, compared to that of cigarette smoke; nevertheless, inflammation alone caused V/Q mismatching similar to that seen with long-term CS exposure. The emphysematous lesions caused by PPE administration were also capable of causing V/Q mismatch in the absence of inflammation. Following CS cessation, inflammatory cell levels returned to those of controls and, similarly, V/Q measures returned to normal despite evidence of persistent mild airspace enlargement.ConclusionsBoth robust inflammation and extensive airspace enlargement, on their own, were capable of producing V/Q mismatch. As CS cessation resulted in a return of V/Q mismatching and inflammatory cell counts to control levels, lung inflammation is likely a major contributor to V/Q mismatch observed in the cigarette smoke exposure model as well as in COPD patients. This return of V/Q mismatching to control values also took place in the presence of mild airspace enlargement, indicating that emphysematous lesions must be of a larger volume before affecting the lung significantly. Early smoking cessation is therefore critical before emphysema has an irreversible impact on gas exchange.

Long-term cigarette smoke exposure and the memory ability of rats of different ages

Context: Cigarette smoke had been implicated in the aetiopathogenesis of certain features seen in children. Aims: The study is aimed at assessing the effects of high doses and long-term exposure to cigarette smoke (CSE) on the memory. Settings and Design: The study involved exposed and non-exposed rats. Materials and Methods: The rats were assigned into groups A, B, C and D respectively. Each group had 5 rats. Groups A was exposed to smoke in utero only and group C from 2 to 5 months of age. Groups B and D served as the controls, respectively. Exposures to cigarette smoke was monitored at 200 PPM of carbon monoxide. Assessment of memory was done by comparing two reference time records separated by 10 minutes, obtained from the water maze task for each rat just after the entire period of exposure to cigarette smoke. Statistical Analysis Used: A paired t-test was used to assess the differences using statistical package for the social sciences software (SPSS version 15). Results: The actual reference time records revealed a statistically significant difference between the initial and final water maze task time records. All the exposed 5-month-old rats showed the lowest repeat time records and also more predictability in their repeat time records. Conclusions: Regardless of the age at the onset and duration of exposure to cigarette smoke, all the rats were able to attain memory for the first session of water task. The 5 months of exposed rat attained more memory. Hence, long-term CSE was beneficial to memory attainment.

TLR and NKG2D Signaling Pathways Mediate CS-Induced Pulmonary Pathologies

Long-term exposure to cigarette smoke (CS) can have deleterious effects on lung epithelial cells including cell death and the initiation of inflammatory responses. CS-induced cell injury can elaborate cell surface signals and cellular byproducts that stimulate immune system surveillance. Our previous work has shown that the expression of ligands for the cytotoxic lymphocyte activating receptor NKG2D is enhanced in patients with COPD and that the induction of these ligands in a mouse model can replicate COPD pathologies. Here, we extend these findings to demonstrate a role for the NKG2D receptor in CS-induced pathophysiology and provide evidence linking nucleic acid-sensing endosomal toll-like receptor (TLR) signaling to COPD pathology through NKG2D activation. Specifically, we show that mice deficient in NKG2D exhibit attenuated pulmonary inflammation and airspace enlargement in a model of CS-induced emphysema. Additionally, we show that CS exposure induces the release of free nucleic acids in the bronchoalveolar lavage and that direct exposure of mouse lung epithelial cells to cigarette smoke extract similarly induces functional nucleic acids as assessed by TLR3, 7, and 9 reporter cell lines. We demonstrate that exposure of mouse lung epithelial cells to TLR ligands stimulates the surface expression of RAET1, a ligand for NKG2D, and that mice deficient in TLR3/7/9 receptor signaling do not exhibit CS-induced NK cell hyperresponsiveness and airspace enlargement. The findings indicate that CS-induced airway injury stimulates TLR signaling by endogenous nucleic acids leading to elevated NKG2D ligand expression. Activation of these pathways plays a major role in the altered NK cell function, pulmonary inflammation and remodeling related to long-term CS exposure.

Die Auswirkungen von Tabakrauch auf Alveolarmakrophagen - ein In-vivo-Kurzzeitrauchmodell zur Mauslunge

The effect of cigarette smoke (CS) on the phagocytosis of alveolar macrophages is discussed controversially on the basis of in vitro experiments. In this short report we describe the in vivo observations that we have performed. For this purpose mice were exposed to CS for three consecutive days. One day later the fluorescent microspheres were administered intratracheally and the lung surface was investigated using long-distance fluorescence microscopy. We found that the numbers of neutrophils which engulfed particles was increased in the CS group as compared to controls. The overall phagocytic activity was not significantly different after CS exposure. In conclusion the phagocytosis of alveolar macrophages and neutrophils after short time CS exposure was not affected. Further investigations will need to look for the effects of long-term CS exposure and the phagocytosis of living bacteria.

NKG2D Mediates NK Cell Hyperresponsiveness and Influenza-Induced Pathologies in a Mouse Model of Chronic Obstructive Pulmonary Disease

Chronic obstructive pulmonary disease (COPD) is characterized by peribronchial and perivascular inflammation and largely irreversible airflow obstruction. Acute disease exacerbations, due frequently to viral infections, lead to enhanced disease symptoms and contribute to long-term progression of COPD pathology. Previously, we demonstrated that NK cells from cigarette smoke (CS)-exposed mice exhibit enhanced effector functions in response to stimulating cytokines or TLR ligands. In this article, we show that the activating receptor NKG2D is a key mediator for CS-stimulated NK cell hyperresponsiveness, because CS-exposed NKG2D-deficient mice (Klrk1(-/-)) did not exhibit enhanced effector functions as assessed by cytokine responsiveness. NK cell cytotoxicity against MHC class I-deficient targets was not affected in a COPD model. However, NK cells from CS-exposed mice exhibit greater cytotoxic activity toward cells that express the NKG2D ligand RAET1ε. We also demonstrate that NKG2D-deficient mice exhibit diminished airway damage and reduced inflammation in a model of viral COPD exacerbation, which do not affect viral clearance. Furthermore, adoptive transfer of NKG2D(+) NK cells into CS-exposed, influenza-infected NKG2D-deficient mice recapitulated the phenotypes observed in CS-exposed, influenza-infected wild-type mice. Our findings indicate that NKG2D stimulation during long-term CS exposure is a central pathway in the development of NK cell hyperresponsiveness and influenza-mediated exacerbations of COPD.

Long-term nose-only cigarette smoke exposure induces emphysema and mild skeletal muscle dysfunction in mice

SUMMARYMouse models of chronic obstructive pulmonary disease (COPD) focus on airway inflammation and lung histology, but their use has been hampered by the lack of pulmonary function data in their assessment. Systemic effects such as muscle dysfunction are also poorly modeled in emphysematous mice. We aimed to develop a cigarette-smoke-induced emphysema mouse model in which serial lung function and muscular dysfunction could be assessed, allowing the disease to be monitored more appropriately. C57Bl6 mice were nose-only exposed to cigarette smoke or filtered air for 3–6 months. Lung function tests were repeated in the same mice after 3 and 6 months of cigarette smoke or air exposure and compared with lung histological changes. Contractile properties of skeletal muscles and muscle histology were also determined at similar time points in separate groups of mice. Serial lung function measurements documented hyperinflation after 3 and 6 months of cigarette smoke exposure, with a significant 31–37% increase in total lung capacity (TLC) and a significant 26–35% increase in compliance (Cchord) when compared with animals exposed to filtered air only (P<0.001 after 3 and after 6 months). These functional changes preceded the changes in mean linear intercept, which became only significant after 6 months of cigarette smoke exposure and which correlated very well with TLC (r=0.74, P=0.004) and Cchord (r=0.79, P=0.001). After 6 months of cigarette smoke exposure, a significant fiber-type shift from IIa to IIx/b was also observed in the soleus muscle (P<0.05), whereas a 20% reduction of force was present at high stimulation frequencies (80 Hz; P=0.09). The extensor digitorum longus (EDL) muscle was not affected by cigarette smoke exposure. These serial pulmonary function variables are sensitive outcomes to detect emphysema progression in a nose-only cigarette-smoke-exposed animal model of COPD. In this model, muscular changes became apparent only after 6 months, particularly in muscles with a mixed fiber-type composition.

Lung-Targeted Overexpression of the NF-κB Member RelB Inhibits Cigarette Smoke–Induced Inflammation

Acute lung inflammation can be caused by a variety of respirable agents, including cigarette smoke. Long-term cigarette smoke exposure can cause chronic obstructive pulmonary disease (COPD), a serious illness that affects >10 million Americans. Cigarette smoke is a known inducer of inflammation and is responsible for approximately 90% of all COPD cases. RelB, a member of the NF-κB family, attenuates cigarette smoke-induced inflammatory mediator production in mouse lung fibroblasts in vitro. We hypothesized that overexpression of RelB in the airways of mice would dampen acute smoke-induced pulmonary inflammation. Mice received a recombinant adenovirus encoding RelB by intranasal aspiration to induce transient RelB overexpression in the lungs and were subsequently exposed to mainstream cigarette smoke. Markers of inflammation were analyzed after smoke exposure. Neutrophil infiltration, normally increased by smoke exposure, was significantly and potently decreased after RelB overexpression. Cigarette smoke-induced proinflammatory cytokine and chemokine production, cyclooxygenase-2 expression, and prostaglandin E(2) production were also significantly decreased in the context of RelB overexpression. The expression of intercellular adhesion molecule 1, an NF-κB-dependent protein, was decreased, indicating a potential mechanism through which RelB can regulate inflammatory cell migration. Therefore, increased expression and/or activation of RelB could be a novel therapeutic strategy against acute lung inflammation caused by respirable agents and possibly against chronic injury, such as COPD.

Long-term exposure to cigarette smoke impairs lung function and increases HMGB-1 expression in mice

Cigarette smoke (CS)-induced emphysema is caused by a continuous inflammatory response in the lower respiratory tract. The development of the condition is believed to be mediated by oxidant-antioxidant imbalance. This paper describes the effects of long-term CS exposure on alveolar cell recruitment, antioxidant defense systems, activity of extracellular matrix metalloelastases, expression of metalloelastase MMP-12, and high mobility group box-1 protein (HMGB-1). Ten C57Bl/6 mice were exposed to 12 cigarettes-a-day for 60 consecutive days, while 10 control animals were exposed to ambient air. After sacrifice, bronchoalveolar lavage fluid (BALF) was removed, and lung tissue underwent biochemical and histological analyses. In CS-exposed animals influx of alveolar macrophages and neutrophils into BALF, lung static elastance, and expression of MMP-12 and HMGB-1 were significantly increased while the activity of antioxidant enzyme was significantly reduced in comparison with control group. Thus, we demonstrated for the first time that long-term CS exposure decreased antioxidant defenses concomitantly with impaired lung function, which was associated with HMGB-1 expression.

Reinke Edema Finding on Positron Emission Tomography

Long-term cigarette smoke exposure has been described as an etiological factor in a myriad of aberrant conditions of the vocal folds, including carcinoma, and a range of benign responses, including Reinke edema (RE). Clinically, RE is described as unilateral or bilateral swelling of the entire vocal fold with hypermobility during phonation. 1 Although historically RE was thought to be a precancerous lesion, current thinking holds that RE is a benign process characterized by altered number and size of microvessels within the subepithelial space and compromised intercellular junctions, especially in basal and spinous layers. 2 Within the lamina propria, the generally organized collagen architecture is disrupted. 3 Cumulatively, these alterations to the delicate, layered structure of the vocal folds often lead to the characteristic rough, low-pitched voice found in RE. In severe cases, airway obstruction due to marked RE has been reported.

Induction of the unfolded protein response by cigarette smoke is primarily an activating transcription factor 4-C/EBP homologous protein mediated process

Purpose:Cigarette smoke is the major risk factor associated with the development of chronic obstructive pulmonary disease (COPD). Recent studies propose a link between endoplasmic reticulum (ER) stress and emphysema, demonstrated by increased ER stress markers under smoking conditions. Here, we investigate whether cigarette smoke-induced ER stress is cell specific and correlates with acute and chronic cigarette smoke exposure.Methods:Gene and protein expression changes in human primary lung cell cultures following cigarette smoke extract (CSE) exposure were monitored by qPCR and Western blot analysis. Mice and guinea pigs were exposed to cigarette smoke and ER stress markers examined in whole lung homogenates. Inflammatory cells from the bronchoalveolar lavage fluid of 10 days smoke exposed mice were also examined.Results:Cigarette smoke induced a trend increase in the ER stress response through an activating transcription factor 4 (ATF4) mediated induction of C/EBP homologous protein (CHOP) in primary small airway epithelial cells. Bronchial epithelial cells and macrophages responded similarly to CSE. Wild-type mice and guinea pigs exposed to acute levels of cigarette smoke exhibited increased levels of CHOP but not at significant levels. However, after long-term chronic cigarette smoke exposure, CHOP expression was reduced. Interestingly, inflammatory cells from smoke exposed mice had a significant increase in CHOP/ATF4 expression.Conclusion:A trend increase in CHOP levels appear in multiple human lung cell types following acute cigarette smoke exposure in vitro. In vivo, inflammatory cells, predominately macrophages, demonstrate significant cigarette smoke-induced ER stress. Early induction of CHOP in cigarette smoke may play a pivotal role in early induction of lung disease, however in vivo long-term cigarette smoke exposure exhibited a reduction in the ER stress response.

N-(2-mercaptopropionyl)-glycine but not Allopurinol prevented cigarette smoke-induced alveolar enlargement in mouse

We investigated the possible protective effects of the Allopurinol (A), N-(2-mercaptopropionyl)-glycine (M) and N-acetylcysteine (N) against lung injury caused by long-term exposure to cigarette smoke (CS) in mouse. C57BL6 mice were exposed to 12 cigarettes a day for 60 days and concomitantly treated with either one of the antioxidant drugs diluted in saline (CS+A-50 mg/kg; CS+M-200 mg/kg/day; CS+N-200 mg/kg/day). Control groups were sham-smoked (AA). Long-term CS exposure results in extensive parenchyma destruction in CS group. Both CS+N and CS+M groups showed preserved alveolar structure and showed preserved lung function when compared to CS group. Macrophage and neutrophil counts were decreased in CS+M, and CS+N groups when compared to CS group (p<0.05). Antioxidant enzyme activities were reduced in all treated groups. CS+A showed the highest reduction in catalase activity (-25%, p<0.01). We conclude that M treatment reduced long-term CS-induced inflammatory lung parenchyma destruction and lung function, comparable to N treatment, however, antioxidant administration did not reverse CS-induced antioxidant enzyme activity reduction.

Long-Term Cigarette Smoke Exposure in a Mouse Model of Ciliated Epithelial Cell Function

Exposure to cigarette smoke is associated with airway epithelial mucus cell hyperplasia and a decrease in cilia and ciliated cells. Few models have addressed the long-term effects of chronic cigarette smoke exposure on ciliated epithelial cells. Our previous in vitro studies showed that cigarette smoke decreases ciliary beat frequency (CBF) via the activation of protein kinase C (PKC). We hypothesized that chronic cigarette smoke exposure in an in vivo model would decrease airway epithelial cell ciliary beating in a PKC-dependent manner. We exposed C57BL/6 mice to whole-body cigarette smoke 2 hours/day, 5 days/week for up to 1 year. Tracheal epithelial cell CBF and the number of motile cells were measured after necropsy in cut tracheal rings, using high-speed digital video microscopy. Tracheal epithelial PKC was assayed according to direct kinase activity. At 6 weeks and 3 months of smoke exposure, the baseline CBF was slightly elevated (~1 Hz) versus control mice, with no change in β-agonist-stimulated CBF between control mice and cigarette smoke-exposed mice. By 6 months of smoke exposure, the baseline CBF was significantly decreased (2-3 Hz) versus control mice, and a β-agonist failed to stimulate increased CBF. The loss of β-agonist-increased CBF continued at 9 months and 12 months of smoke exposure, and the baseline CBF was significantly decreased to less than one third of the control rate. In addition to CBF, ciliated cell numbers significantly decreased in response to smoke over time, with a significant loss of tracheal ciliated cells occurring between 6 and 12 months. In parallel with the slowing of CBF, significant PKC activation from cytosol to the membrane of tracheal epithelial cells was detected in mice exposed to smoke for 6-12 months.