Apoptosis Of Airway Epithelial Cells Research Articles

Herb pair of Ephedrae Herba-Armeniacae Semen Amarum alleviates airway injury in asthmatic rats

Ethnopharmacological relevanceEphedrae Herba (EH, Ephedra sinica Stapf.) and Armeniacae Semen Amarum (ASA, Prunus armeniaca L. var. ansu Maxim.) have been used to treat asthma, cold, fever, and cough in China for thousands of years. Aim of the studyIn this study, we aimed to investigate the optimal ratio of EH and ASA compatibility (EAC) to reduce airway injury in asthmatic rats and its possible mechanism. MethodsRats were sensitized with a mixture of acetylcholine chloride and histamine bisphosphate 1 h before sensitization by intragastric administration of EAC or dexamethasone or saline for 7 days. Subsequently, the ultrastructure of rat airway epithelial tissue changes, apoptosis of the airway epithelial cells, and the expression of mRNA and protein of EGRF and Bcl-2 were detected. ResultsTransmission electron microscope: EAC (groups C and E) had the most prominent effect on repairing airway epithelial cells' ultrastructural changes in asthmatic rats. TUNEL: dexamethasone and EAC (groups B、C、E and F) inhibited the apoptosis of airway epithelial cells in asthmatic rats (P < 0.05). In situ hybridization: EAC (group E) inhibited the overexpression of EGFR and Bcl-2 mRNA (P < 0.05).Western Blotting: EAC (groups A、B、C、E and F) inhibited the upregulation of airway epithelial EGFR and Bcl-2 protein expression (P < 0.01). ConclusionsOur findings indicate that EAC can inhibit abnormal changes in airway epithelial structure and apoptosis of airway epithelial cells, thereby alleviating airway injury. In this study, the best combination of EH and ASA to alleviate airway epithelial injury in asthmatic rats was group E (EH: ASA = 8: 4.5).

Effect of family with sequence similarity 13 member A gene interference on apoptosis and proliferation of human airway epithelial cells and its relationship with small airway remodeling in patients with chronic obstructive pulmonary disease

Objective: To explore the relationship between family with sequence similarity 13 member A (FAM13A) gene and small airway remodeling in chronic obstructive pulmonary disease (COPD), and the effect of interference with FAM13A gene expression on the apoptosis and proliferation phenotype of human airway epithelial cells (16HBE). Methods: From January 2018 to January 2020, 74 patients in the Department of Thoracic Surgery of General Hospital of Ningxia Medical University were treated by surgery for lung tumors or pulmonary bullae. According to the lung function and smoking history, the 74 patients were divided into four groups: non-smoking group with normal lung function (normal group, 23 patients), smoking group with normal lung function (smoking group, 24 patients), non-smoking group with COPD (11 patients) and smoking group with COPD (16 patients). The expression of FAM13A in small airway of each group was detected by immunohistochemistry, and the correlation between FAM13A and the airflow restriction indexes by pulmonary function was analyzed. The shRNA fragment of FAM13A gene was designed, and the shRNA lentivirus vector of FAM13A gene was constructed and packaged. The expression level of FAM13A gene was detected by real-time fluorescent quantitative PCR (qRT-PCR) and Western blot, and the best shRNA sequence was screened. Flow cytometry was used to detect apoptosis rate and the fluorescence intensity of proliferation marker Ki-67 in 16HBE cells. Results: FAM13A was mainly expressed in the cytoplasm of small airway epithelial cells. The levels of FAM13A absorbance (A) of small airway epithelial cells in non-smoking group and smoking group with COPD were higher than those in normal group and smoking group (0.365±0.026, 0.412±0.053 to 0.113±0.018, 0.105±0.009, all P<0.05), and they were negatively correlated with forced expiratory volume in 1s/forced vital capacity (FEV(1)/FVC) and FEV(1)% pre (r=-0.48 and r=-0.40, all P<0.05). The FAM13A shRNA lentiviral vector was successfully constructed, and FAM13A interference was successfully achieved in the 16HBE cell line. After infection of 16HBE cells, the results of qRT-PCR and Western blot showed that the expression of FAM13A in shRNA-target-2 group decreased (all P<0.01). Compared with the negative control group (shRNA-NC), the apoptosis rate of FAM13A shRNA group decreased (P=0.023), and the fluorescence intensity of Ki-67 also decreased (P=0.042). Conclusions: FAM13A gene expression is increased in COPD small airway epithelial cells, and it is related to COPD airflow limitation. FAM13A gene may participate in the process of COPD remodeling by affecting the apoptosis and proliferation of human airway epithelial cells.

Panax notoginseng saponin R1 modulates TNF-α/NF-κB signaling and attenuates allergic airway inflammation in asthma

BackgroudPanax notoginseng saponin R1 (PNS-R1) is one of the most important chemical monomers derived from the panax notoginseng, and our previous study found that PNS-R1 reduced glucocorticoid-induced apoptosis in asthmatic airway epithelial cells. Thus, in this study, we explored the effects of the PNS-R1 on inflammation of allergic asthma. MethodsThe asthmatic mice were administered 15 mg/kg PNS-R1 by intraperitoneal injection three days before sensitized to OVA. The effects of PNS-R1 on asthmatic mice were detected by airway hyperresponsiveness, inflammation, mucus hypersecretion and inflammatory cytokines such as interleukin (IL)-13, IL-4, IL-5, IL-8 and tumor necrosis factor (TNF)-α were studied. We also treated human bronchial epithelial cells (16HBE) with house dust mites (HDM) and then detected the secretion of cellular inflammatory factors (IL-13 and TNF-α). Western blot and immunofluorescence were used to examine the effect of PNS-R1 on TNF-α/NF-κB pathway. TNF-α/NF-κB/IKK signal pathway activator was used in PNS-R1-treated asthmatic mice. ResultsPNS-R1 significantly reduced the airway inflammatory, mucus secretion and hyperresponsiveness in asthma model. It also reduced the levels of IL-13, IL-4, IL-5 and IL-8 in bronchoalveolar lavage fluid (BALF) and IgE and OVA-specific IgE in serum for asthma mice. PNS-R1 reduced IL-13 and TNF-α secretion in HDM-treated 16HBE cells. In addition, PNS-R1 suppressed TNF-α/NF-κB pathway in both asthmatic mice and 16HBE. Activation of NF-kB pathway reversed the therapeutic effect of PNS-R1 on asthmatic mice. ConclusionThe results indicated that PNS-R1 effectively suppresses allergic airway inflammation of asthma partly through TNF-α/NF-κB pathway. PNS-R1 may play a potential role in allergic asthma treatment in the future.

Respiratory exposure to PM2.5 soluble extract disrupts mucosal barrier function and promotes the development of experimental asthma

BackgroundAir pollutants are important factors that contribute to the development and exacerbation of asthma, but experimental evidence still needs to be collected and the mechanisms still need to be addressed. In this study, we aimed to assess the association between PM2.5 exposure and asthma development. The effects of PM2.5 exposure on the barrier functions of airway epithelial cells were also determined. MethodsPM2.5 was collected from Nanjing, China, and its soluble extract was prepared. Human lung epithelial cells (BEAS-2B) were treated with different concentrations of soluble PM2.5 extract, and cell viability was detected by FACS using Annexin V-FITC staining. PM2.5-induced oxidative stress and inflammatory events were assessed by DCF-DA staining and qPCR. PM2.5-induced dysfunction of the airway epithelial barrier was assessed by measuring the expression of tight junction molecules. In vivo, BALB/c mice were treated with OVA in the presence or absence of PM2.5 solution, followed by exposure to OVA aerosols. Allergy-induced airway inflammation and lung injury were assessed by histopathological analyses. ResultsSoluble PM2.5 extract exposure in vitro decreased the viability and increased apoptosis of airway epithelial cells. Soluble PM2.5 extract induced oxidative stress and enhanced pro-inflammatory factor expression by activating the NF-κB and MAPK signalling pathways, which were accompanied by reduced airway barrier function. The in vivo data demonstrated that PM2.5 exposure increased the effects of allergy sensitization after respiratory exposure to allergens, which led to the development of asthma. ConclusionThis study suggests that exposure to soluble PM2.5 extract contributes to airway barrier dysfunction. The soluble mediators generated by airway epithelial cells in response to PM2.5 exposure orchestrate the breaking of inhalational tolerance and sensitization to allergic antigens, leading to the exacerbated development of asthma.

The Protective Effect of HBO1 on Cigarette Smoke Extract-Induced Apoptosis in Airway Epithelial Cells.

PurposeEpigenetic modification is one of most important mechanisms underlying the pathogenesis of chronic obstructive pulmonary disease (COPD). The purpose of this study was to determine whether histone acetyltransferase binding to ORC1 (HBO1) can protect against cigarette smoke (CS)-induced cell apoptosis and sustain normal histone acetylation in COPD.MethodsHuman lung tissue samples were obtained from patients who underwent lung resection. The emphysema mouse model and HBO1 overexpressing mice were each established by intraperitoneal injection with cigarette smoke extract (CSE) or intratracheal lentiviral vectors instillation. TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assays were used to assess apoptotic ratio in mice. The apoptosis of human bronchial epithelial cells (HBECs) was assayed by flow cytometry. HBO1, B-cell lymphoma-2 (BCL-2), and H3K14ac protein expression were detected by Western blotting. HBO1 mRNA expression was measured by quantitative real-time polymerase chain reaction.ResultsProtein expression of HBO1 was decreased significantly in lung tissue from COPD patients and CSE-treated emphysema mouse models. Overexpression of HBO1 attenuated CSE-induced emphysematous changes, as well as apoptosis in the lungs of COPD mice. In vitro, the HBO1 protein degraded in a time- and dose-dependent course with CSE treatment. With flow cytometry, we proved that HBO1 could reverse the apoptosis of HBECs induced by CSE. Furthermore, HBO1 overexpression promoted the expression of anti-apoptotic BCL-2 protein and enhanced H3K14 acetylation in airway epithelial cells.ConclusionThese findings demonstrate that the key histone modulator HBO1 plays a protective role in COPD pathogenesis that may shed light on potential therapeutic targets to inhibit the progress of COPD.

Downregulated Rac1 promotes apoptosis and inhibits the clearance of apoptotic cells in airway epithelial cells, which may be associated with airway hyper-responsiveness in asthma.

The accumulation of airway apoptotic cells may be an important factor causing airway hyper-responsiveness (AHR). Whether the apoptotic cells can be promptly removed is related to the occurrence and course of asthma. In recent years, studies have shown that Rac1 is involved in many cellular biological activities including the formation and elimination of apoptotic cells. In this study, based on the analysis of airway local cells and related factors in asthmatic mice, we evaluated the expression of Rac1 in airway epithelial cells or phagocytes and analysed its relationship with the incidence of apoptosis or scavenging of apoptotic cells. Our data showed that the expression level of Rac1 in asthmatic mice decreased significantly, while the expression of IL-33 increased obviously. The airway epithelial cell line was stimulated by curcumin at 50μmol/L for 24-48hours; more than 50% of the cells were apoptotic, and of which, about 20% were late apoptosis. Rac1 inhibitor (NSC23766) can enhance the apoptosis effect. In addition, the ability of phagocytosis and migration in the epithelial cells or macrophages was increased following the application of Rac1 inhibitors or specific siRNA in a dose-dependent manner, and the expression level of IL-33 was simultaneously increased after blocking Rac1. It is suggested that the down regulation of Rac1 in asthma may contribute to the apoptosis of airway epithelial cells and affect the clearance of apoptotic cells, which will lead to the aggregation of the apoptotic cells in the respiratory tract and participate in AHR.

MiR-145-5p is associated with smoke-related chronic obstructive pulmonary disease via targeting KLF5

Increasing evidence illustrate that dysregulation of microRNAs (miRNAs) is involved in the pathogenesis of chronic obstructive pulmonary disease (COPD), which is mainly resulted from cigarette smoke (CS) exposure. However, the role of miR-145-5p in CS-mediated COPD remains largely unknown. Thus, the aim of this study was to investigate the expression level of miR-145-5p in 31 human lung tissues samples, and to explore its regulatory role in the apoptosis and inflammation of human bronchial epithelial cells (HBECs) following CS extract (CSE) exposure. We found that miR-145-5p was significantly down-regulated in lung tissues from smokers without or with COPD compared to non-smokers. Functional assays showed that miR-145-5p overexpression remarkably alleviated CSE-induced apoptosis and inflammation response by regulating p53-mediated apoptotic signaling and pre-inflammatory factors such as necrosis factor-α (TNF-α), interleukins (IL)-6, IL-8 in HBECs, whereas, down-regulation of miR-145-5p showed opposite effects. Furthermore, luciferase reporter assays verified that Kruppel-like 5 (KLF5) was a direct target of miR-145-5p. Western blot assay also confirmed that KLF5 was up-regulated in COPD tissues and was negatively associated with miR-145-5p expression. Restoration of miR-145-5p expression significantly abrogated the suppressive effect of miR-145-5p on CSE-stimulated apoptosis and inflammation. In addition, the CSE-induced NF-κB signaling activation was suppressed by miR-145-5p overexpression. Therefore, our data suggested that miR-145-5p conferred protection against CSE-induced airway epithelial cell apoptosis and inflammation partially via targeting KLF5, which might be a potential therapeutic biomarker in COPD treatment.

Silencing of Gal-7 inhibits TGF-β1-induced apoptosis of human airway epithelial cells through JNK signaling pathway

Apoptosis of epithelial cells is regarded as the initial pathological process of many lung diseases, including asthma. Previous studies have identified that galectin-7 (Gal-7), a regulator of apoptosis, was overexpressed in bronchial epithelial cells in asthma. However, the effect and mechanism of Gal-7 in the progression of asthma is still unclear. In this study, we investigated the expression and role of Gal-7 in the apoptosis of bronchial epithelial cells BEAS-2B upon TGF-β1 stimulation. TGF-β1 significantly induced apoptosis of BEAS-2B cells, as determined by flow cytometry. Western blot results revealed that the mRNA and protein expression of Gal-7 were obviously increased after TGF-β1 stimulation. Small interfering RNA (siRNA)-mediated knockdown of Gal-7 abrogated TGF-β1-evoked cell apoptosis. Simultaneously, increased Bcl-2 expression, decreased Bax expression and the cleavage of poly ADP-ribose polymerase (PARP) and caspase-3 activity were also monitored in TGF-β1-treated cells after Gal-7 siRNA transfection. Gal-7 silence also inhibited TGF-β1-induced c-Jun N-terminal kinase (JNK) phosphorylation in BEAS-2B cells. Furthermore, anisomycin, a specific activator for JNK, reversed the effect of Gal-7 siRNA on cell apoptosis induced by TGF-β1. These results demonstrate that Gal-7 silence attenuates TGF-β1-induced apoptosis in bronchial epithelial cells through the inactivation of JNK pathway. Therefore, Gal-7 may act as a potential target for asthma treatment.

Heme oxygenase-1 protects airway epithelium against apoptosis by targeting the proinflammatory NLRP3–RXR axis in asthma

Asthma is thought to be caused by malfunction of type 2 T helper cell (Th2)-mediated immunity, causing excessive inflammation, mucus overproduction, and apoptosis of airway epithelial cells. Heme oxygenase-1 (HO-1) functions in heme catabolism and is both cytoprotective and anti-inflammatory. We hypothesized that this dual function may be related to asthma's etiology. Using primary airway epithelial cells (pAECs) and an asthma mouse model, we demonstrate that severe lung inflammation is associated with rapid pAEC apoptosis. Surprisingly, NOD-like receptor protein 3 (NLRP3) inhibition, retinoid X receptor (RXR) deficiency, and HO-1 induction were associated with abrogated apoptosis. MCC950, a selective small-molecule inhibitor of canonical and noncanonical NLRP3 activation, reduced RXR expression, leading to decreased pAEC apoptosis that was reversed by the RXR agonist adapalene. Of note, HO-1 induction in a mouse model of ovalbumin-induced eosinophilic asthma suppressed Th2 responses and reduced apoptosis of pulmonary pAECs. In vitro, HO-1 induction desensitized cultured pAECs to ovalbumin-induced apoptosis, confirming the in vivo observations. Critically, the HO-1 products carbon monoxide and bilirubin suppressed the NLRP3-RXR axis in pAECs. Furthermore, HO-1 impaired production of NLRP3-RXR-induced cytokines (interleukin [IL]-25, IL-33, thymic stromal lymphopoietin, and granulocyte-macrophage colony-stimulating factor) in pAECs and lungs. Finally, we demonstrate that HO-1 binds to the NACHT domain of NLRP3 and the RXRα and RXRβ subunits and that this binding is not reversed by Sn-protoporphyrin. Our findings indicate that HO-1 and its products are essential for pAEC survival to maintain airway epithelium homeostasis during NLRP3-RXR-mediated apoptosis and inflammation.

Role of reciprocal interaction between autophagy and endoplasmic reticulum stress in apoptosis of human bronchial epithelial cells induced by cigarette smoke extract.

Endoplasmic reticulum stress (ERS)-induced apoptosis of airway epithelial cells plays an important role in the pathogenesis of chronic obstructive pulmonary disease (COPD). Furthermore, autophagy is closely related to ERS under apoptosis. Here, this study aimed to investigate the role of the reciprocal interaction between autophagy and ERS in the cigarette smoke extract (CSE)-induced apoptosis of human bronchial epithelial (HBE) cells. Cell apoptosis was detected by flow cytometry analysis. Protein expression was examined by Western blot. The mRNA expression was detected using real-time quantitative reverse transcription PCR (qRT-PCR). The results showed that CSE treatment induced apoptosis, autophagy, and expression of ERS-related proteins in HBE cells. Furthermore, autophagy inhibition by 3-MA significantly decreased protein expression of GRP78, p-PERK, and p-eIF2α and increased CHOP, ATF4, and caspase-4, whereas ERS inhibition by 4-PBA led to autophagy suppression. Moreover, the CSE-induced autophagy was diminished by knockdown of GRP78, PERK, or eIF2α but enhanced by knockdown of ATF4 or CHOP; however, the CSE-induced HBE apoptosis was enhanced by knockdown of GRP78, PERK, or eIF2α but was attenuated by knockdown of ATF4 or CHOP. Additionally, both sodium hydrosulfide (NaHS) and melatonin attenuated the CSE-induced apoptosis, enhanced the CSE-induced autophagy, increased GRP78, p-PERK, and p-eIF2α, and decreased CHOP, ATF4, and caspase-4, via SIRT1/ORP150 pathway. Collectively, this study provided evidence about the role of the reciprocal interaction between autophagy and ERS in CSE-induced apoptosis of HBE cells. © 2018 IUBMB Life, 71(1):66-80, 2019.

Poly-L-Arginine Induces Apoptosis of NCI-H292 Cells via ERK1/2 Signaling Pathway.

Cationic protein is a cytotoxic protein secreted by eosinophils and takes part in the damage of airway epithelium in asthma. Poly-L-arginine (PLA), a synthetic cationic protein, is widely used to mimic the biological function of the natural cationic protein in vitro. Previous studies demonstrated the damage of the airway epithelial cells by cationic protein, but the molecular mechanism is unclear. The purpose of this study aimed at exploring whether PLA could induce apoptosis of human airway epithelial cells (NCI-H292) and the underlying mechanism. Methods. The morphology of apoptotic cells was observed by transmission electron microscopy. The rate of apoptosis was analyzed by flow cytometry (FCM). The expressions of the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), Bcl-2/Bax, and cleaved caspase-3 were assessed by western blot. Results. PLA can induce apoptosis in NCI-H292 cells in a concentration-dependent manner. Moreover, the phosphorylation of the ERK1/2 and the unbalance of Bcl2/Bax, as well as the activation of caspase-3, were involved in the PLA-induced apoptosis. Conclusions. PLA can induce the apoptosis in NCI-H292 cells, and this process at least involved the ERK1/2 and mitochondrial pathway. The results could have some indications in revealing the apoptotic damage of the airway epithelial cells. Besides, inhibition of cationic protein-induced apoptotic death in airway epithelial cells could be considered as a potential target of anti-injury or antiremodeling in asthmatics.

Cystic fibrosis epithelial cells are primed for apoptosis as a result of increased Fas (CD95)

BackgroundPrevious work suggests that apoptosis is dysfunctional in cystic fibrosis (CF) airways with conflicting results. We evaluated the relationship between dysfunctional cystic fibrosis transmembrane conductance regulator (CFTR) and apoptosis in CF airway epithelial cells. MethodsApoptosis and associated caspase activity were analysed in non-CF and CF tracheal and bronchial epithelial cell lines. ResultsBasal levels of apoptosis and activity of caspase-3 and caspase-8 were significantly increased in CF epithelial cells compared to controls, suggesting involvement of extrinsic apoptosis signalling, which is mediated by the activation of death receptors, such as Fas (CD95). Increased levels of Fas were observed in CF epithelial cells and bronchial brushings from CF patients compared to non-CF controls. Neutralisation of Fas significantly inhibited caspase-3 activity in CF epithelial cells compared to untreated cells. In addition, activation of Fas significantly increased caspase-3 activity and apoptosis in CF epithelial cells compared to control cells. ConclusionsOverall, these results suggest that CF airway epithelial cells are more sensitive to apoptosis via increased levels of Fas and subsequent activation of the Fas death receptor pathway, which may be associated with dysfunctional CFTR.

AMPK alleviates endoplasmic reticulum stress by inducing the ER-chaperone ORP150 via FOXO1 to protect human bronchial cells from apoptosis

Chronic obstructive pulmonary disease (COPD), is characterized by inflammation of airways accompanied by a progressive destruction of lung parenchyma. This process is initiated in most cases by cigarette smoking. In this study we investigated the role of AMP activated protein kinase (AMPK) in cigarette smoke extract (CSE)-induced airway epithelial cell apoptosis as a consequence of endoplasmic reticulum stress (ER stress). Exposure of human bronchial epithelial cells (HBEpC) to CSE resulted in apoptosis as detected using Annexin V-PI flow cytometry. However, co-treatment with N1-(β-d-ribofuranosyl)-5-aminoimidazole-4-carboxamide (AICAR), a pharmacological activator of AMPK, significantly increased cell protection against ER stress-induced apoptosis by upregulating the 150 kDa oxygen-regulated protein (ORP150), which functions as an ER-associated chaperone, with concomitant elevation of FOXO1, a critical transcription factor regulating ORP150 expression. Lentiviral silencing of AMPK or FOXO1 using short hairpin (sh) RNA resulted in a significant decrease of ORP150 and an elevation of CCAAT/enhancer-binding protein-homologous protein (CHOP) resulting in ER stress and apoptosis of HBEpC. Together, our results strongly suggest that AMPK can activate ORP150 through FOXO1 pathway and confer protection against ER stress-induced apoptosis of airway epithelial cells following exposure to CSE. Thus, AMPK may serve as a likely therapeutic target for clinical and sub-clinical interventions in COPD.

MicroRNA-150 protects against cigarette smoke-induced lung inflammation and airway epithelial cell apoptosis through repressing p53: MicroRNA-150 in CS-induced lung inflammation.

Cigarette smoke (CS) exposure is an important risk factor for chronic obstructive pulmonary disease (COPD). MicroRNA-150 (miR-150) is involved in several inflammatory diseases. However, little is known about the role of miR-150 in the pathogenesis of COPD. In this study, we established a CS-related mouse model of COPD and evaluated the impact of miR-150 on CS-induced lung inflammation. We further investigated the effects of miR-150 overexpression on pro-inflammatory cytokine production and apoptosis in airway epithelial cells exposed to CS extract (CSE). It was found that miR-150 was significantly ( p < 0.05) downregulated in the lungs of CS-exposed mice, compared to control mice under normal air. The CSE-exposed BEAS-2B airway epithelial cells displayed a four- to six-fold reduction in miR-150 levels, compared to control cells ( p < 0.05). Delivery of miR-150 mimic attenuated CS-induced lung inflammation and accumulation of neutrophils, lymphocytes, and macrophages in bronchoalveolar lavage fluid. Moreover, miR-150 overexpression prevented the induction of interleukin-6, tumor necrosis factor alpha, and interleukin-8 expression and nuclear factor kappa B (NF-κB) transcriptional activity in BEAS-2B cells by CSE. Additionally, miR-150 protected BEAS-2B cells from CSE-induced apoptosis, which was associated with reduced p53 expression. Co-expression of p53 restored apoptotic response to CSE in miR-150-overexpressing BEAS-2B cells. Collectively, miR-150 suppresses CS-induced lung inflammation and airway epithelial cell apoptosis, which is causally linked to repression of p53 expression and NF-κB activity. Restoration of miR-150 expression may represent a potential therapeutic strategy for CS-related COPD.

Sinomenine ameliorates the airway remodelling, apoptosis of airway epithelial cells, and Th2 immune response in a murine model of chronic asthma

BackgroundSinomenine (SIN), an alkaloid isolated from the root of Sinomenium acutum which has a variety of pharmacological effects, including anti-inflammation, immunosuppression and anti-angiogenesis. The present study aimed to evaluate the effects of SIN on airway remodelling, epithelial apoptosis, and T Helper (Th)-2 derived cytokine levels in a murine model of chronic asthma. MethodsTwenty-two BALB/c mice were divided into four groups; I (control), II (placebo), III, IV. Mice in groups III and IV received the SIN (100mg/kg), and dexamethasone (1mg/kg) respectively. Epithelium thickness, sub-epithelial smooth muscle thickness, number of mast and goblet cells of samples isolated from the lung were measured. Immunohistochemical scorings of the lung tissue for matrix metalloproteinase-9 (MMP-9), vascular endothelial growth factor (VEG-F), transforming growth factor-beta (TGF-β), terminal deoxynucleotidyl transferase-mediated dUTP nick endlabeling (TUNEL) and cysteine-dependent aspartate-specific proteases (caspase)-3 were determined. IL-4, IL-5, IL-13, Nitric oxide in bronchoalveolar lavage fluid (BALF) and ovalbumin-specific immunoglobulin (Ig) E in serum were quantified by standard ELISA protocols. ResultsThe dose of 100mg/kg SIN treatment provided beneficial effects on all of the histopathological findings of airway remodelling compared to placebo (p<0.05). All cytokine levels in BALF and serum and immunohistochemical scores were significantly lower in 100mg/kg SIN treated group compared to the placebo (p<0.05). ConclusionsThese findings suggested that the dose of 100mg/kg SIN improved all histopathological changes of airway remodelling and its beneficial effects might be related to modulating Th-2 derived cytokines and the inhibition of apoptosis of airway epithelial cells.

Beneficial effects of ursodeoxycholic acid via inhibition of airway remodelling, apoptosis of airway epithelial cells, and Th2 immune response in murine model of chronic asthma

Background and aimsIn previous studies, anti-inflammatory, anti-apoptotic and immunomodulatory effects of ursodeoxycholic acid (UDCA) on liver diseases have been shown. In this study, we aimed to investigate the effects of UDCA on airway remodelling, epithelial apoptosis, and T Helper (Th)-2 derived cytokine levels in a murine model of chronic asthma. MethodsTwenty-seven BALB/c mice were divided into five groups; PBS-Control, OVA-Placebo, OVA-50mg/kg UDCA, OVA-150mg/kg UDCA, OVA-Dexamethasone. Mice in groups OVA-50mg/kg UDCA, OVA-150mg/kg UDCA, OVA-Dexamethasone received the UDCA (50mg/kg), UDCA (150mg/kg), and dexamethasone, respectively. Epithelium thickness, sub-epithelial smooth muscle thickness, number of mast and goblet cells of samples isolated from the lung were measured. Immunohistochemical scorings of the lung tissue for matrix metalloproteinase-9 (MMP-9), vascular endothelial growth factor (VEG-F), transforming growth factor-beta (TGF-β), terminal deoxynucleotidyl transferase-mediated dUTP nick endlabeling (TUNEL) and cysteine-dependent aspartate-specific proteases (caspase)-3 were determined. IL-4, IL-5, IL-13, Nitric oxide, ovalbumin-specific immunoglobulin (Ig) E levels were quantified. ResultsThe dose of 150mg/kg UDCA treatment led to lower epithelial thickness, sub-epithelial smooth muscle thickness, goblet and mast cell numbers compared to placebo. Except for MMP-9 and TUNEL all immunohistochemical scores were similar in both UDCA treated groups and the placebo. All cytokine levels were significantly lower in group IV compared to the placebo. ConclusionsThese findings suggested that the dose of 150mg/kg UDCA improved all histopathological changes of airway remodelling and its beneficial effects might be related to modulating Th-2 derived cytokines and the inhibition of apoptosis of airway epithelial cells.

Azithromycin ameliorates airway remodeling via inhibiting airway epithelium apoptosis

AimsAzithromycin can benefit treating allergic airway inflammation and remodeling. In the present study, we hypothesized that azithromycin alleviated airway epithelium injury through inhibiting airway epithelium apoptosis via down regulation of caspase-3 and Bax/Bcl2 ratio in vivo and in vitro. Main methodsOvalbumin induced rat asthma model and TGF-β1-induced BEAS-2B cell apoptosis model were established, respectively. In vivo experiments, airway epithelium was stained with hematoxylin and eosin (HE) and periodic acid–Schiff (PAS) to histologically evaluate the airway inflammation and remodeling. Airway epithelium apoptotic index (AI) was further analyzed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), while expression of apoptosis related gene (Bax, Bcl2, Caspase-3) in lungs were measured by qRT-PCR and western blotting, respectively. In vitro experiments, apoptosis were evaluated by Flow cytometry (FCM) and TUNEL. Above apoptosis related gene were also measured by qRT-PCR and western blotting. Key findingsCompared with the OVA group, azithromycin significantly reduced the inflammation score, peribronchial smooth muscle layer thickness, epithelial thickening and goblet cell metaplasia (P<0.05), and effectively suppressed AI of airway epithelium (P<0.05). Moreover, the increasing mRNA and protein expressions of Caspase-3 and Bax/Bcl-2 ratio in lung tissue were all significantly decreased in azithromycin-treated rats (P<0.05). In vitro, azithromycin significantly suppressed TGF-β1-induced BEAS-2B cells apoptosis (P<0.05) and reversed TGF-β1 elevated Caspase-3 mRNA level and Bax/Bcl-2 ratio (P<0.05). SignificanceAzithromycin is an attractive treatment option for reducing airway epithelial cell apoptosis by improving the imbalance of Bax/Bcl-2 ratio and inhibiting Caspase-3 level in airway epithelium.

Cigarette smoke extract induces placental growth factor release from human bronchial epithelial cells via ROS/MAPK (ERK-1/2)/Egr-1 axis.

Etiological evidence demonstrates that there is a significant association between cigarette smoking and chronic airway inflammatory disease. Abnormal expression of placental growth factor (PlGF) has been reported in COPD, and its downstream signaling molecules have been reported to contribute to the pathogenesis of airway epithelial cell apoptosis and emphysema. However, the signaling mechanisms underlying cigarette smoke extract (CSE)-induced PlGF expression in airway microenvironment remain unclear. Herein, we investigated the effects of reactive oxygen species (ROS)-dependent activation of the mitogen-activated protein kinase (MAPK) (extracellular signal-regulated kinase1/2 [ERK-1/2])/early growth response-1 (Egr-1) pathway on CSE-induced PlGF upregulation in human bronchial epithelium (HBE). The data obtained with quantitative reverse transcription polymerase chain reaction, Western blot, enzyme-linked immunosorbent assay (ELISA) and immunofluorescence staining analyses showed that CSE-induced Egr-1 activation was mainly mediated through production of ROS and activation of the MAPK (ERK-1/2) cascade. The binding of Egr-1 to the PlGF promoter was corroborated by an ELISA-based DNA binding activity assay. These results demonstrate that ROS activation of the MAPK (ERK-1/2)/Egr-1 pathway is a main player in the regulatory mechanism for CSE-induced PlGF production and that the use of an antioxidant could partly abolish these effects. Understanding the mechanisms of PlGF upregulation by CSE in the airway microenvironment may provide rational therapeutic interventions for cigarette smoking-related airway inflammatory diseases.

Region-specific role for Pten in maintenance of epithelial phenotype and integrity.

Previous studies have demonstrated resistance to naphthalene-induced injury in proximal airways of mice with lung epithelial-specific deletion of the tumor-suppressor gene Pten, attributed to increased proliferation of airway progenitors. We tested effects of Pten loss following bleomycin injury, a model typically used to study distal lung epithelial injury, in conditional PtenSFTPC-cre knockout mice. Pten-deficient airway epithelium exhibited marked hyperplasia, particularly in small bronchioles and at bronchoalveolar duct junctions, with reduced E-cadherin and β-catenin expression between cells toward the luminal aspect of the hyperplastic epithelium. Bronchiolar epithelial and alveolar epithelial type II (AT2) cells in PtenSFTPC-cre mice showed decreased expression of epithelial markers and increased expression of mesenchymal markers, suggesting at least partial epithelial-mesenchymal transition at baseline. Surprisingly, and in contrast to previous studies, mutant mice were exquisitely sensitive to bleomycin, manifesting rapid weight loss, respiratory distress, increased early mortality (by day 5), and reduced dynamic lung compliance. This was accompanied by sloughing of the hyperplastic airway epithelium with occlusion of small bronchioles by cellular debris, without evidence of increased parenchymal lung injury. Increased airway epithelial cell apoptosis due to loss of antioxidant defenses, reflected by decreased expression of superoxide dismutase 3, in combination with deficient intercellular adhesion, likely predisposed to airway sloughing in knockout mice. These findings demonstrate an important role for Pten in maintenance of airway epithelial phenotype integrity and indicate that responses to Pten deletion in respiratory epithelium following acute lung injury are highly context-dependent and region-specific.

TNF-α inhibits the growth of Legionella pneumophila in airway epithelial cells by inducing apoptosis.

TNF-α plays an important role in the pathogenesis of Legionella pneumophila (Lp)-induced pneumonia. Patients undergoing anti-TNF-α therapy are at an increased risk of Lp infection. Lp infects both phagocytic and non-phagocytic cells such as airway epithelial cells; however, the role of TNF-α in airway epithelial cells is unknown. Human airway epithelial cell line NCI-H292 was infected with Lp NUL1 strain. After infection, both intracellular growth of Lp and cell death were evaluated after treating the cells with or without TNF-α. Apoptosis was examined by performing activated caspase-3/7 staining and by using a pan-caspase inhibitor. Lp infected and replicated in NCI-H292 cells in a time-dependent manner, and TNF-α treatment of Lp-infected NCI-H292 cells inhibited Lp replication. Inhibitory effects of TNF-α on Lp replication were suppressed after treatment with a TNF-α-neutralizing antibody. Lp infection increased extracellular lactate dehydrogenase levels and decreased the number of living cells. Increased number of Lp-infected NCI-H292 cells showed caspase-3/7 activation, indicating they underwent apoptosis. TNF-α treatment inhibited Lp replication by increasing the apoptosis of NCI-H292 cells. Thus, our results suggested that airway epithelial cells were involved in the pathogenesis of Lp infection and that TNF-α played a protective role by inhibiting the intracellular replication of Lp and by increasing the apoptosis of Lp-infected airway epithelial cells. However, Lp infection should be investigated further in patients undergoing anti-TNF-α therapy who develop pneumonia.