Mechanosensitive channels in lung disease.

Mechanosensitive channels (MS channels) are membrane proteins capable of responding to mechanical stress over a wide dynamic range of external mechanical stimuli. In recent years, it has been found that MS channels play an important role as "sentinels" in the process of cell sensing and response to extracellular and intracellular force signals. There is growing appreciation for mechanical activation of ion channels and their subsequent initiation of downstream signaling pathways. Members of the transient receptor potential (TRP) superfamily and Piezo channels are broadly expressed in human tissues and contribute to multiple cellular functions. Both TRP and Piezo channels are thought to play key roles in physiological homeostasis and pathophysiology of disease states including in the lung. Here, we review the current state of knowledge on the expression, regulation, and function of TRP and Piezo channels in the context of the adult lung across the age spectrum, and in lung diseases such as asthma, COPD and pulmonary fibrosis where mechanical forces likely play varied roles in the structural and functional changes characteristic of these diseases. Understanding of TRP and Piezo in the lung can provide insights into new targets for treatment of pulmonary disease.

Interactions between calcium regulatory pathways and mechanosensitive channels in airways.

INTRODUCTION: Asthma is a chronic lung disease influenced by environmental and inflammatory triggers and involving complex signaling pathways across resident airway cells such as epithelium, airway smooth muscle, fibroblasts, and immune cells. While our understanding of asthma pathophysiology is continually progressing, there is a growing realization that cellular microdomains play critical roles in mediating signaling relevant to asthma in the context of contractility and remodeling. Mechanosensitive pathways are increasingly recognized as important to microdomain signaling, with Piezo and transient receptor protein (TRP) channels at the plasma membrane considered important for converting mechanical stimuli into cellular behavior. Given their ion channel properties, particularly Ca2+ conduction, a question becomes whether and how mechanosensitive channels contribute to Ca2+ microdomains in airway cells relevant to asthma. AREAS COVERED: Mechanosensitive TRP and Piezo channels regulate key Ca2+ regulatory proteins such as store operated calcium entry (SOCE) involving STIM and Orai channels, and sarcoendoplasmic (SR) mechanisms such as IP3 receptor channels (IP3Rs), and SR Ca2+ ATPase (SERCA) that are important in asthma pathophysiology including airway hyperreactivity and remodeling. EXPERT OPINION: Physical and/or functional interactions between Ca2+ regulatory proteins and mechanosensitive channels such as TRP and Piezo can toward understanding asthma pathophysiology and identifying novel therapeutic approaches.

Changes in the gut microbiome and metabolome in a rat model of pulmonary arterial hypertension.

The gut microbiota is widely considered to be involved in several diseases, including atherosclerosis, obesity, chronic obstructive pulmonary disease (COPD) and pulmonary arterial hypertension (PAH). This study aimed to determine if changes in the gut microbiome and metabolome play a major role in the early pathogenesis of PAH. Male Wistar rats were injected with monocrotaline (MCT) (55 mg/kg) at day 1 and injected with calcium-sensing receptor (CaSR) antagonist NPS2143 (4.5 mg/kg/d) from days 1 to 21. Fecal samples were obtained. The gut microbiota and metabolome were analyzed by 16S rRNA gene sequencing and mass spectrometry-based analysis to investigate the effect of PAH in this rat model. MCT injection had a marked effect on the composition of the gut microbiota. This finding was further confirmed by metabolomic analysis with identification of several metabolites relevant to the gut microflora. However, NPS2143 partially abrogated this intestinal flora disorder and reversed fecal metabolite abnormalities. In conclusion, our study shows correlations between changes in the gut microbiome and the metabolome in PAH, which are affected by NPS2143.

Long noncoding RNA IL6-AS1 is highly expressed in chronic obstructive pulmonary disease and is associated with interleukin 6 by targeting miR-149-5p and early B-cell factor 1.

Chronic obstructive pulmonary disease is a complex condition with multiple etiologies, including inflammation. We identified a novel long noncoding RNA (lncRNA), interleukin 6 antisense RNA 1 (IL6-AS1), which is upregulated in this disease and is associated with airway inflammation. We found that IL6-AS1 promotes the expression of inflammatory factors, especially interleukin (IL) 6. Mechanistically, cytoplasmic IL6-AS1 acts as an endogenous sponge by competitively binding to the microRNA miR-149-5p to stabilize IL-6 mRNA. Nuclear IL6-AS1 promotes IL-6 transcription by recruiting early B-cell factor 1 to the IL-6 promoter, which increases the methylation of the H3K4 histone and acetylation of the H3K27 histone. We propose a model of lncRNA expression in both the nucleus and cytoplasm that exerts similar effects through differing mechanisms, and IL6-AS1 probably increases inflammation via multiple pathways.

Long Noncoding RNA COPDA1 Promotes Airway Smooth Muscle Cell Proliferation in Chronic Obstructive Pulmonary Disease.

Abnormal expression of long noncoding RNAs (lncRNAs) has been confirmed to be associated with many diseases, including chronic obstructive pulmonary disease (COPD). To gain better understanding of the mechanism of COPD, we investigated the lncRNA and mRNA profiles in the lung tissue of patients with COPD. According to the analysis, one of the significantly different lncRNAs, COPDA1, might participate in the occurrence and development of COPD. Lung tissues were collected from nonsmokers, smokers, or smokers with COPD for RNA sequencing. Bioinformatic analysis and cell experiments were used to define the function of COPDA1, and the effects of COPDA1 on intracellular Ca2+ concentration and cell proliferation were examined after knockdown or overexpression of COPDA1. A number of variations of lncRNAs were found in the comparison of nonsmokers, smokers, and smokers with COPD. GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analyses indicated that smoking was involved in the activation of cytokines and the cell cycle, which is associated with COPD. According to the lncRNA-mRNA-coexpressing network and enrichment analysis, COPDAz1 and one of its target genes, MS4A1 (membrane-spanning 4-domains family, subfamily A) were investigated, and we discovered that the expression of MS4A1 was closely associated with lncRNA COPDA1 expression in human bronchial smooth muscle cells (HBSMCs). Further study showed that lncRNA COPDA1 upregulated the expression of MS4A1 to increase store-operated calcium entry in the HBSMCs, resulting in the promotion of the proliferation of smooth muscle cells as well as of airway remodeling. COPDA1 might be involved in the regulation of certain signaling pathways in COPD, might promote the proliferation of HBSMCs, and might also be involved in facilitating airway remodeling.

Tiotropium discontinuation in patients with early-stage COPD: a prospective observational cohort study.

BACKGROUND: Tiotropium improves lung function and ameliorates the annual decline in forced expiratory volume in 1 s (FEV1) after bronchodilator use in patients with mild to moderate chronic obstructive pulmonary disease (COPD). However, whether these benefits persist in patients with early-stage COPD after tiotropium discontinuation is unknown. METHODS: In this prospective cohort observational follow-up study, patients who had completed the Tiotropium in Early-Stage COPD (Tie-COPD) trial were followed for a maximum of 3 years, continuing or discontinuing treatment according to their willingness. The outcomes measured were spirometry parameters, COPD exacerbations, COPD Assessment Test (CAT) scores, Clinical COPD Questionnaire (CCQ) scores, modified Medical Research Council (mMRC) scores and the use of respiratory medications. RESULTS: Out of 376 patients, 262 (126 in the post-placebo group and 136 in the post-tiotropium group) completed the maximum 3-year follow-up after the study medication was withdrawn. After discontinuation, the decrease in FEV1 and forced vital capacity (FVC) did not differ significantly between the two groups, and neither did their annual decline. In addition, the frequency of acute COPD exacerbations and the mMRC scores were similar between the two groups after medication withdrawal. Both the mean CAT and CCQ scores were significantly lower in the post-tiotropium group than in the post-placebo group (p<0.05 for all comparisons) at the 1-year follow-up after withdrawal, but they were not different at the next follow-up. CONCLUSION: Withdrawal of tiotropium treatment in early-stage COPD resulted in difference reduction of both FEV1 and FVC, indicating that treatment should be continued.

Identification of abnormally expressed lncRNAs induced by PM2.5 in human bronchial epithelial cells.

To investigate the effect of stimulation of human bronchial epithelial cells (HBECs) by arterial traffic ambient PM2.5 (TAPM2.5) and wood smoke PM2.5 (WSPM2.5) on the expression of long non-coding RNAs (lncRNAs) in order to find new therapeutic targets for treatment of chronic obstructive pulmonary disease (COPD). HBECs were exposed to TAPM2.5 and WSPM2.5 at a series of concentrations. The microarray analysis was used to detect the lncRNA and mRNA expression profiles. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and gene ontology (GO) enrichment were conducted to analyze the differentially expressed lncRNAs and mRNAs. Quantitative real-time PCR (qRT-PCR) was performed to confirm the differential expression of lncRNAs. Western blot was performed to study the expression of autophagy and apoptosis-associated proteins. Flow cytometry was used to detect the apoptotic cells. The results indicated that fine particulate matter (PM2.5)-induced cell damage of HBECs occurred in a dose-dependent manner. The microarray analysis indicated that treatment with TAPM2.5 and WSPM2.5 led to the alteration of lncRNA and mRNA expression profiles. LncRNA maternally expressed gene 3 (MEG3) was significantly up-regulated in HBECs after PM2.5 treatment. The results of Western blot showed that PM2.5 induced cell apoptosis and autophagy by up-regulating apoptosis-associated gene, caspase-3, and down-regulating autophagy-associated markers, Bcl-2 and LC3 expression. In addition, we demonstrated that TAPM2.5 and WSPM2.5 accelerated apoptosis of human bronchial (HBE) cells, silencing of MEG3 suppressed apoptosis and autophagy of HBE cells. These findings suggested that the lncRNA MEG3 mediates PM2.5-induced cell apoptosis and autophagy, and probably through regulating the expression of p53.

PM2.5 promotes human bronchial smooth muscle cell migration via the sonic hedgehog signaling pathway.

BACKGROUND: The contribution of airway remodeling in chronic obstructive pulmonary disease (COPD) has been well documented, with airway smooth muscle cell proliferation and migration playing a role in the remodeling process. Here, we aimed to verify the effects of fine particulate matter (PM2.5) on human bronchial smooth muscle cell (HBSMC) migration and to explore the underlying signaling pathways. METHODS: HBSMC apoptosis, proliferation and migration were measured using flow cytometry, cell counting and transwell migration assays, respectively. The role of the hedgehog pathway in cell migration was assessed by western blotting to measure the expression of Sonic hedgehog (Shh), Gli1 and Snail. Furthermore, siRNA was used to knock down Gli1 or Snail expression. RESULTS: PM2.5 induced HBSMC apoptosis in a dose-dependent manner, although certain concentrations of PM2.5 did not induce HBSMC proliferation or apoptosis. Interestingly, cell migration was stimulated by PM2.5 doses far below those that induced apoptosis. Additional experiments revealed that these PM2.5 doses enhanced the expression of Shh, Gli1 and Snail in HBSMCs. Furthermore, PM2.5-induced cell migration and protein expression were enhanced by recombinant Shh and attenuated by cyclopamine. Similar results were obtained by knocking down Gli1 or Snail. CONCLUSIONS: These findings suggest that PM2.5, which may exert its effects through the Shh signaling pathway, is necessary for the migration of HBSMCs. These data define a novel role for PM2.5 in airway remodeling in COPD.

Nicotine-Induced Airway Smooth Muscle Cell Proliferation Involves TRPC6-Dependent Calcium Influx Via α7 nAChR.

BACKGROUND/AIMS: The proliferation of human bronchial smooth muscle cells (HBSMCs) is a key pathophysiological component of airway remodeling in chronic obstructive pulmonary disease (COPD) for which pharmacotherapy is limited, and only slight improvements in survival have been achieved in recent decades. Cigarette smoke is a well-recognized risk factor for COPD; however, the pathogenesis of cigarette smoke-induced COPD remains incompletely understood. This study aimed to investigate the mechanisms by which nicotine affects HBSMC proliferation. METHODS: Cell viability was assessed with a CCK-8 assay. Proliferation was measured by cell counting and EdU immunostaining. Fluorescence calcium imaging was performed to measure intracellular Ca2+ concentration ([Ca2+]i). RESULTS: The results showed that nicotine promotes HBSMC proliferation, which is accompanied by elevated store-operated calcium entry (SOCE), receptor-operated calcium entry (ROCE) and basal [Ca2+]i in HBSMCs. Moreover, we also confirmed that canonical transient receptor potential protein 6 (TRPC6) and α7 nicotinic acetylcholine receptor (α7 nAChR) are involved in nicotine-induced upregulation of cell proliferation. Furthermore, we verified that activation of the PI3K/Akt signaling pathway plays a pivotal role in nicotine-enhanced proliferation and calcium influx in HBSMCs. Inhibition of α7 nAChR significantly decreased Akt phosphorylation levels, and LY294002 inhibited the protein expression levels of TRPC6. CONCLUSION: Herein, these data provide compelling evidence that calcium entry via the α7 nAChR-PI3K/Akt-TRPC6 signaling pathway plays an important role in the physiological regulation of airway smooth muscle cell proliferation, representing an important target for augmenting airway remodeling.

Exposure to Ambient Particulate Matter Induced COPD in a Rat Model and a Description of the Underlying Mechanism.

While the health effects of air pollution have been an international public health concern since at least the 1950s, recent research has focused on two broad sources of air pollution, namely, biomass fuel (BMF) and motor vehicle exhaust (MVE). Many studies have shown associations between air pollution PM and exacerbations of pre-existing COPD, but the role of air pollution PM in the development and progression of COPD is still uncertain. The current study indicates that rats can develop pronounced COPD following chronic exposure to air pollution PM (BMF and MVE), as characterized by lung function reduction, mucus metaplasia, lung and systemic inflammation, emphysema, and small airway remodeling. Comparative analyses demonstrate that both BMF and MVE activate similar pathogenesis that are linked to the development of COPD. These findings also show that some differences are found in the lungs of rats exposed to BMF or MVE, which might result in different phenotypes of COPD.