Fibroblast-derived Extracellular Vesicles Induce Lung Cancer Progression in the Idiopathic Pulmonary Fibrosis Microenvironment.

Idiopathic pulmonary fibrosis (IPF) is a progressive aging-related lung disease associated with increased lung cancer risk. Although previous studies have shown that IPF worsens the survival of patients with lung cancer, whether IPF independently affects cancer malignancy and prognosis remains inconclusive. Extracellular vesicles (EVs) have recently emerged as active carriers of molecular biomarkers and mediators of intercellular communication in lung homeostasis and pathogenesis. EV cargo-mediated fibroblast-tumor cell communication might participate in the development and progression of lung cancer by modulating various signaling pathways. In this study, we examined the impact of lung fibroblast (LF)-derived EVs on non-small cell lung cancer (NSCLC) malignancy in the IPF microenvironment. Here, we showed that LFs derived from patients with IPF have phenotypes of myofibroblast differentiation and cellular senescence. Furthermore, we found that IPF LF-derived EVs have markedly altered microRNA compositions and exert proproliferative functions on NSCLC cells. Mechanistically, the phenotype was attributed mainly to the enrichment of miR-19a in IPF LF-derived EVs. As a downstream signaling pathway, mir-19a in IPF LF-derived EVs regulates ZMYND11-mediated c-Myc activation in NSCLC, potentially contributing to the poor prognosis of patients with NSCLC with IPF. Our discoveries provide novel mechanistic insights for understanding lung cancer progression in the IPF microenvironment. Accordingly, blocking the secretion of IPF LF-derived EV miR-19a and their signaling pathways is a potential therapeutic strategy for managing IPF and lung cancer progression.

Impaired TRIM16-Mediated Lysophagy in Chronic Obstructive Pulmonary Disease Pathogenesis.

Insufficient autophagic degradation has been implicated in accelerated cellular senescence during chronic obstructive pulmonary disease (COPD) pathogenesis. Aging-linked and cigarette smoke (CS)-induced functional deterioration of lysosomes may be associated with impaired autophagy. Lysosomal membrane permeabilization (LMP) is indicative of damaged lysosomes. Galectin-3 and tripartite motif protein (TRIM) 16 play a cooperative role in recognizing LMP and inducing lysophagy, a lysosome-selective autophagy, to maintain lysosome function. In this study, we sought to examine the role of TRIM16-mediated lysophagy in regulating CS-induced LMP and cellular senescence during COPD pathogenesis by using human bronchial epithelial cells and lung tissues. CS extract (CSE) induced lysosomal damage via LMP, as detected by galectin-3 accumulation. Autophagy was responsible for modulating LMP and lysosome function during CSE exposure. TRIM16 was involved in CSE-induced lysophagy, with impaired lysophagy associated with lysosomal dysfunction and accelerated cellular senescence. Airway epithelial cells in COPD lungs showed an increase in lipofuscin, aggresome and galectin-3 puncta, reflecting accumulation of lysosomal damage with concomitantly reduced TRIM16 expression levels. Human bronchial epithelial cells isolated from COPD patients showed reduced TRIM16 but increased galectin-3, and a negative correlation between TRIM16 and galectin-3 protein levels was demonstrated. Damaged lysosomes with LMP are accumulated in epithelial cells in COPD lungs, which can be at least partly attributed to impaired TRIM16-mediated lysophagy. Increased LMP in lung epithelial cells may be responsible for COPD pathogenesis through the enhancement of cellular senescence.

Chaperone-Mediated Autophagy Suppresses Apoptosis via Regulation of the Unfolded Protein Response during Chronic Obstructive Pulmonary Disease Pathogenesis.

Cigarette smoke (CS) induces accumulation of misfolded proteins with concomitantly enhanced unfolded protein response (UPR). Increased apoptosis linked to UPR has been demonstrated in chronic obstructive pulmonary disease (COPD) pathogenesis. Chaperone-mediated autophagy (CMA) is a type of selective autophagy for lysosomal degradation of proteins with the KFERQ peptide motif. CMA has been implicated in not only maintaining nutritional homeostasis but also adapting the cell to stressed conditions. Although recent papers have shown functional cross-talk between UPR and CMA, mechanistic implications for CMA in COPD pathogenesis, especially in association with CS-evoked UPR, remain obscure. In this study, we sought to examine the role of CMA in regulating CS-induced apoptosis linked to UPR during COPD pathogenesis using human bronchial epithelial cells (HBEC) and lung tissues. CS extract (CSE) induced LAMP2A expression and CMA activation through a Nrf2-dependent manner in HBEC. LAMP2A knockdown and the subsequent CMA inhibition enhanced UPR, including CHOP expression, and was accompanied by increased apoptosis during CSE exposure, which was reversed by LAMP2A overexpression. Immunohistochemistry showed that Nrf2 and LAMP2A levels were reduced in small airway epithelial cells in COPD compared with non-COPD lungs. Both Nrf2 and LAMP2A levels were significantly reduced in HBEC isolated from COPD, whereas LAMP2A levels in HBEC were positively correlated with pulmonary function tests. These findings suggest the existence of functional cross-talk between CMA and UPR during CSE exposure and also that impaired CMA may be causally associated with COPD pathogenesis through enhanced UPR-mediated apoptosis in epithelial cells.

Real-life effectiveness of dupilumab in patients with mild to moderate bronchial asthma comorbid with CRSwNP.

BACKGROUND: Dupilumab, an anti-IL-4α receptor antibody, is a new treatment for severe or refractory asthma. However, real-world evidence on the efficacy of dupilumab in patients with mild to moderate bronchial asthma is lacking. METHODS: We retrospectively evaluated the effects of dupilumab in 62 patients who received dupilumab for eosinophilic sinusitis comorbid with asthma at a single centre in Japan. Type 2 inflammatory markers, ACT, respiratory function tests, and forced oscillation technique (FOT) were analysed before, three months after, and one year after dupilumab administration, mainly in patients with mild to moderate asthma. RESULTS: FEV1, %FEV1, %FVC, treatment steps for asthma and ACT improved significantly after three months of dupilumab treatment. FeNO was markedly decreased, whereas IgE and eosinophil counts showed no significant changes. Pre- and post-treatment respiratory resistance (Rrs) and respiratory reactance (Xrs) correlated significantly with FEV1. Improvement in %FEV1 was associated with higher FeNO and higher serum IgE before dupilumab treatment. CONCLUSION: Dupilumab treatment for sinusitis may improve respiratory functions, asthma symptoms, and asthma treatment reduction, even if the associated bronchial asthma is not severe.

Involvement of GPx4-Regulated Lipid Peroxidation in Idiopathic Pulmonary Fibrosis Pathogenesis.

The imbalanced redox status in lung has been widely implicated in idiopathic pulmonary fibrosis (IPF) pathogenesis. To regulate redox status, hydrogen peroxide must be adequately reduced to water by glutathione peroxidases (GPx). Among GPx isoforms, GPx4 is a unique antioxidant enzyme that can directly reduce phospholipid hydroperoxide. Increased lipid peroxidation products have been demonstrated in IPF lungs, suggesting the participation of imbalanced lipid peroxidation in IPF pathogenesis, which can be modulated by GPx4. In this study, we sought to examine the involvement of GPx4-modulated lipid peroxidation in regulating TGF-ß-induced myofibroblast differentiation. Bleomycin-induced lung fibrosis development in mouse models with genetic manipulation of GPx4 were examined. Immunohistochemical evaluations for GPx4 and lipid peroxidation were performed in IPF lung tissues. Immunohistochemical evaluations showed reduced GPx4 expression levels accompanied by increased 4-hydroxy-2-nonenal in fibroblastic focus in IPF lungs. TGF-ß-induced myofibroblast differentiation was enhanced by GPx4 knockdown with concomitantly enhanced lipid peroxidation and SMAD2/SMAD3 signaling. Heterozygous GPx4-deficient mice showed enhancement of bleomycin-induced lung fibrosis, which was attenuated in GPx4-transgenic mice in association with lipid peroxidation and SMAD signaling. Regulating lipid peroxidation by Trolox showed efficient attenuation of bleomycin-induced lung fibrosis development. These findings suggest that increased lipid peroxidation resulting from reduced GPx4 expression levels may be causally associated with lung fibrosis development through enhanced TGF-ß signaling linked to myofibroblast accumulation of fibroblastic focus formation during IPF pathogenesis. It is likely that regulating lipid peroxidation caused by reduced GPx4 can be a promising target for an antifibrotic modality of treatment for IPF.

Involvement of Lamin B1 Reduction in Accelerated Cellular Senescence during Chronic Obstructive Pulmonary Disease Pathogenesis.

Downregulation of lamin B1 has been recognized as a crucial step for development of full senescence. Accelerated cellular senescence linked to mechanistic target of rapamycin kinase (MTOR) signaling and accumulation of mitochondrial damage has been implicated in chronic obstructive pulmonary disease (COPD) pathogenesis. We hypothesized that lamin B1 protein levels are reduced in COPD lungs, contributing to the process of cigarette smoke (CS)-induced cellular senescence via dysregulation of MTOR and mitochondrial integrity. To illuminate the role of lamin B1 in COPD pathogenesis, lamin B1 protein levels, MTOR activation, mitochondrial mass, and cellular senescence were evaluated in CS extract (CSE)-treated human bronchial epithelial cells (HBEC), CS-exposed mice, and COPD lungs. We showed that lamin B1 was reduced by exposure to CSE and that autophagy was responsible for lamin B1 degradation in HBEC. Lamin B1 reduction was linked to MTOR activation through DEP domain-containing MTOR-interacting protein (DEPTOR) downregulation, resulting in accelerated cellular senescence. Aberrant MTOR activation was associated with increased mitochondrial mass, which can be attributed to peroxisome proliferator-activated receptor γ coactivator-1ß-mediated mitochondrial biogenesis. CS-exposed mouse lungs and COPD lungs also showed reduced lamin B1 and DEPTOR protein levels, along with MTOR activation accompanied by increased mitochondrial mass and cellular senescence. Antidiabetic metformin prevented CSE-induced HBEC senescence and mitochondrial accumulation via increased DEPTOR expression. These findings suggest that lamin B1 reduction is not only a hallmark of lung aging but is also involved in the progression of cellular senescence during COPD pathogenesis through aberrant MTOR signaling.

Regulated Necrosis in Pulmonary Disease. A Focus on Necroptosis and Ferroptosis.

To date, increasing evidence suggests the possible involvement of various types of cell death in lung diseases. The recognized regulated cell death includes necrotic cell death that is immunogenic, releasing damage-associated molecular patterns and driving tissue inflammation. Necroptosis is a well-understood form of regulated necrosis that is executed by RIPK3 (receptor-interacting protein kinase 3) and the pseudokinase MLKL (mixed lineage kinase domain-like protein). Ferroptosis is a newly described caspase-independent form of regulated necrosis that is characterized by the increase of detrimental lipid reactive oxygen species produced via iron-dependent lipid peroxidation. The role of these two cell death pathways differs depending on the disease, cell type, and microenvironment. Moreover, some experimental cell death models have demonstrated shared ferroptotic and necroptotic cell death and the synergistic effect of simultaneous inhibition. This review examines the role of regulated necrotic cell death, particularly necroptosis and ferroptosis, in lung disease pathogenesis in the context of recent insights into the roles of the key effector molecules of these two cell death pathways.

Extracellular Vesicles from Fibroblasts Induce Epithelial-Cell Senescence in Pulmonary Fibrosis.

Aberrant epithelial-mesenchymal interactions have critical roles in regulating fibrosis development. The involvement of extracellular vesicles (EVs), including exosomes, remains to be elucidated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Here, we found that lung fibroblasts (LFs) from patients with IPF induce cellular senescence via EV-mediated transfer of pathogenic cargo to lung epithelial cells. Mechanistically, IPF LF-derived EVs increased mitochondrial reactive oxygen species and associated mitochondrial damage in lung epithelial cells, leading to activation of the DNA damage response and subsequent epithelial-cell senescence. We showed that IPF LF-derived EVs contain elevated levels of microRNA-23b-3p (miR-23b-3p) and miR-494-3p, which suppress SIRT3, resulting in the epithelial EV-induced phenotypic changes. Furthermore, the levels of miR-23b-3p and miR-494-3p found in IPF LF-derived EVs correlated positively with IPF disease severity. These findings reveal that the accelerated epithelial-cell mitochondrial damage and senescence observed during IPF pathogenesis are caused by a novel paracrine effect of IPF fibroblasts via microRNA-containing EVs.

Chaperone-mediated autophagy receptor modulates tumor growth and chemoresistance in non-small cell lung cancer.

Chaperone-mediated autophagy (CMA) is a lysosomal degradation pathway of selective soluble proteins. Lysosome-associated membrane protein type 2a (LAMP2A) is the key receptor protein of CMA; downregulation of LAMP2A leads to CMA blockade. Although CMA activation has been involved in cancer growth, CMA status and functions in non-small cell lung cancer (NSCLC) by focusing on the roles in regulating chemosensitivity remain to be clarified. In this study, we found that LAMP2A expression is elevated in NSCLC cell lines and patient's tumors, conferring poor survival and platinum resistance in NSCLC patients. LAMP2A knockdown in NSCLC cells suppressed cell proliferation and colony formation and increased the sensitivity to chemotherapeutic drugs in vitro. Furthermore, we found that intrinsic apoptosis signaling is the mechanism of cell death involved with CMA blockade. Remarkably, LAMP2A knockdown repressed tumorigenicity and sensitized the tumors to cisplatin treatment in NSCLC-bearing mice. Our discoveries suggest that LAMP2A is involved in the regulation of cancer malignant phenotypes and represents a promising new target against chemoresistant NSCLC.

Successful treatment of steroid-refractory immune checkpoint inhibitor-related pneumonitis with triple combination therapy: a case report.

Immune checkpoint inhibitor (ICI)-related pneumonitis is a relatively rare but clinically serious and potentially life-threatening adverse event. The majority of cases can be managed by drug discontinuation, with the administration of corticosteroids added in severe cases. However, worsening of pneumonitis can develop in a subset of patients despite treatment with high doses of corticosteroids. We herein report a case of steroid-refractory ICI-related pneumonitis in a recurrent non-small cell lung cancer (NSCLC) patient treated with pembrolizumab that was successfully improved by triple combination therapy (high-dose corticosteroids, tacrolimus, and cyclophosphamide). After 3 weeks of initial pembrolizumab administration, the patient was diagnosed with ICI-related pneumonitis. Chest computed tomography (CT) showed patchy distributed bilateral consolidation and ground-glass opacities (GGOs) with traction bronchiectasis and bronchiolectasis resembling the diffuse alveolar damage (DAD) radiographic pattern. Although methylprednisolone pulse therapy was initiated, worsening of respiratory failure resulted in the patient being transferred to the intensive care unit. Because of an insufficient therapeutic response to high-dose corticosteroids, tacrolimus and cyclophosphamide pulse therapy were additively performed as triple combination therapy according to the treatment strategy for pulmonary complications of clinically amyopathic dermatomyositis (CADM). In response to this triple combination therapy, the patient's respiratory condition gradually improved, and chest CT showed the marked amelioration of pulmonary opacities. This is the first report suggesting the efficacy of triple combination therapy (high-dose corticosteroids, tacrolimus, and cyclophosphamide) for steroid-refractory ICI-related pneumonitis complicated with respiratory failure.

Efficacy of benralizumab for patients with severe eosinophilic asthma: a retrospective, real-life study.

BACKGROUND: Benralizumab, an anti-interleukin-5 (IL-5) receptor α monoclonal antibody, significantly reduces the number of annual exacerbations and oral corticosteroid (OCS) maintenance doses for patients with severe eosinophilic asthma (SEA). However, few studies on the efficacy of this biologic in real life are available. The aim was to elucidate the efficacy of benralizumab by evaluating changes in clinical parameters after benralizumab treatment in patients with SEA. METHODS: From July 2018 to December 2019, 24 Japanese patients with SEA received benralizumab at Jikei University Hospital. We retrospectively evaluated the patients' characteristics, parameters, numbers of exacerbations and maintenance OCS doses. RESULTS: Among the 24 patients, eleven patients had received mepolizumab treatment and were directly switched to benralizumab. The peripheral blood eosinophil and basophil counts significantly decreased after benralizumab treatment regardless of previous mepolizumab treatment. Pulmonary function, Asthma Control Test scores, the numbers of annual exacerbations and maintenance OCS doses in patients without previous mepolizumab treatment tended to improve without significant differences. Fourteen patients (58%) were responders according to the Global Evaluation of Treatment Effectiveness (GETE) score. The proportion of GETE responders among patients with aspirin-exacerbated respiratory disease (AERD) tended to be lower than that among patients without AERD (p = 0.085). After benralizumab treatment, the change in the forced expiratory volume in 1 s from baseline was 200 ml or greater in eight patients (33%), including three patients who were switched from mepolizumab. CONCLUSION: Benralizumab treatment improved and controlled asthma symptoms based on the GETE score.

Efficacy of mepolizumab for patients with severe asthma and eosinophilic chronic rhinosinusitis.

BACKGROUND: Several major randomized control studies have demonstrated that mepolizumab, an anti-IL-5 monoclonal antibody, is effective for patients with severe eosinophilic asthma who show exacerbation or require systemic corticosteroid maintenance therapy. However, the predictive factors of the response to mepolizumab other than blood eosinophil count are unclear in clinical practice. OBJECTIVE: To elucidate the predictive factors of the response to mepolizumab for patients with severe eosinophilic asthma. METHODS: From July 2016 to December 2017, 28 patients with severe asthma received mepolizumab in our hospital. To determine the predictive factors, we retrospectively evaluated patient characteristics, comorbidities, biomarkers, pulmonary function, maintenance dose of systemic corticosteroids and number of exacerbations. RESULTS: The response rate to mepolizumab treatment was 70% (19/27; one pregnant woman was excluded from analysis). Compared with 11 patients without eosinophilic chronic rhinosinusitis (ECRS), 16 patients with ECRS showed significantly improved systemic corticosteroid-sparing effects [- 71.3 ± 37.0% vs - 10.7 ± 20.1%, P = 0.006], change from baseline FeNO [- 19 ± 57 (%) vs 30 ± 77 (%), P = 0.023] and symptoms [14 patients (88%) vs five patients (45%), P = 0.033]. ECRS was identified as a predictive factor of the response to mepolizumab in a multivariate logistic regression analysis [odds ratio = 22.5, 95% CI (1.5-336), P = 0.024]. Of the eight patients previously administered omalizumab, five responded to mepolizumab. Staphylococcus aureus enterotoxin B IgE results were negative in 80% of responders (P = 0.14). CONCLUSION: Both groups showed improved symptom scores and a decreased number of exacerbations. Mepolizumab substantially improved the clinical variables of patients with eosinophilic asthma complicated with ECRS.

Involvement of Alveolar Epithelial Cell Necroptosis in Idiopathic Pulmonary Fibrosis Pathogenesis.

Alveolar epithelial cell (AEC) injury leading to cell death is involved in the process of fibrosis development during idiopathic pulmonary fibrosis (IPF). Among regulated/programmed cell death, the excessive apoptosis of AECs has been widely implicated in IPF pathogenesis. Necroptosis is a type of regulated/programmed necrosis. A multiprotein complex composed of receptor-interacting protein kinase (RIPK)-1 and -3 plays a key regulatory role in initiating necroptosis. Although necroptosis participates in disease pathogeneses through the release of damage-associated molecular patterns, its association with IPF progression remains elusive. In this study, we attempted to illuminate the involvement of RIPK3-regulated necroptosis in IPF pathogenesis. IPF lung tissues were used to detect necroptosis, and the role of RIPK3 was determined using cell culturing models of AECs. Lung fibrosis models of bleomycin (BLM) treatment were also used. RIPK3 expression levels were increased in IPF lungs, and both apoptosis and necroptosis were detected mainly in AECs. Necrostatin-1 and RIPK3 knockout experiments in AECs revealed the participation of necroptosis in BLM and hydrogen peroxide-induced cell death. BLM treatment induced RIPK3 expression in AECs and increased high-mobility group box 1 and IL-1ß levels in mouse lungs. The efficient attenuation of BLM-induced lung inflammation and fibrosis was determined in RIPK3 knockout mice and by necrostatin-1 with a concomitant reduction in high-mobility group box 1 and IL-1ß. RIPK3-regulated necroptosis in AECs is involved in the mechanism of lung fibrosis development through the release of damage-associated molecular patterns as part of the pathogenic sequence of IPF.

Involvement of PARK2-Mediated Mitophagy in Idiopathic Pulmonary Fibrosis Pathogenesis.

Fibroblastic foci, known to be the leading edge of fibrosis development in idiopathic pulmonary fibrosis (IPF), are composed of fibrogenic myofibroblasts. Autophagy has been implicated in the regulation of myofibroblast differentiation. Insufficient mitophagy, the mitochondria-selective autophagy, results in increased reactive oxygen species, which may modulate cell signaling pathways for myofibroblast differentiation. Therefore, we sought to investigate the regulatory role of mitophagy in myofibroblast differentiation as a part of IPF pathogenesis. Lung fibroblasts were used in in vitro experiments. Immunohistochemical evaluation in IPF lung tissues was performed. PARK2 was examined as a target molecule for mitophagy regulation, and a PARK2 knockout mouse was employed in a bleomycin-induced lung fibrosis model. We demonstrated that PARK2 knockdown-mediated mitophagy inhibition was involved in the mechanism for activation of the platelet-derived growth factor receptor (PDGFR)/PI3K/AKT signaling pathway accompanied by enhanced myofibroblast differentiation and proliferation, which were clearly inhibited by treatment with both antioxidants and AG1296, a PDGFR inhibitor. Mitophagy inhibition-mediated activation of PDGFR signaling was responsible for further autophagy suppression, suggesting the existence of a self-amplifying loop of mitophagy inhibition and PDGFR activation. IPF lung demonstrated reduced PARK2 with concomitantly increased PDGFR phosphorylation. Furthermore, bleomycin-induced lung fibrosis was enhanced in PARK2 knockout mice and subsequently inhibited by AG1296. These findings suggest that insufficient mitophagy-mediated PDGFR/PI3K/AKT activation, which is mainly attributed to reduced PARK2 expression, is a potent underlying mechanism for myofibroblast differentiation and proliferation in fibroblastic foci formation during IPF pathogenesis.

Risk factors of postoperative pulmonary complications in patients with asthma and COPD.

BACKGROUND: Postoperative pulmonary complications (PPC) in patients with pulmonary diseases remain to be resolved clinical issue. However, most evidence regarding PPC has been established more than 10 years ago. Therefore, it is necessary to evaluate perioperative management using new inhalant drugs in patients with obstructive pulmonary diseases. METHODS: April 2014 through March 2015, 346 adult patients with pulmonary diseases (257 asthma, 89 chronic obstructive pulmonary disease (COPD)) underwent non-pulmonary surgery except cataract surgery in our university hospital. To analyze the risk factors for PPC, we retrospectively evaluated physiological backgrounds, surgical factors and perioperative specific treatment for asthma and COPD. RESULTS: Finally, 29 patients with pulmonary diseases (22 asthma, 7 COPD) had PPC. In patients with asthma, smoking index (≥ 20 pack-years), peripheral blood eosinophil count (≥ 200/mm3) and severity (Global INitiative for Asthma(GINA) STEP ≥ 3) were significantly associated with PPC in the multivariate logistic regression analysis [odds ratio (95% confidence interval) = 5.4(1.4-20.8), 0.31 (0.11-0.84) and 3.2 (1.04-9.9), respectively]. In patients with COPD, age, introducing treatment for COPD, upper abdominal surgery and operation time (≥ 5 h) were significantly associated with PPC [1.18 (1.00-1.40), 0.09 (0.01-0.81), 21.2 (1.3-349) and 9.5 (1.2-77.4), respectively]. CONCLUSIONS: History of smoking or severe asthma is a risk factor of PPC in patients with asthma, and age, upper abdominal surgery, or long operation time is a risk factor of PPC in patients with COPD. Adequate inhaled corticosteroids treatment in patients with eosinophilic asthma and introducing treatment for COPD in patients with COPD could reduce PPCs.

Role of IL-17A in murine models of COPD airway disease.

Small airway fibrosis is a major pathological feature of chronic obstructive pulmonary disease (COPD) and is refractory to current treatments. Chronic inflammatory cells accumulate around small airways in COPD and are thought to play a major role in small airway fibrosis. Mice deficient in α/ß T cells have recently been shown to be protected from both experimental airway inflammation and fibrosis. In these models, CD4+Th17 cells and secretion of IL-17A are increased. However, a pathogenic role for IL-17 in specifically mediating fibrosis around airways has not been demonstrated. Here a role for IL-17A in airway fibrosis was demonstrated using mice deficient in the IL-17 receptor A (il17ra) Il17ra-deficient mice were protected from both airway inflammation and fibrosis in two different models of airway fibrosis that employ COPD-relevant stimuli. In these models, CD4+ Th17 are a major source of IL-17A with other expressing cell types including γδ T cells, type 3 innate lymphoid cells, polymorphonuclear cells, and CD8+ T cells. Antibody neutralization of IL-17RA or IL-17A confirmed that IL-17A was the relevant pathogenic IL-17 isoform and IL-17RA was the relevant receptor in airway inflammation and fibrosis. These results demonstrate that the IL-17A/IL-17 RA axis is crucial to murine airway fibrosis. These findings suggest that IL-17 might be targeted to prevent the progression of airway fibrosis in COPD.

Pirfenidone inhibits myofibroblast differentiation and lung fibrosis development during insufficient mitophagy.

BACKGROUND: Pirfenidone (PFD) is an anti-fibrotic agent used to treat idiopathic pulmonary fibrosis (IPF), but its precise mechanism of action remains elusive. Accumulation of profibrotic myofibroblasts is a crucial process for fibrotic remodeling in IPF. Recent findings show participation of autophagy/mitophagy, part of the lysosomal degradation machinery, in IPF pathogenesis. Mitophagy has been implicated in myofibroblast differentiation through regulating mitochondrial reactive oxygen species (ROS)-mediated platelet-derived growth factor receptor (PDGFR) activation. In this study, the effect of PFD on autophagy/mitophagy activation in lung fibroblasts (LF) was evaluated, specifically the anti-fibrotic property of PFD for modulation of myofibroblast differentiation during insufficient mitophagy. METHODS: Transforming growth factor-ß (TGF-ß)-induced or ATG5, ATG7, and PARK2 knockdown-mediated myofibroblast differentiation in LF were used for in vitro models. The anti-fibrotic role of PFD was examined in a bleomycin (BLM)-induced lung fibrosis model using PARK2 knockout (KO) mice. RESULTS: We found that PFD induced autophagy/mitophagy activation via enhanced PARK2 expression, which was partly involved in the inhibition of myofibroblast differentiation in the presence of TGF-ß. PFD inhibited the myofibroblast differentiation induced by PARK2 knockdown by reducing mitochondrial ROS and PDGFR-PI3K-Akt activation. BLM-treated PARK2 KO mice demonstrated augmentation of lung fibrosis and oxidative modifications compared to those of BLM-treated wild type mice, which were efficiently attenuated by PFD. CONCLUSIONS: These results suggest that PFD induces PARK2-mediated mitophagy and also inhibits lung fibrosis development in the setting of insufficient mitophagy, which may at least partly explain the anti-fibrotic mechanisms of PFD for IPF treatment.

A critical role for dendritic cells in the evolution of IL-1ß-mediated murine airway disease.

Chronic airway inflammation and fibrosis, known as airway remodeling, are defining features of chronic obstructive pulmonary disease and are refractory to current treatments. How and whether chronic inflammation contributes to airway fibrosis remain controversial. In this study, we use a model of chronic obstructive pulmonary disease airway disease utilizing adenoviral delivery of IL-1ß to determine that adaptive T cell immunity is required for airway remodeling because mice deficient in α/ß T cells (tcra(-/-)) are protected. Dendritic cells (DCs) accumulate around chronic obstructive pulmonary disease airways and are critical to prime adaptive immunity, but they have not been shown to directly influence airway remodeling. We show that DC depletion or deficiency in the crucial DC chemokine receptor ccr6 both protect from adenoviral IL-1ß-induced airway adaptive T cell immune responses and fibrosis in mice. These results provide evidence that chronic airway inflammation, mediated by accumulation of α/ß T cells and driven by DCs, is critical to airway fibrosis.

TGF-ß-Dependent Dendritic Cell Chemokinesis in Murine Models of Airway Disease.

Small airway chronic inflammation is a major pathologic feature of chronic obstructive pulmonary disease (COPD) and is refractory to current treatments. Dendritic cells (DCs) accumulate around small airways in COPD. DCs are critical mediators of Ag surveillance and Ag presentation and amplify adaptive immune responses. How DCs accumulate around airways remains largely unknown. We use 2-photon DC imaging of living murine lung sections to directly visualize the dynamic movement of living DCs around airways in response to either soluble mediators (IL-1ß) or environmental stimuli (cigarette smoke or TLR3 ligands) implicated in COPD pathogenesis. We find that DCs accumulate around murine airways primarily by increasing velocity (chemokinesis) rather than directional migration (chemotaxis) in response to all three stimuli. DC accumulation maximally occurs in a specific zone located 26-50 µm from small airways, which overlaps with zones of maximal DC velocity. Our data suggest that increased accumulation of DCs around airways results from increased numbers of highly chemokinetic DCs entering the lung from the circulation with balanced rates of immigration and emigration. Increases in DC accumulation and chemokinesis are partially dependent on ccr6, a crucial DC chemokine receptor, and fibroblast expression of the integrin αvß8, a critical activator of TGF-ß. αvß8-Mediated TGF-ß activation is known to enhance IL-1ß-dependent fibroblast expression of the only known endogenous ccr6 chemokine ligand, ccl20. Taken together, these data suggest a mechanism by which αvß8, ccl20, and ccr6 interact to lead to DC accumulation around airways in response to COPD-relevant stimuli.

Detection of pathogens by real-time PCR in adult patients with acute exacerbation of bronchial asthma.

BACKGROUND: Respiratory tract infection is a major cause of acute exacerbation of bronchial asthma (AEBA). Although recent findings suggest that common bacteria are causally associated with AEBA, a comprehensive epidemiologic analysis of infectious pathogens including common/atypical bacteria and viruses in AEBA has not been performed. Accordingly, we attempted to detect pathogens during AEBA by using real-time polymerase chain reaction (PCR) in comparison to conventional methods. METHODS: We prospectively enroled adult patients with AEBA from August 2012 to March 2014. Infectious pathogens collected in nasopharyngeal swab and sputum samples were examined in each patient by conventional methods and real-time PCR, which can detect 6 bacterial and 11 viral pathogens. The causal association of these pathogens with AEBA severity and their frequency of monthly distribution were also examined. RESULTS: Among the 64 enroled patients, infectious pathogens were detected in 49 patients (76.6%) using real-time PCR and in 14 patients (21.9%) using conventional methods (p < 0.001). Real-time PCR detected bacteria in 29 patients (45.3%) and respiratory viruses in 28 patients (43.8%). Haemophilus influenzae was the most frequently detected microorganism (26.6%), followed by rhinovirus (15.6%). Influenza virus was the significant pathogen associated with severe AEBA. Moreover, AEBA occurred most frequently during November to January. CONCLUSIONS: Real-time PCR was more useful than conventional methods to detect infectious pathogens in patients with AEBA. Accurate detection of pathogens with real-time PCR may enable the selection of appropriate anti-bacterial/viral agents as a part of the treatment for AEBA.