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Background: Triggering Receptor Expressed on Myeloid cells-1 (TREM-1) plays an important role in innate immune system. However, whether and how TREM-1 contributes to obliterative bronchiolitis (OB) progression remains unclear., Methods: A murine orthotopic tracheal transplantation model was constructed to mimic the pathogenesis of OB. qPCR and immunoblotting were used to measure TREM-1 expression. RNA sequencing was used to investigate the impact of TREM-1 on proinflammatory phenotype of macrophages. Trem-1 knockout mice and Nlrp3 knockout mice were generated to investigate the role of the TREM-1/NLRP3 pathway in the proinflammatory phenotype of macrophages. The infiltration of immune cells within the grafts was quantified using immunofluorescence staining. Flow cytometry was used to detect the proportion of different immune cells in mice spleen and the expression levels of iNOS and co-stimulatory molecules in macrophages., Results: The expression of TREM-1 was upregulated in the mouse OB model. Genetic ablation or pharmacological inhibition of TREM-1 ameliorated OB, whereas the stimulation of TREM-1 using anti-TREM-1 agonistic antibody exacerbated OB. Moreover, Trem-1 ablation reduced the infiltration of iNOS + macrophages and limited the T cell responses. In vitro studies revealed that Trem-1 deletion impaired the proinflammatory function and antigen presentation ability of macrophages. Additionally, Trem-1 knockout inhibited the activation of NLRP3 signaling pathway. NLRP3 overexpression restored the proinflammatory phenotype of Trem-1 knockout macrophages., Conclusions: These findings indicated that TREM-1 could promote the proinflammatory phenotype of macrophages through NLRP3 inflammasome activation, thereby exacerbating OB progression. These findings indicated that TREM-1 may serve as a therapeutic target for OB treatment., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Long-term survival after lung transplantation (LTx) remains limited by chronic lung allograft dysfunction (CLAD), which includes 2 main phenotypes: bronchiolitis obliterans syndrome (BOS) and restrictive allograft syndrome (RAS), with possible overlap. We aimed to detail and quantify pathological features of these CLAD sub-types., Methods: Peripheral and central paraffin-embedded explanted lung samples were obtained from 20 consecutive patients undergoing a second LTx for CLAD, from 3 lobes. Thirteen lung samples, collected from non-transplant lobectomies or donor lungs, were used as controls. Blinded semi-quantitative grading was performed to assess airway fibrotic changes, parenchymal and pleural fibrosis, and epithelial and vascular abnormalities., Results: CLAD lung samples had higher scores for all airway- and lung-related parameters compared to controls. There was a notable overlap in histologic scores between BOS and RAS, with a wide range of scores in both conditions. Parenchymal and vascular fibrosis scores were significantly higher in RAS compared to BOS (p = 0.003 for both). We observed a significant positive correlation between the degree of inflammation around each airway, the severity of epithelial changes, and airway fibrosis. Immunofluorescence staining demonstrated a trend toward a lower frequency of club cells in CLAD and a higher frequency of apoptotic club cells in BOS samples (p = 0.01)., Conclusions: CLAD is a spectrum of airway, parenchymal, and pleural fibrosis, as well as epithelial, vascular, and inflammatory pathologic changes, where BOS and RAS overlap significantly. Our semi-quantitative grading score showed a generally high inter-reader reliability and may be useful for future CLAD histologic assessments., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Highly selective RET inhibitor selpercatinib has demonstrated notable efficacy in advanced/progressive RET-mutant medullary thyroid cancer (MTC) patients. However, despite a more tolerable toxicity profile than multikinase inhibitors, peculiar adverse events (AEs) have been described. Obliterative bronchiolitis (OB) is a respiratory disease characterized by inflammation and fibrosis in small conducting airways. We evaluated a 70-year-old man with advanced RET-mutant MTC who developed OB during treatment with selpercatinib. Radiological features of OB occurred early and persisted during selpercatinib treatment, with a waxing and waning pattern. Notably, a partial response of MTC was achieved during the treatment, and selpercatinib was never reduced or interrupted. The almost complete absence of symptoms and the fluctuating trend, without specific treatment for OB, suggested that it is necessary to carefully evaluate the risks mediated by this AE with the risks of modifying or discontinuing the anti-cancer therapy.

Introduction: Chronic rejection is a major complication post-transplantation. Within lung transplantation, chronic rejection was considered as airway centred. Chronic Lung Allograft Dysfunction (CLAD), defined to cover all late chronic complications, makes it more difficult to understand chronic rejection from an immunological perspective. This study investigated the true nature, timing and location of chronic rejection as a whole, within mouse lung transplantation., Methods: 40 mice underwent an orthotopic left lung transplantation, were sacrificed at day 70 and evaluated by histology and in vivo µCT. For timing and location of rejection, extra grafts were sacrificed at day 7, 35, 56 and investigated by ex vivo µCT or single cell RNA (scRNA) profiling., Results: Chronic rejection originated as innate inflammation around small arteries evolving toward adaptive organization with subsequent end-arterial fibrosis and obliterans. Subsequently, venous and pleural infiltration appeared, followed by airway related bronchiolar folding and rarely bronchiolitis obliterans was observed. Ex vivo µCT and scRNA profiling validated the time, location and sequence of events with endothelial destruction and activation as primary onset., Conclusion: Against the current belief, chronic rejection in lung transplantation may start as an arterial response, followed by responses in venules, pleura, and, only in the late stage, bronchioles, as may be seen in some but not all patients with CLAD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Heigl, Kaes, Aelbrecht, Serré, Yamada, Geudens, Van Herck, Vanstapel, Sacreas, Ordies, Frick, Saez Gimenez, Van Slambrouck, Beeckmans, Acet Oztürk, Orlitova, Vaneylen, Claes, Schols, Vande Velde, Schupp, Kaminski, Boesch, Korf, van der Merwe, Dupont, Vanoirbeek, Godinas, Van Raemdonck, Janssens, Gayan-Ramirez, Ceulemans, McDonough, Verbeken, Vos and Vanaudenaerde.)

Background: Pulmonary neutrophils may play a crucial role in the development of bronchiolitis obliterans (BO) following measles virus infection. IL-27 could potentially have a negative regulatory effect on the release of reactive oxygen species and cytotoxic granules in neutrophils., Objective: To investigate the levels of IL-27 in the bronchoalveolar lavage fluid (BALF) of children with post-infectious bronchiolitis obliterans (PIBO) and analyze the relationship between IL-27 levels and neutrophil proportions., Methods: A total of 24 children with PIBO were recruited for the experimental group, while 23 children with bronchial foreign bodies were included in the control group. Bronchoscopic alveolar lavage was performed in both groups. The levels of IL-27 in BALF were measured using enzyme-linked immunosorbent assay (ELISA). The proportions of neutrophils in BALF were determined by smear staining. The relationship between the levels of IL-27 in BALF and the neutrophil proportions was analyzed by the Pearson test., Results: The levels of IL-27 in BALF were significantly lower in children with PIBO compared to children with bronchial foreign bodies (p<0.05). Additionally, the proportions of neutrophils in BALF were significantly higher in children with PIBO compared to children with bronchial foreign bodies (p<0.05). The levels of IL-27 were negatively correlated with the neutrophil proportions in BALF in children with PIBO (p<0.05), but not in children with bronchial foreign bodies (p>0.05)., Conclusion: The present study suggests that a decrease in IL-27 may be associated with an increase in neutrophils in BALF and may contribute to the pathogenesis of PIBO.

Our purpose of this study was to explore the application effect of respiratory flora regulation in bronchiolitis obliterans after lung transplantation, and its regulatory effect on the microbial environment of the lesion and the IL-10/STAT3 signaling pathway. 25 clean-grade C57BL/6 male mice and 5 BALB/c male mice were selected for orthotopic tracheal transplantation and postoperative respiratory flora regulation in a hospital animal room from Jan 2019 to Dec 2021. Next, the changes in the microbial environment and the IL-10/STAT3 signaling pathway before and after respiratory flora regulation were compared, so as to evaluate the regulatory effect of this method. The Simpson index did not show a significant difference before and after respiratory flora regulation intervention (P>0.05). However, the Chao1, ACE, Shannon, and Actinobacteria dominant indices were higher after the intervention. There were significant changes in the abundance of Proteobacteria, Bacteroidetes, Acidobacteria, Firmicutes, Propionibacterium, Corynebacterium, Staphylococcus, and Streptococcus after the intervention (P<0.05). Additionally, IL-10 and STAT3 levels were higher after the intervention and showed significant differences (P<0.05) compared to before. Regulating the respiratory tract flora can improve the microbial environment of bronchiolitis obliterans post-lung transplantation. This helps balance the respiratory flora, increase IL-10 and STAT3 levels, and aid in the recovery of inflammatory responses.

Diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH) is a rare, but increasingly recognized entity that primarily affects middle-aged and elderly women. It is characterized by abnormal proliferation of pulmonary neuroendocrine cells (PNECs) and is considered a preinvasive lesion for carcinoid tumorlets/tumors. Sometimes, DIPNECH is accompanied by constrictive bronchiolitis which usually manifests as chronic cough and/or dyspnea, along with airflow limitation on spirometry. The telltale imaging sign of DIPNECH is the presence of multiple noncalcified pulmonary nodules and mosaic attenuation on CT. However, these clinico-radiologic features of DIPNECH are characteristic but nonspecific; thus, histopathologic confirmation is usually necessary. DIPNECH has an indolent course and only rarely leads to respiratory failure or death; progression to overt neuroendocrine tumor (carcinoid) of the lung occurs in a minority of patients. Of available therapies, somatostatin analogs and mechanistic target of rapamycin inhibitors are the most promising. In this review, we provide an update regarding the diagnosis and management of DIPNECH and describe critical gaps in our understanding of this entity, including the central terms 'diffuse' and 'idiopathic.' We also summarize the inconsistencies in definitions employed by recent studies and discuss the pitfalls of the DIPNECH definitions proposed by the World Health Organization in 2021. In this context, we propose an objective and reproducible radio-pathologic case definition intended for implementation in the research realm and seeks to enhance homogeneity across cohorts. Furthermore, we discuss aspects of PNECs biology which suggest that PNEC hyperplasia may contribute to the pathogenesis of phenotypes of lung disease aside from constrictive bronchiolitis and carcinoid tumorlets/tumors. Finally, we steer attention to some of the most pressing and impactful research questions awaiting to be unraveled.

Purpose: Childhood interstitial lung disease (chILD) is an umbrella concept covering a wide range of rare lung diseases, many of which are unique to childhood. The diagnosis is based on clinical presentation, multidetector computed tomography (MDCT), genetic testing, lung-function testing, and lung biopsy. Because knowledge of the usefulness of MDCT pattern recognition in ChILD is at present limited, we examined the occurrence of MDCT patterns in children with histologically confirmed interstitial lung disease., Method: We searched the biopsy, MDCT, and clinical information database of a single national paediatric referral hospital for 2004-2020. Data were from affected children under age 18. MDCT images we reanalysed while blinded to the identity and referral information., Results: We included 90 patients, of whom 63 (70 %) were male. The median age at biopsy was 1.3 years (interquartile range 0.1-16.8). Biopsy findings fell into 26 histological classes covering all nine chILD classification categories. We recognized six distinct MDCT patterns: neuroendocrine cell hyperplasia of infancy (23), organizing pneumonia (5), non-specific interstitial pneumonia (4), bronchiolitis obliterans (3), pulmonary alveolar proteinosis (2), and bronchopulmonary dysplasia (n = 2). Of the total 90, in 51 (57 %) children, none of these six MDCT patterns appeared. Of those 39 children with a recognizable MDCT pattern, in 34 (87 %), that pattern predicted their final diagnosis., Conclusions: Among cases of chILD, we identified a specific predefined MDCT pattern in only 43 %. However, when such a recognizable pattern occurred, it was predictive of the final chILD diagnosis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Bronchiolitis obliterans (BO) is a debilitating disease of the small airways that can develop following exposure to toxic chemicals as well as respiratory tract infections. BO development is strongly associated with diacetyl (DA) inhalation exposures at occupationally relevant concentrations or severe influenza A viral (IAV) infections. However, it remains unclear whether lower dose exposures or more mild IAV infections can result in similar pathology. In the current work, we combined these two common environmental exposures, DA and IAV, to test whether shorter DA exposures followed by sublethal IAV infection would result in similar airways disease. Adult mice exposed to DA vapors 1 h/day for 5 consecutive days followed by infection with the airway-tropic IAV H3N2 (HKx31) resulted in increased mortality, increased bronchoalveolar lavage (BAL) neutrophil percentage, mixed obstruction and restriction by lung function, and subsequent airway remodeling. Exposure to DA or IAV alone failed to result in significant pathology, whereas mice exposed to DA + IAV showed increased α-smooth muscle actin (αSMA) and epithelial cells coexpressing the basal cell marker keratin 5 (KRT5) with the club cell marker SCGB1A1. To test whether DA exposure impairs epithelial repair after IAV infection, mice were infected first with IAV and then exposed to DA during airway epithelial repair. Mice exposed to IAV + DA developed similar airway remodeling with increased subepithelial αSMA and epithelial cells coexpressing KRT5 and SCGB1A1. Our findings reveal an underappreciated concept that common environmental insults while seemingly harmless by themselves can have catastrophic implications on lung function and long-term respiratory health when combined.

Objective: Critically ill patients with solid tumors complicated with paraneoplastic pemphigus are usually treated in intensive care units (ICU) for perioperative management after surgical treatment. In this study, the clinical characteristics and predictors of long-term prognosis of these critically ill patients were analyzed., Methods: the clinical and laboratory data of 63 patients with solid tumors complicated with paraneoplastic pemphigus admitted to ICU from 2005 to 2020 were retrospectively analyzed, and the survival status of the patients were followed up., Results: Among the 63 patients, 79.4% had Castleman disease as the primary tumor, and 20.6% with other pathological types; 69.8% had severe-extensive skin lesions, and 30.2% had other skin lesions; the patients with bronchiolitis obliterans accounted for 44.4%, and 55.6% were not merged. Postoperative fungal infection occurred in 23.8% of the patients, and 76.2% without fungal infection. The median follow-up time was 95 months, and 25 patients died during the study period. The 1-year, 3-year and 5-year survival rates were 74.6% (95% CI 63.8%-85.4%), 67.4% (95% CI 55.6%-79.2%) and 55.1% (95% CI 47.9%-62.3%), respectively. The log-rank univariate analysis showed that the patients had age>40 years ( P =0.042), preoperative weight loss>5 kg ( P =0.002), preoperative albumin < 30 g/L ( P < 0.001), paraneoplastic pemphigus complicated with bronchiolitis obliterans ( P =0.002), and perioperative fungal infection ( P < 0.001) had increased mortality. Cox univariate analysis showed that preoperative weight loss >5 kg ( P =0.005), preoperative albumin < 30 g/L ( P < 0.001), paraneoplastic pemphigus complicated with bronchiolitis obliterans ( P =0.009), preoperative bacterial pulmonary infection ( P =0.007), prolonged surgical time ( P =0.048), postoperative oxygenation index ( P =0.012) and low albumin ( P =0.010) and hemoglobin concentration ( P =0.035) in ICU, acute physiology and chronic health evaluation (APACHE Ⅱ) score ( P =0.001); sequential organ failure assessment (SOFA) score ( P =0.010), and postoperative fungal infection ( P < 0.001) were risk factors for long-term survival. Cox regression model for multivariate analysis showed that preoperative weight loss > 5 kg ( HR 4.44; 95% CI 1.47-13.38; P =0.008), and preoperative albumin < 30 g/L ( HR 4.38; 95% CI 1.72-11.12; P =0.002), bronchiolitis obliterans ( HR 2.69; 95% CI 1.12-6.50; P =0.027), and postoperative fungal infection ( HR 4.85; 95% CI 2.01-11.72; P < 0.001) were independent risk factors for postoperative mortality., Conclusion: The 5-year survival rate of critically ill patients undergoing surgery for paraneoplastic pemphigus combined with solid tumors is approximately 55.1%, with preoperative weight loss > 5 kg, albumin < 30 g/L, bronchiolitis obliterans and postoperative fungal infection were associated with an increased risk of near- and long-term postoperative mortality.

Idiopathic obliterative bronchiolitis (OB) is a rare disease that usually requires a surgical lung biopsy. A 25-year-old woman with progressive exertional dyspnea for several months showed a severe mixed restrictive and obstructive pattern on spirometry. Chest computed tomography showed a mosaic pattern, and pulmonary ventilation-perfusion scintigraphy showed a matched defect. The bronchoscopic specimens obtained from both the alveolar and bronchiolar regions of the predicted lesion area contributed to the diagnosis of OB. She had no underlying causes of secondary OB, and she was diagnosed with idiopathic OB. Since lung transplantation was indicated, she was referred to a lung transplantation-certified hospital.

This descriptive case series retrospectively reviewed medical records from thirty-one previously healthy, war-fighting veterans who self-reported exposure to airborne hazards while serving in Iraq and Afghanistan between 2003 and the present. They all noted new-onset dyspnea, which began during deployment or as a military contractor. Twenty-one subjects underwent non-invasive pulmonary diagnostic testing, including maximum expiratory pressure (MEP) and impulse oscillometry (IOS). In addition, five soldiers received a lung biopsy; tissue results were compared to a previously published sample from a soldier in our Iraq Afghanistan War Lung Injury database and others in our database with similar exposures, including burn pits. We also reviewed civilian control samples (5) from the Stony Brook University database. Military personnel were referred to our International Center of Excellence in Deployment Health and Medical Geosciences, Donald and Barbara Zucker School of Medicine at Hofstra/Northwell under the auspices of Northwell IRB: 17-0140-FIMR Feinstein Institution for Medical Research "Clinicopathologic characteristics of Iraq Afghanistan War Lung Injury." We retrospectively examined medical records, including exposure data, radiologic imaging, and non-invasive pulmonary function testing (MGC Diagnostic Platinum Elite Plethysmograph) using the American Thoracic Society (ATS) standard interpretation based on Morgan et al., and for a limited cohort, biopsy data. Lung tissue, when available, was examined for carbonaceous particles, polycyclic aromatic hydrocarbons (Raman spectroscopy), metals, titanium connected to iron (Brookhaven National Laboratory, National Synchrotron Light Source II, Beamline 5-ID), oxidized metals, combustion temperature, inflammatory cell accumulation and fibrosis, neutrophil extracellular traps, Sirius red, Prussian Blue, as well as polarizable crystals/particulate matter/dust. Among twenty-one previously healthy, deployable soldiers with non-invasive pulmonary diagnostic tests, post-deployment, all had severely decreased MEP values, averaging 42% predicted. These same patients concurrently demonstrated abnormal airways reactance (X5Hz) and peripheral/distal airways resistance (D5-D20%) via IOS, averaging - 1369% and 23% predicted, respectively. These tests support the concept of airways hyperresponsiveness and distal airways narrowing, respectively. Among the five soldiers biopsied, all had constrictive bronchiolitis. We detected the presence of polycyclic aromatic hydrocarbons (PAH)-which are products of incomplete combustion-in the lung tissue of all five warfighters. All also had detectable titanium and iron in the lungs. Metals were all oxidized, supporting the concept of inhaling burned metals. Combustion temperature was consistent with that of burned petrol rather than higher temperatures noted with cigarettes. All were nonsmokers. Neutrophil extracellular traps were reported in two biopsies. Compared to our prior biopsies in our Middle East deployment database, these histopathologic results are similar, since all database biopsies have constrictive bronchiolitis, one has lung fibrosis with titanium bound to iron in fixed mathematical ratios of 1:7 and demonstrated polarizable crystals. These results, particularly constrictive bronchiolitis and polarizable crystals, support the prior data of King et al. (N. Engl. J. Med. 365:222-230, 2011) Soldiers in this cohort deployed to Iraq and Afghanistan since 2003, with exposure to airborne hazards, including sandstorms, burn pits, and improvised explosive devices, are at high risk for developing chronic clinical respiratory problems, including: (1) reduction in respiratory muscle strength; (2) airways hyperresponsiveness; and (3) distal airway narrowing, which may be associated with histopathologic evidence of lung damage, reflecting inhalation of burned particles from burn pits along with particulate matter/dust. Non-invasive pulmonary diagnostic tests are a predictor of burn pit-induced lung injury., (© 2022. The Author(s).)

Background: The major obstacle for long-term survival after successful lung transplantation is the development of bronchiolitis obliterans (BO) which is one phenotype of chronic lung allograft dysfunction (CLAD). Nintedanib has beneficial effects treating neoplastic diseases and idiopathic pulmonary fibrosis by blocking tyrosine kinase receptors. These receptors play an important role in alloimmune-mediated proliferative diseases. The aim of this study was to determine the effect of nintedanib on proliferative airway changes after orthotopic trachea transplantation in mice., Methods: C57BL/6 mice (H-2 b ) donor tracheas were orthotopically transplanted into CBA/J mice (H-2 k ). After transplantation, recipients were daily treated with nintedanib (60 mg/kg; p.o.). Histological and immunofluorescence analysis were performed after 30 days and intragraft gene expression measurements after 14 days of treatment, respectively., Results: Tracheal allografts from mice treated with nintedanib showed significantly less features of chronic rejection than untreated allografts reflected in a higher epithelium/lamina propria ratio (ELR) [ELR: 0.65 ± 0.13 nintedanib vs. 0.50 ± 0.07 untreated controls; p < 0.05] and a reduced submucosal smooth muscle actin (SMA) content [SMA: 1.26% ± 0.78% nintedanib vs. 2.18% ± 1.01% untreated controls; p < 0.01]. Furthermore, lower T cell, macrophage and dendritic cell infiltration was detected in the nintedanib treated grafts. The protein and intragraft mRNA expression of receptor subtypes was considerably decreased in grafts of nintedanib treated mice. The mRNA expression of relevant immune mediators was affected by nintedanib treatment., Conclusion: Receptor blocking by nintedanib reduced alloimmune-induced inflammation and chronic airway changes in mouse trachea allografts and might be a promising approach to diminish the development of BO in lung transplants., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Since December 2019, the SARS-CoV-2 pandemic significantly has impacted the medical community. When infected with SARS-CoV-2, most of the patients developed bilateral pneumonia. We have herein presented the atypical case of a patient who developed unilateral SARS-CoV-2 pneumonia, affecting only the second lung allograft re-transplanted (re-LTX)., Case Presentation: A SARS-CoV-2 infection occurred in a 2-dose vaccinated patient with LTx with a history of second unilateral lung transplantation performed after an end-stage bronchiolitis obliterans syndrome. The first symptoms started with a flu-like syndrome, and the patient's clinical condition worsened with nonsevere acute respiratory failure requiring conventional oxygen therapy. Treatment consisted in administrating specific anti-SARS-CoV-2 monoclonal antibodies along with probabilistic antibiotherapy, anticoagulation, and steroids. On day 7, the patient was discharged from hospital. We aimed to assess this atypical unilateral pneumonia based on different explorations. A ventilation scintigraphy showed a severe ventilation decrease owing to end-stage bronchiolitis obliterans syndrome within the left first allograft, which may be associated with asymmetrical virus diffusion between the 2 lungs. We did not identify any other relevant differences with respect to the 2 donors' clinical characteristics. Using specific immunohistochemistry staining against angiotensin converting enzyme-2 receptor, the main known receptor for SARS-CoV-2 binding on airway epithelial cells, no staining difference was observed between the 2 lung biopsies that were collected at re-LTx from each lung., Conclusions: With the present case report, we aimed to highlight how this kind of unusual presentation may be caused by the difference of ventilation between the 2 lungs., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Despite advances in the field, chronic graft-versus-host-disease (cGVHD) remains a leading cause of morbidity and mortality following allogenic hematopoietic stem cell transplant. Because treatment options remain limited, we tested efficacy of anticancer, chromatin-modifying enzyme inhibitors in a clinically relevant murine model of cGVHD with bronchiolitis obliterans (BO). We observed that the novel enhancer of zeste homolog 2 (EZH2) inhibitor JQ5 and the BET-bromodomain inhibitor JQ1 each improved pulmonary function; impaired the germinal center (GC) reaction, a prerequisite in cGVHD/BO pathogenesis; and JQ5 reduced EZH2-mediated H3K27me3 in donor T cells. Using conditional EZH2 knockout donor cells, we demonstrated that EZH2 is obligatory for the initiation of cGVHD/BO. In a sclerodermatous cGVHD model, JQ5 reduced the severity of cutaneous lesions. To determine how the 2 drugs could lead to the same physiological improvements while targeting unique epigenetic processes, we analyzed the transcriptomes of splenic GCB cells (GCBs) from transplanted mice treated with either drug. Multiple inflammatory and signaling pathways enriched in cGVHD/BO GCBs were reduced by each drug. GCBs from JQ5- but not JQ1-treated mice were enriched for proproliferative pathways also seen in GCBs from bone marrow-only transplanted mice, likely reflecting their underlying biology in the unperturbed state. In conjunction with in vivo data, these insights led us to conclude that epigenetic targeting of the GC is a viable clinical approach for the treatment of cGVHD, and that the EZH2 inhibitor JQ5 and the BET-bromodomain inhibitor JQ1 demonstrated clinical potential for EZH2i and BETi in patients with cGVHD/BO., (© 2022 by The American Society of Hematology.)

Histopathologic evidence of deployment-related constrictive bronchiolitis (DRCB) has been identified in soldiers deployed to Southwest Asia. While inhalational injury to the airway epithelium is suspected, relatively little is known about the pathogenesis underlying this disabling disorder. Club cells are local progenitors critical for repairing the airway epithelium after exposure to various airborne toxins, and a prior study using an inducible transgenic murine model reported that 10 days of sustained targeted club cell injury causes constrictive bronchiolitis. To further understand the mechanisms leading to small airway fibrosis, a murine model was employed to show that sustained club cell injury elicited acute weight loss, caused increased local production of proinflammatory cytokines, and promoted accumulation of numerous myeloid cell subsets in the lung. Transition to a chronic phase was characterized by up-regulated expression of oxidative stress-associated genes, increased activation of transforming growth factor-β, accumulation of alternatively activated macrophages, and enhanced peribronchiolar collagen deposition. Comparative histopathologic analysis demonstrated that sustained club cell injury was sufficient to induce epithelial metaplasia, airway wall thickening, peribronchiolar infiltrates, and clusters of intraluminal airway macrophages that recapitulated key abnormalities observed in DRCB. Depletion of alveolar macrophages in mice decreased activation of transforming growth factor-β and ameliorated constrictive bronchiolitis. Collectively, these findings implicate sustained club cell injury in the development of DRCB and delineate pathways that may yield biomarkers and treatment targets for this disorder., (Copyright © 2022 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Obliterative bronchiolitis (OB) after lung transplantation is a nonreversible, life-threatening complication. Herein, the role of vascular endothelial growth factor receptor (Vegfr)-1 and -2 was investigated in the development of obliterative airway disease (OAD), an experimental model for OB. The nonimmunosuppressed recipients underwent transplantation with fully major histocompatibility complex mismatched heterotopic tracheal allografts and received Vegfr1 and -2-specific monoclonal antibodies either alone or in combination, or rat IgG as a control. The treatment with Vegfr1- or -2-blocking antibody significantly decreased intragraft mRNA expression of natural killer cell activation markers early after transplantation. This was followed by reduced infiltration of Cd11b + cells and Cd4 + T cells as well as down-regulated mRNA expression of proinflammatory chemokines and profibrotic growth factors. However, blocking of both Vegfr1 and -2 was necessary to reduce luminal occlusion. Furthermore, concomitant inhibition of the calcineurin activation pathway almost totally abolished the development of OAD. This study proposes that blocking of Vegf receptors blunted natural killer cell and innate immune responses early after transplantation and attenuated the development of OAD. The results of this study suggest that further studies on the role of Vegfr1 and -2 blocking in development of obliterative airway lesions might be rewarding., (Copyright © 2022 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Background: Chronic Lung Allograft Dysfunction (CLAD) is a major obstacle for long term survival after lung transplantation (LTx). Besides Bronchiolitis Obliterans Syndrome, two other phenotypes of CLAD, restrictive allograft syndrome (RAS) and mixed phenotype, have been described. Trials to test in these conditions are desperately needed and analyzing natural outcome to plan such trials is essential., Methods: We performed a retrospective analysis of functional outcome in bilateral LTx recipients with RAS and mixed phenotype, transplanted between 2009 and 2018 in five large European centers with follow- up spirometry up to 12 months after diagnosis. Based on these data, sample size and power calculations for randomized therapeutic trial was estimated using two imputation methods for missing values., Results: Seventy patients were included (39 RAS and 31 mixed phenotype), median 3.1 years after LTx when CLAD was diagnosed. Eight, 13 and 25 patients died within 6, 9 and 12 months after diagnosis and a two patients underwent re-transplantation within 12 months leading to a graft survival of 89, 79 and 61% six, nine and 12 months after diagnosis, respectively. Observed FEV1 decline was 451 ml at 6 months and stabilized at 9 and 12 months, while FVC showed continuous decline. Using two methods of imputation, a progressive further decline after 6 months for FEV1 was noted., Conclusion: The poor outcome of these two specific CLAD phenotypes suggests the urgent need for future therapeutic randomized trials. The number of missing values in a potential trial seems to be high and most frequently attributed to death. Survival may be used as an endpoint in clinical trials in these distinct phenotypes and imputation techniques are relevant if graft function is used as a surrogate of disease progression in future trials., Competing Interests: The authors have declared that no competing interests exist.

After deployment to Southwest Asia, some soldiers develop persistent respiratory symptoms, including exercise intolerance and exertional dyspnea. We identified 50 soldiers with a history of deployment to Southwest Asia who presented with unexplained dyspnea and underwent an unrevealing clinical evaluation followed by surgical lung biopsy. Lung tissue specimens from 17 age-matched, nonsmoking subjects were used as controls. Quantitative histomorphometry was performed for evaluation of inflammation and pathologic remodeling of small airways, pulmonary vasculature, alveolar tissue and visceral pleura. Compared with control subjects, lung biopsies from affected soldiers revealed a variety of pathologic changes involving their distal lungs, particularly related to bronchovascular bundles. Bronchioles from soldiers had increased thickness of the lamina propria, smooth muscle hypertrophy, and increased collagen content. In adjacent arteries, smooth muscle hypertrophy and adventitial thickening resulted in increased wall-to-lumen ratio in affected soldiers. Infiltration of CD4 and CD8 T lymphocytes was noted within airway walls, along with increased formation of lymphoid follicles. In alveolar parenchyma, collagen and elastin content were increased and capillary density was reduced in interalveolar septa from soldiers compared to control subjects. In addition, pleural involvement with inflammation and/or fibrosis was present in the majority (92%) of soldiers. Clinical follow-up of 29 soldiers (ranging from 1 to 15 y) showed persistence of exertional dyspnea in all individuals and a decline in total lung capacity. Susceptible soldiers develop a postdeployment respiratory syndrome that includes exertional dyspnea and complex pathologic changes affecting small airways, pulmonary vasculature, alveolar tissue, and visceral pleura., Competing Interests: Conflicts of Interest and Source of Funding: Supported by the Department of Defense W81XWH-17-1-0503, National Institutes of Health HL126176, generous donation by Ms. Carol Odess. S.I.R. was an employee of AstraZeneca from 2015 to 2019 and continues to own shares that were received as part of his compensation. As part of that employment, he represented AstraZeneca on the Board of Directors of Dizal Pharma without additional compensation. In the last 3 years, he has consulted for Bergenbio, GlaxoSmithKline, NovoVentures and Verona. Between 1996 and 2007, his university received funding from tobacco companies that supported studies relating to harm reduction and to the impact of tobacco smoke on stem cells. As part of this work, he consulted with RJ Reynolds without personal fee on the topic of harm reduction, received funding from RJ Reynolds to evaluate the effect of a harm reduction product in normal smokers (1996) and in subjects with chronic bronchitis (1999) and to assess the effect of smoking cessation on lower respiratory tract inflammation (2000); he participated in a Philip Morris multi-center study to assess biomarkers of smoke exposure (2002); he received funding for a clinical trial from the Institute for Science and Health (2005), which receives support from the tobacco industry, to evaluate biomarkers in exhaled breath associated with smoking cessation and reduction. This study was supplemented with funding from Lorillard and RJ Reynolds to expand the spectrum of biomarkers assessed. He received a grant from the Philip Morris External Research Program (2005) to assess the impact of cigarette smoking on circulating stem cells in the mouse. There are no active tobacco-industry funded projects. All ties with tobacco industry companies and entities supported by tobacco companies were terminated in 2007. For the remaining authors none were declared., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc.)

Bronchiolitis obliterans syndrome, a common form of chronic lung allograft dysfunction, is the major limitation to long-term survival after lung transplantation. The histologic correlate is progressive, fibrotic occlusion of small airways, obliterative bronchiolitis lesions, which ultimately lead to organ failure. The molecular composition of these lesions is unknown. In this sutdy, the protein composition of the lesions in explanted lungs from four end-stage bronchiolitis obliterans syndrome patients was analyzed using laser-capture microdissection and optimized sample preparation protocols for mass spectrometry. Immunohistochemistry and immunofluorescence were used to determine the spatial distribution of commonly identified proteins on the tissue level, and protein signatures for 14 obliterative bronchiolitis lesions were established. A set of 39 proteins, identified in >75% of lesions, included distinct structural proteins (collagen types IV and VI) and cellular components (actins, vimentin, and tryptase). Each respective lesion exhibited a unique composition of proteins (on average, n = 66 proteins), thereby mirroring the morphologic variation of the lesions. Antibody-based staining confirmed these mass spectrometry-based findings. The 14 analyzed obliterative bronchiolitis lesions showed variations in their protein content, but also common features. This study provides molecular and morphologic insights into the development of chronic rejection after lung transplantation. The protein patterns in the lesions were correlated to pathways of extracellular matrix organization, tissue development, and wound healing processes., (Copyright © 2021 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Bronchiolitis obliterans (BO) is a devastating lung disease seen commonly after lung transplant, following severe respiratory tract infection or chemical inhalation exposure. Diacetyl (DA; 2,3-butanedione) is a highly reactive alpha-diketone known to cause BO when inhaled, however, the mechanisms of how inhalation exposure leads to BO development remains poorly understood. In the current work, we combined two clinically relevant models for studying the pathogenesis of DA-induced BO: (1) an in vivo rat model of repetitive DA vapor exposures with recovery and (2) an in vitro model of primary human airway epithelial cells exposed to pure DA vapors. Rats exposed to 5 consecutive days 200 parts-per-million DA 6 h per day had worsening survival, persistent hypoxemia, poor weight gain, and histologic evidence of BO 14 days after DA exposure cessation. At the end of exposure, increased expression of the ubiquitin stress protein ubiquitin-C accumulated within DA-exposed rat lung homogenates and localized primarily to the airway epithelium, the primary site of BO development. Lung proteasome activity increased concurrently with ubiquitin-C expression after DA exposure, supportive of significant proteasome stress. In primary human airway cultures, global proteomics identified 519 significantly modified proteins in DA-exposed samples relative to controls with common pathways of the ubiquitin proteasome system, endosomal reticulum transport, and response to unfolded protein pathways being upregulated and cell-cell adhesion and oxidation-reduction pathways being downregulated. Collectively, these two models suggest that diacetyl inhalation exposure causes abundant protein damage and subsequent ubiquitin proteasome stress prior to the development of chemical-induced BO pathology.

Background: Chronic lung allograft dysfunction (CLAD) and its obstructive form, the obliterative bronchiolitis (OB), are the main long-term complications related to high mortality rate postlung transplantation. CLAD treatment lacks a significant success in survival. Here, we investigated a new strategy through inhibition of the proinflammatory mitogen- and stress-activated kinase 1 (MSK1) kinase., Methods: MSK1 expression was assessed in a mouse OB model after heterotopic tracheal allotransplantation. Pharmacological inhibition of MSK1 (H89, fasudil, PHA767491) was evaluated in the murine model and in a translational model using human lung primary fibroblasts in proinflammatory conditions. MSK1 expression was graded over time in biopsies from a cohort of CLAD patients., Results: MSK1 mRNA progressively increased during OB (6.4-fold at D21 posttransplantation). Inhibition of MSK1 allowed to counteract the damage to the epithelium (56% restoration for H89), and abolished the recruitment of MHCII+ (94%) and T cells (100%) at the early inflammatory phase of OB. In addition, it markedly decreased the late fibroproliferative obstruction in allografts (48%). MSK1 inhibitors decreased production of IL-6 (whose transcription is under the control of MSK1) released from human lung fibroblasts (96%). Finally, we confirmed occurrence of a 2.9-fold increased MSK1 mRNA expression in lung biopsies in patients at 6 months before CLAD diagnosis as compared to recipients with stable lung function., Conclusions: These findings suggest the overall interest of the MSK1 kinase either as a marker or as a potential therapeutic target in lung dysfunction posttransplantation., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Immunosuppression therapy is ineffective at preventing bronchiolitis obliterans syndrome (BOS), primarily a disease of the small airways (SAs). Our previous reports show increased senescent CD28null T and natural killer T (NKT)-like cells in the peripheral blood of patients with BOS and increased cytotoxic, proinflammatory lymphocytes in the SAs. We hypothesized that the cytotoxic, proinflammatory lymphocytes in the SAs would be steroid-resistant senescent CD28null lymphocytes., Methods: Intracellular cytotoxic mediator granzyme B, interferon (IFN)-γ and tumor necrosis factor (TNF)-α proinflammatory cytokines, and CD28 were measured in the blood, bronchoalveolar lavage, large airway, and SA brushing T and NKT-like cells from 10 patients with BOS, 11 stable lung transplant recipients, and 10 healthy age-matched controls. SA brushings were cultured in the presence of ±1 µmol/liter prednisolone, ±5 mg/liter theophylline, and ±2.5 ng/ml cyclosporine A, and IFN-γ and TNF-α proinflammatory cytokines were assessed using flow cytometry., Results: Increased SA CD28null T and NKT-like cells were identified in patients with BOS compared with that in the controls and stable transplant recipients. Loss of CD28 was associated with increased T and NKT-like cells expressing granzyme B, IFN-γ, and TNF-α. Loss of CD28 expression by CD8+ T cells was significantly associated with forced expiratory volume in 1 sec (R = 0.655, p = 0.006) and with time after transplantation (R = -0.552, p = 0.041). Treatment with prednisolone + theophylline + cyclosporin A inhibited IFN-γ and TNF-α production by SA CD28null CD8+ T and NKT-like cells additively., Conclusions: BOS is associated with the loss of CD28 in SA cytotoxic, proinflammatory senescent T and NKT-like lymphocytes. Treatment options that target the proinflammatory nature of these cells in the SAs may improve graft survival., (Copyright © 2020 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Transplantation of solid organs and hematopoietic stem cells represents an important therapeutic option for a variety of end-stage pulmonary diseases, aggressive hematopoietic neoplasms, or severe immunodeficiencies. Although the overall survival following transplantation has generally improved over recent decades, long-time survival of lung and stem-cell transplant recipients is still alarmingly low with an average 5‑year survival rate of only 50-60%. Chronic allo-immunoreactions in general and pulmonary allo-immunoreactions with subsequent fibrosis in particular are major reasons for this poor outcome. Comparable patterns of fibrotic lung remodeling are observed following both lung and hematopoietic stem-cell transplantation. Besides the meanwhile well-established obliterative and functionally obstructive remodeling of the small airways - obliterative bronchiolitis - a specific restrictive subform of fibrosis, namely alveolar fibroelastosis, has been identified. Despite their crucial impact on patient outcome, both entities can be very challenging to detect by conventional histopathological analysis. Their underlying mechanisms are considered overreaching aberrant repair attempts to acute lung injuries with overactivation of (myo-) fibroblasts and excessive and irreversible deposition of extracellular matrix. Of note, the underlying molecular mechanisms are widely divergent between these two morphological entities and are independent of the underlying clinical setting.Further comprehensive investigations of these fibrotic alterations are key to the development of much-needed predictive diagnostics and curative concepts, considering the high mortality of pulmonary fibrosis following transplantation.

Bronchiolitis obliterans is a clinical entity which results from a variety of etiologies and has a detrimental impact on patients' quality of life when it remains undiagnosed and untreated. Due to its non-pathognomic clinical symptoms and signs, physicians often proceed to radiological examination, especially with high resolution chest tomography. Histological examination reveals constrictive bronchiolitis, the pathological definition of bronchiolitis obliterans. Due to an almost normal aspect of the lung parenchyma this condition can be missed. However, its recognition and the identification of a possible exposure are important for removing the patient from the incriminating agent. We present a case of constrictive bronchiolitis in a metal-cutting worker, highlighting the principal findings of this disease., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Chronic allograft dysfunction (CAD) resulting from fibrosis is the major limiting factor for long-term survival of lung transplant patients. Myofibroblasts promote fibrosis in multiple organs, including the lungs. In this study, we identified PLK1 as a promoter of myofibroblast differentiation and investigated the mechanism by which its inhibition alleviates transplant-associated obliterative bronchiolitis (OB) during CAD. High-throughput bioinformatic analyses and experiments using the murine heterotopic tracheal transplantation model revealed that PLK1 is upregulated in grafts undergoing CAD as compared with controls, and that inhibiting PLK1 alleviates OB in vivo . Inhibition of PLK1 in vitro reduced expression of the specific myofibroblast differentiation marker α-smooth muscle actin (α-SMA) and decreased phosphorylation of both MEK and ERK. Importantly, we observed a similar phenomenon in human primary fibroblasts. Our results thus highlight PLK1 as a promising therapeutic target for alleviating transplant-associated OB through suppression of TGF-β1-mediated myofibroblast differentiation.

Background: Localization of inflammatory stimuli may direct lung allografts to different phenotypes of chronic dysfunction, such as bronchiolitis obliterans syndrome (BOS) or restrictive allograft syndrome (RAS). We hypothesized that airway stimulation with lipopolysaccharide (LPS) in rats leads to airway-centered inflammation similar to human BOS., Methods: Rat left lung transplantation was conducted (donor: Brown Norway, recipient: Lewis). Allotransplant recipients received cyclosporine A (CsA) until postoperative day 56 with airway instillation of LPS (Allo-LPS, n = 8), phosphate buffered saline (Allo-PBS, n = 5) from days 35 to 46 (3 times a wk), or no further treatment (n = 4). Some allotransplant recipients received CsA until day 14 and were immunosuppression free after day 15 until day 56. Bronchial and pleural fibrosis were semiquantified; alveolar fibrosis was evaluated with a histological scale., Results: The Allo-LPS group had significantly increased International Society for Heart and Lung Transplantation rejection grades (grade A, P = 0.005; grade B, P = 0.004), bronchial obstructive proportion (0.34 ± 0.04% [Allo-LPS] versus 0.11 ± 0.04% [Allo-PBS], P = 0.006), and airway resistance (3.05 ± 1.78 cm H2O·s/mL [Allo-LPS] versus 0.83 ± 0.58 cm H2O·s/mL [Allo-PBS], P = 0.007) compared with other groups. Allotransplant recipients that underwent a short course of CsA developed RAS-like fibrosis involving the airways, alveoli, and pleura., Conclusions: Airway instillation of LPS in allografts under immunosuppression resulted in BOS-like airway-centered inflammation and fibrosis distinct from RAS-like diffuse fibrosis, which was induced by a shortened course of immunosuppression. We propose novel animal models for BOS and RAS after lung transplantation.

Background: Dysregulated airway epithelial repair following injury is a proposed mechanism driving posttransplant bronchiolitis obliterans (BO), and its clinical correlate bronchiolitis obliterans syndrome (BOS). This study compared gene and cellular characteristics of injury and repair in large (LAEC) and small (SAEC) airway epithelial cells of transplant patients., Methods: Subjects were recruited at the time of routine bronchoscopy posttransplantation and included patients with and without BOS. Airway epithelial cells were obtained from bronchial and bronchiolar brushing performed under radiological guidance from these patients. In addition, bronchial brushings were also obtained from healthy control subjects comprising of adolescents admitted for elective surgery for nonrespiratory-related conditions. Primary cultures were established, monolayers wounded, and repair assessed (±) azithromycin (1 µg/mL). In addition, proliferative capacity as well as markers of injury and dysregulated repair were also assessed., Results: SAEC had a significantly dysregulated repair process postinjury, despite having a higher proliferative capacity than large airway epithelial cells. Addition of azithromycin significantly induced repair in these cells; however, full restitution was not achieved. Expression of several genes associated with epithelial barrier repair (matrix metalloproteinase 7, matrix metalloproteinase 3, the integrins β6 and β8, and β-catenin) were significantly different in epithelial cells obtained from patients with BOS compared to transplant patients without BOS and controls, suggesting an intrinsic defect., Conclusions: Chronic airway injury and dysregulated repair programs are evident in airway epithelium obtained from patients with BOS, particularly with SAEC. We also show that azithromycin partially mitigates this pathology.

Background: Lung transplantation (LTx) is a last treatment option for patients with an end-stage pulmonary disease. Chronic lung allograft dysfunction, which generally manifests as bronchiolitis obliterans syndrome (BOS), is a major long-term survival limitation. During injury, inflammation and BOS monocytes are recruited. We determined whether changes in count, subset distribution, and functionality by surface marker expression coincided with BOS development., Methods: Fresh whole-blood samples were analyzed from 44 LTx patients, including 17 patients diagnosed with BOS, and compared with 10 age-matched healthy controls and 9 sarcoidosis patients as positive controls. Monocytes were quantified and analyzed using flow cytometry. Based on surface marker expression, classical, intermediate, and nonclassical subsets were determined, and functional phenotypes were investigated., Results: The absolute count of monocytes was decreased in LTx and slightly increased in BOS patients. The relative count shifted toward classical monocytes at the expense of nonclassical monocytes in LTx and BOS. Surface marker expression was highest on intermediate monocytes. The expression of both CD36 and CD163 was significantly increased in the LTx and BOS cohort. The difference between the BOS cohort and the LTx cohort was only subtle, with a significant decrease in HLA-DR expression on nonclassical monocytes in BOS., Conclusions: Monocyte subsets and surface marker expression changed significantly in transplantation patients, while BOS-specific changes were understated. More research is needed to determine whether and how monocytes influence the disease process and how current immunosuppressants affect their normal function in vivo., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Long term survival post lung transplantation (LTx) is limited by the occurrence of bronchiolitis obliterans syndrome (BOS). One mechanism involved is the epithelial-mesenchymal transition (EMT). Membrane microparticles (MPs) are known to be involved in some respiratory diseases and in other organs allograft rejection episodes. We hypothesized that leukocyte-derived MPs likely contribute to EMT. To emphasize this physiological concept, our objectives were to: (1) confirm the presence of EMT on explanted lungs from patients who underwent a second LTx for BOS; 2) characterize circulating MPs in transplanted patients, with or without BOS; (3) evaluate in vitro the effect of monocyte-derived MPs in EMT of human bronchial epithelial cells. Our IHC analysis on explanted graft lungs revealed significant pathological signs of EMT with an inhomogeneous destruction of the bronchial epithelium, with decreased expression of the epithelial protein E-cadherin and increased expression of the mesenchymal protein Vimentin. The immunophenotyping of MPs demonstrated that the concentration of MPs carrying E-cadherin was lower in patients affected by BOS (p = .007). In vitro, monocyte-derived MPs produced with LPS were associated with decreased E-cadherin expression (p < .05) along with significant morphological and functional cell modifications. MPs may play a role in EMT onset in bronchial epithelium following LTx., Competing Interests: Declaration of Competing Interest None of the authors has a financial relationship with a commercial entity that has an interest in the subject of the presented manuscript or other conflicts of interest to disclose., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Three major histologic patterns of bronchiolitis: obliterative bronchiolitis, follicular bronchiolitis, and diffuse panbronchiolitis, are reviewed in detail. These distinct patterns of primary bronchiolar injury provide a useful starting point for formulating a differential diagnosis and considering possible causes. In support of the aim toward a cause-based classification system of small airway disease, a simple diagnostic algorithm is provided for further subclassification of the above 3 bronchiolitis patterns according to the major associated etiologic subgroups., Competing Interests: Disclosure The authors have nothing to disclose., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: Pathological gastroesophageal reflux (GER) is a known risk factor for bronchiolitis obliterans syndrome (BOS) after lung transplantation. This study aimed at determining whether functional esophageal evaluation might predict BOS occurrence and survival in this setting., Methods: Ninety-three patients who underwent esophageal high-resolution manometry and 24-hour pH-impedance monitoring within the first year after lung transplantation were retrospectively included. A univariable analysis was performed to evaluate the parameters associated with GER disease and BOS occurrence. The Cox regression model was used to identify the prognostic factors of death or retransplantation., Results: Thirteen percent of patients exhibited major esophageal motility disorders and 20% pathological GER. GER occurrence was associated with younger age, cystic fibrosis, and hypotensive esophagogastric junction. Within a median follow-up of 62 months, 10 patients (11%) developed BOS, and no predictive factors were identified. At the end of the follow-up, 10 patients died and 1 underwent retransplantation. The 5-year cumulative survival rate without retransplantation was lower in patients with major esophageal motility disorders compared with that in those without (75% vs 90%, P = 0.01) and in patients who developed BOS compared with that in those without (66% vs 91%; P = 0.005). However, in multivariable analysis, major esophageal motility disorders and BOS were no longer significant predictors of survival without retransplantation., Discussion: Major esophageal motility disorders and BOS were associated with allograft survival in lung transplantation in the univariable analysis. Although the causes of this association remain to be determined, this observation confirms that esophageal motor dysfunction should be evaluated in the context of lung transplantation.

Introduction: Long-term pulmonary complications are one of the major long-term consequences of sulfur mustard (SM) exposure. Toll-like receptor 4 (TLR4) involves in the pathogenesis of several pulmonary disorders. Surfactant protein-A (SP-A) regulates LPS-induced TLR4 localization and activation responses. However, the intensity and significance of TLR4 and SP-A expression by lung cells in SM-exposed patients is not clear., Methods: The gene expression of TLR4 (through real-time PCR) and TLR4 and SP-A positive cells and alveolar type II cells, as SP-A producers, (using IHC) were assessed in formalin fixed paraffin embedded (FFPE) specimens from SM-exposed (n = 17), and non-SM exposed individuals (n = 12)., Results: TLR4 gene expression did not change between study groups. However, its cell surface presentation was significantly reduced in SM-exposed patients and particularly in which with constrictive bronchiolitis compared with the control group (P < 0.001 and P = 0.002, respectively). Frequency of alveolar type II cells was lower in the case group rather than the control group while the number of SP-A positive cells did not alter., Conclusions: These findings suggest that reduced TLR4 cell surface presentation may have anti-inflammatory function and SP-A may have a critical role in regulation of inflammatory responses in SM-exposed patients. Further investigation on other possible mechanisms involved in TLR4 internalization maybe help to illustrate the modulatory or inflammatory activity of TLR4 in these patients., Competing Interests: Declaration of Competing Interest All of the authors declare no conflict of interest., (Copyright © 2019. Published by Elsevier B.V.)

Background: Although electronic cigarettes (e-cigarettes) were initially marketed as a potential smoking-cessation aid and a safer alternative to smoking, the long-term health effect of e-cigarette use ("vaping") is unknown. Vaping e-liquids expose the user to several potentially harmful chemicals, including diacetyl, a flavouring compound known to cause bronchiolitis obliterans with inhalational exposure ("popcorn worker's lung")., Case Description: We report the case of a 17-year-old male who presented with intractable cough, progressive dyspnea and malaise after vaping flavoured e-liquids and tetrahydrocannabinol intensively. Initial physical examination showed fever, tachycardia, hypoxemia, and bibasilar inspiratory crackles on lung auscultation. Computed tomography of the chest showed diffuse centrilobular "tree-inbud" nodularity, consistent with acute bronchiolitis. Multiple cultures, including from 2 bronchoalveolar lavage samples, and biopsy stains, were negative for infection. He required intubation, invasive mechanical ventilation and venovenous extracorporeal membrane oxygenation (ECMO) for refractory hypercapnia. The patient's condition improved with high-dose corticosteroids. He was weaned off ECMO and mechanical ventilation, and discharged home after 47 days in hospital. Several months after hospital discharge, his exercise tolerance remained limited and pulmonary function tests showed persistent, fixed airflow obstruction with gas trapping. The patient's clinical picture was suggestive of possible bronchiolitis obliterans, thought to be secondary to inhalation of flavouring agents in the e-liquids, although the exact mechanism of injury and causative agent are unknown., Interpretation: This case of severe acute bronchiolitis, causing near-fatal hypercapnic respiratory failure and chronic airflow obstruction in a previously healthy Canadian youth, may represent vaping-associated bronchiolitis obliterans. This novel pattern of pulmonary disease associated with vaping appears distinct from the type of alveolar injury predominantly reported in the recent outbreak of cases of vaping-associated pulmonary illness in the United States, underscoring the need for further research into all potentially toxic components of e-liquids and tighter regulation of e-cigarettes., Competing Interests: Competing interests: None declared., (© 2019 Joule Inc. or its licensors.)

Flavorings-related lung disease is a potentially disabling and sometimes fatal lung disease of workers making or using flavorings. First identified almost 20 years ago in microwave popcorn workers exposed to butter-flavoring vapors, flavorings-related lung disease remains a concern today. In some cases, workers develop bronchiolitis obliterans, a severe form of fixed airways disease. Affected workers have been reported in microwave popcorn, flavorings, and coffee production workplaces. Volatile α-dicarbonyl compounds, particularly diacetyl (2,3-butanedione) and 2,3-pentanedione, are implicated in the etiology. Published studies on diacetyl and 2,3-pentanedione document their ability to cause airway epithelial necrosis, damage biological molecules, and perturb protein homeostasis. With chronic exposure in rats, they produce airway fibrosis resembling bronchiolitis obliterans. To add to this knowledge, we recently evaluated airway toxicity of the 3-carbon α-dicarbonyl compound, methylglyoxal. Methylglyoxal inhalation causes epithelial necrosis at even lower concentrations than diacetyl. In addition, we investigated airway toxicity of mixtures of diacetyl, acetoin, and acetic acid, common volatiles in butter flavoring. At ratios comparable to workplace scenarios, the mixtures or diacetyl alone, but not acetic acid or acetoin, cause airway epithelial necrosis. These new findings add to existing data to implicate α-dicarbonyl compounds in airway injury and flavorings-related lung disease.

Collagen Tissue Disease-associated Interstitial Lung Fibrosis (CTD-ILDs) and Bronchiolitis Obliterans Syndrome (BOS) represent severe lung fibrogenic disorders, characterized by fibro-proliferation with uncontrolled extracellular matrix deposition. Hyaluronic acid (HA) plays a key role in fibrosis with its specific receptor, CD44, overexpressed by CTD-ILD and BOS cells. The aim is to use HA-liposomes to develop an inhalatory treatment for these diseases. Liposomes with HA of two molecular weights were prepared and characterized. Targeting efficiency was assessed toward CTD-ILD and BOS cells by flow cytometry and confocal microscopy and immune modulation by RT-PCR and ELISA techniques. HA-liposomes were internalized by CTD-ILD and BOS cells expressing CD44, and this effect increased with higher HA MW. In THP-1 cells, HA-liposomes decreased pro-inflammatory cytokines IL-1β, IL-12, and anti-fibrotic VEGF transcripts but increased TGF-β mRNA. However, upon analyzing TGF-β release from healthy donors-derived monocytes, we found liposomes did not alter the release of active pro-fibrotic cytokine. All liposomes induced mild activation of neutrophils regardless of the presence of HA. HA liposomes could be also applied for lung fibrotic diseases, being endowed with low pro-inflammatory activity, and results confirmed that higher MW HA are associated to an increased targeting efficiency for CD44 expressing LFs-derived from BOS and CTD-ILD patients.

Chronic lung allograft dysfunction (CLAD) still remains a major drawback in the outcome following lung transplantation (LTx). New therapeutic strategies are warranted. Growth factors and their receptors like platelet-derived growth factor-receptor (PDGFR) and vascular endothelial growth factor-receptor (VEGFR), may play a crucial role in the development of CLAD, especially bronchiolitis obliterans (BO) and vasculopathy. In this study, we used an orthotopic left lung transplantation model from Fischer (F344) to Wystar Kyoto (WKY) rats to investigate the effect of the receptor tyrosine kinase inhibitor (RTKI) vatalanib alone, the dual combination of vatalanib and imatinib and a triple therapy consisting of vatalanib, imatinib and the mammalian target of rapamycin inhibitor (mTORI) everolimus on the development of CLAD after LTx in rats. With this trial we demonstrated that monotherapy with vatalanib attenuated mild and severe chronic vascular rejection, whereas dual therapy (vatalanib and imatinib) after LTx also showed a significant reduction of chronic bronchiolar rejection and interstitial fibrosis. By adding everolimus, the effect of vatalanib and imatinib could additionally be increased. In conclusion, the combination of mTORI and RTKIs might be a possible strategy in the prevention of CLAD and BO.

Objective: To evaluate the diagnostic value of function volume/morphological volume ratio calculated from ventilation/perfusion single-photon emission computed tomography/computed tomography in distinguishing the lungs with bronchiolitis obliterans syndrome (BOS) from the lungs without this syndrome after lung transplantation and to assess its relationship with spirometry parameters., Materials and Methods: We retrospectively identified 84 consecutive lung transplant recipients and 13 donors who underwent ventilation/perfusion single-photon emission computed tomography/computed tomography. Differences in the function volume/morphological volume ratio of unilateral lungs were tested for significance between the lungs with and without BOS. Receiver operating characteristics and correlations between function volume/morphological volume ratios of bilateral lungs and forced expiratory volume in 1 s, forced vital capacity, and total lung capacity were analyzed., Results: The function volume/morphological volume ratios of ventilation and perfusion images of unilateral lungs were significantly lower in lungs with BOS (each P<0.0001). The area under the curve values of ventilation and perfusion images were 0.97 and 0.99, respectively. Significant correlations were identified between the function volume/morphological volume ratios of ventilation and perfusion images and forced expiratory volume in 1 s (r=0.54, P<0.0001 and r=0.45, P<0.0001, respectively). The function volume/morphological volume ratio of ventilation image had a significantly weak correlation with forced vital capacity., Conclusion: The function volume/morphological volume ratio enables a semiquantitative assessment of ventilation and perfusion lung functions and is useful for diagnosing BOS after lung transplantation.

A 63-year-old man presented with persistent cough and progressive dyspnea. Computed tomography showed irregular pleural thickening and fibrotic changes with volume loss in the upper lobes, and subtle reticulation in the lower lobes. Pleuroparenchymal fibroelastosis (PPFE) was diagnosed based on the findings of a surgical lung biopsy. Bronchiolar lesions, including proliferative bronchiolitis, constrictive bronchiolitis obliterans, and peribronchiolar metaplasia were evident on pathology. A usual interstitial pneumonia (UIP) pattern was also observed in the lower lobes. Three weeks after the biopsy, an acute exacerbation occurred. We herein describe a rare case of idiopathic PPFE with various bronchiolar lesions and a UIP pattern in which an acute exacerbation developed.

Purpose of Review: Lung transplantation (LTx) can be a life saving treatment in end-stage pulmonary diseases, but survival after transplantation is still limited. Posttransplant development of chronic lung allograft dysfunction with bronchiolits obliterans syndrome (BOS) as the major subphenotype, is the main cause of morbidity and mortality. Early identification of high-risk patients for BOS is a large unmet clinical need. In this review, we discuss gene polymorphisms and gene expression related to the development of BOS., Recent Findings: Candidate gene studies showed that donor and recipient gene polymorphisms affect transplant outcome and BOS-free survival after LTx. Both selective and nonselective gene expression studies revealed differentially expressed fibrosis and apoptosis-related genes in BOS compared with non-BOS patients. Significantly, recent microarray expression analysis of blood and broncho-alveolar lavage suggest a role for B-cell and T-cell responses prior to the development of BOS. Furthermore, 6 months prior to the development of BOS differentially expressed genes were identified in peripheral blood cells., Summary: Genetic polymorphisms and gene expression changes are associated with the development of BOS. Future genome wide studies are needed to identify easily accessible biomarkers for prediction of BOS toward precision medicine.

Obliterative bronchiolitis (OB) is a poorly understood airway disease characterized by the generation of fibrotic bronchiolar occlusions. In the lung transplant setting, OB is a pathological manifestation of bronchiolitis obliterans syndrome (BOS), which is a major impediment to long-term recipient survival. Club cells play a key role in bronchiolar epithelial repair, but whether they promote lung transplant tolerance through preventing OB remains unclear. We determined if OB occurs in mouse orthotopic lung transplants following conditional transgene-targeted club cell depletion. In syngeneic lung transplants club cell depletion leads to transient epithelial injury followed by rapid club cell-mediated repair. In contrast, allogeneic lung transplants develop severe OB lesions and poorly regenerate club cells despite immunosuppression treatment. Lung allograft club cell ablation also triggers the recognition of alloantigens, and pulmonary restricted self-antigens reported associated with BOS development. However, CD8+ T cell depletion restores club cell reparative responses and prevents OB. In addition, ex-vivo analysis reveals a specific role for alloantigen-primed effector CD8+ T cells in preventing club cell proliferation and maintenance. Taken together, we demonstrate a vital role for club cells in maintaining lung transplant tolerance and propose a new model to identify the underlying mechanisms of OB.