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Controlled hypothermic storage (CHS) is a recent advance in lung transplantation (LTx) allowing preservation at temperatures higher than those achieved with traditional ice storage. The mechanisms explaining the benefits of CHS compared to conventional static ice storage (SIS) remain unclear and clinical data on safety and feasibility of lung CHS are limited. Therefore, we aimed to provide a focus review on animal experiments, molecular mechanisms, CHS devices, current clinical experience, and potential future benefits of CHS. Rabbit, canine and porcine experiments showed superior lung physiology after prolonged storage at 10°C vs. =4°C. In recent molecular analyses of lung CHS, better protection of mitochondrial health and higher levels of antioxidative metabolites were observed. The acquired insights into the underlying mechanisms and development of CHS devices allowed clinical application and research using CHS for lung preservation. The initial findings are promising; however, further data collection and analysis are required to draw more robust conclusions. Extended lung preservation with CHS may provide benefits to both recipients and healthcare personnel. Reduced time pressure between procurement and transplantation introduces flexibility allowing better decision-making and overnight bridging by delaying transplantation to daytime without compromising outcome. [ABSTRACT FROM AUTHOR]

To improve outcomes following lung transplantation, it is essential to understand the immunological mechanisms that result in chronic graft failure. The associated clinical syndrome is termed chronic lung allograft dysfunction (CLAD), which is known to be induced by alloimmune-dependent (i.e., rejection) and alloimmune-independent factors (e.g., infections, reflux and environmental factors). We aimed to explore the alloimmune-related mechanism, i.e., pulmonary rejection. In this study, we use a murine orthotopic left lung transplant model using isografts and allografts (C57BL/6 or BALB/c as donors to C57BL/6 recipients), with daily immunosuppression (10 mg/kg cyclosporin A and 1.6 mg/kg methylprednisolone). Serial sacrifice was performed at days 1, 7 and 35 post-transplantation (n = 6 at each time point for each group). Left transplanted lungs were harvested, a single-cell suspension was made and absolute numbers of immune cells were quantified using multicolor flow cytometry. The rejection process followed the principles of a classic immune response, including innate but mainly adaptive immune cells. At day 7 following transplantation, the numbers of interstitial macrophages, monocytes, dendritic cells, NK cells, NKT cells, CD4+ T cells and CD8+ T and B cells were increased in allografts compared with isografts. Only dendritic cells and CD4+ T cells remained elevated at day 35 in allografts. Our study provides insights into the immunological mechanisms of true pulmonary rejection after murine lung transplantation. These results might be important in further research on diagnostic evaluation and treatment for CLAD. [ABSTRACT FROM AUTHOR]

Lung transplantation (LTx) is a challenging procedure. Following the process of ischemia-reperfusion injury, the transplanted pulmonary graft might become severely damaged, resulting in primary graft dysfunction. In addition, during the intraoperative window, the right ventricle (RV) is at risk of acute failure. The interaction of right ventricular function with lung injury is, however, poorly understood. We aimed to address this interaction in a translational porcine model of pulmonary ischemia-reperfusion injury. Advanced pulmonary and hemodynamic assessment was used, including right ventricular pressure-volume loop analysis. The acute model was based on clamping and unclamping of the left lung hilus, respecting the different hemodynamic phases of a clinical lung transplantation. We found that forcing entire right ventricular cardiac output through a lung suffering from ischemia-reperfusion injury increased afterload (pulmonary vascular resistance from baseline to end experiment P < 0.0001) and induced right ventricular failure (RVF) in 5/9 animals. Notably, we identified different compensation patterns in failing versus nonfailing ventricles (arterial elastance P = 0.0008; stroke volume P < 0.0001). Furthermore, increased vascular pressure and flow produced by the right ventricle resulted in higher pulmonary injury, as measured by ex vivo CT density (correlation: pressure r = 0.8; flow r = 0.85). Finally, RV ischemia as measured by troponin-T was negatively correlated with pulmonary injury (r = -0.76); however, troponin-T values did not determine RVF in all animals. In conclusion, we demonstrate a delicate balance between development of pulmonary ischemia-reperfusion injury and right ventricular function during lung transplantation. Furthermore, we provide a physiological basis for potential benefit of extracorporeal life support technology. NEW & NOTEWORTHY: In contrast to the abundant literature of mechanical pulmonary artery clamping to increase right ventricular afterload, we developed a model adding a biological factor of pulmonary ischemia-reperfusion injury. We did not only focus on the right ventricular behavior, but also on the interaction with the injured lung. We are the first to describe this interaction while addressing the hemodynamic intraoperative phases of clinical lung transplantation. [ABSTRACT FROM AUTHOR]

To date, investigations of the microbiota in the lungs of people with Cystic Fibrosis (PWCF) have primarily focused on microbial community composition in luminal mucus, with fewer studies observing the microbiota in tissue samples from explanted lung tissue. Here, we analysed both tissue and airway luminal mucus samples extracted from whole explanted lungs of PWCF and unused donor lungs. We determined if the lung microbiota in end-stage CF varied within and between patients, was spatially heterogeneous and related to localized structural damage. Microbial community composition was determined by Illumina MiSeq sequencing and related to the CF-Computed Tomography (CT) score and features of end-stage lung disease on micro-CT. Ninety-eight CF tissue (n=11 patients), 20 CF luminal mucus (n=8 patients) and 33 donor tissue (n=4 patients) samples were analysed. Additionally, we compared 20 paired CF tissue and luminal mucus samples that enabled a direct "geographical" comparison of the microbiota in these two niches. Significant differences in microbial communities were apparent between the 3 groups. However, overlap between the three groups, particularly between CF and donor tissue and CF tissue and CF luminal mucus was also observed. Microbial diversity was lower in CF luminal mucus compared to CF tissue, with dominance higher in luminal mucus. For both CF and donor tissue, intra- and inter-patient variability in ecological parameters was observed. No relationships were observed between ecological parameters and CF-CT score, or features of end-stage lung disease. The end-stage CF lung is characterised by a low diversity microbiota, differing within and between individuals. No clear relationship was observed between regional microbiota variation and structural lung damage. [ABSTRACT FROM AUTHOR]

Since microRNA (miR)-223-3p modulates inflammatory responses and chronic obstructive pulmonary disease (COPD) is associated with amplified pulmonary inflammation, we hypothesized that miR-223-3p plays a role in COPD pathogenesis. Expression of miR-223-3p was measured in lung tissue of two independent cohorts with patients with GOLD stage II-IV COPD, never smokers, and smokers without COPD. The functional role of miR-223-3p was studied in deficient mice and on overexpression in airway epithelial cells from COPD and controls. We observed higher miR-223-3p levels in patients with COPD stage II-IV compared with (non)-smoking controls, and levels were associated with higher neutrophil numbers in bronchial biopsies of patients with COPD. MiR-223-3p expression was also increased in lungs and bronchoalveolar lavage of cigarette smoke (CS)-exposed mice. CS-induced neutrophil and monocyte lung infiltration was stronger in miR-223-deficient mice on acute (5 days) exposure, but attenuated on subchronic (4 wk) exposure. Additionally, miR-223 deficiency attenuated acute and subchronic CS-induced lung infiltration of dendritic cells and T lymphocytes. Finally, in vitro overexpression of miR-223-3p in non-COPD airway epithelial cells suppressed C-X-C motif chemokine ligand 8 (CXCL8) and granulocyte monocyte-colony stimulation factor (GM-CSF) secretion and gene expression of the proinflammatory transcription factor TRAF6. Importantly, this suppressive effect of miR-223-3p was compromised in COPD-derived cultures. In conclusion, we demonstrate that miR-223-3p is increased in lungs of patients with COPD and CS-exposed mice and is associated with neutrophilic inflammation. In vivo data indicate that miR-223 acts as negative regulator of acute CS-induced neutrophilic and monocytic inflammation. In vitro data suggest that miR-223-3p does so by suppressing proinflammatory airway epithelial responses, which is less effective in COPD epithelium. [ABSTRACT FROM AUTHOR]

Background: Connective tissue growth factor (CTGF) is an important mediator in several fibrotic diseases, including lung fibrosis. We investigated CTGF-expression in chronic lung allograft dysfunction (CLAD) and pulmonary graft-versus-host disease (GVHD). Materials and Methods: CTGF expression was assessed by quantitative real-time polymerase chain reaction (qPCR) and immunohistochemistry in end-stage CLAD explant lung tissue (bronchiolitis obliterans syndrome (BOS), n=20; restrictive allograft syndrome (RAS), n=20), pulmonary GHVD (n=9). Unused donor lungs served as control group (n=20). Next, 60 matched lung transplant recipients (BOS, n=20; RAS, n=20; stable lung transplant recipients, n=20) were included for analysis of CTGF protein levels in plasma and broncho-alveolar lavage (BAL) fluid at 3 months post-transplant, 1 year post-transplant, at CLAD diagnosis or 2 years post-transplant in stable patients. Results: qPCR revealed an overall significant difference in the relative content of CTGF mRNA in BOS, RAS and pulmonary GVHD vs. controls (p=0.014). Immunohistochemistry showed a significant higher percentage and intensity of CTGF-positive respiratory epithelial cells in BOS, RAS and pulmonary GVHD patients vs. controls (p<0.0001). BAL CTGF protein levels were significantly higher at 3 months post-transplant in future RAS vs. stable or BOS (p=0.028). At CLAD diagnosis, BAL protein content was significantly increased in RAS patients vs. stable (p=0.0007) and BOS patients (p=0.042). CTGF plasma values were similar in BOS, RAS, and stable patients (p=0.74). Conclusions: Lung CTGF-expression is increased in end-stage CLAD and pulmonary GVHD; and higher CTGF-levels are present in BAL of RAS patients at CLAD diagnosis. Our results suggest a potential role for CTGF in CLAD, especially RAS, and pulmonary GVHD. [ABSTRACT FROM AUTHOR]

Physiological lung ageing is associated with a gradual decline in dynamic lung volumes and a progressive increase in residual volume due to diminished elastic recoil of the lung, loss of alveolar tissue, and lower chest wall compliance. However, the effects of ageing on the small airways (ie, airways <2·0 mm in diameter) remain largely unknown. By using a combination of ex-vivo conventional CT (resolution 1 mm), whole lung micro-CT (resolution 150 μm), and micro-CT of extracted cores (resolution 10 μm), we aimed to provide a multiresolution assessment of the small airways in lung ageing in a large cohort of never smokers. For this cross-sectional study, we included donor lungs collected from 32 deceased never-smoking donors (age range 16–83 years). Ex-vivo CT and whole lung high-resolution CT (micro-CT) were used to determine total airway numbers, stratified by airway diameter. Micro-CT was used to assess the number, length, and diameter of terminal bronchioles (ie, the last generation of conducting airways); mean linear intercept; and surface density in four lung tissue cores from each lung, extracted using a uniform sampling approach. Regression β coefficients are calculated using linear regression and polynomial models. Ex-vivo CT analysis showed an age-dependent decrease in the number of airways of diameter 2·0 mm to less than 2·5 mm (β coefficient per decade −0·119, 95% CI −0·193 to −0·045; R 2=0·29) and especially in airways smaller than 2·0 mm in diameter (−0·158, −0·233 to −0·084; R 2=0·47), between 30 and 80 years of age, but not of the larger (≥2·5 mm) diameter airways (−0·00781, −0·04409 to 0·02848; R 2=0·0007). In micro-CT analysis of small airways, the total number of terminal bronchioles per lung increased until the age of 30 years, after which an almost linear decline in the number of terminal bronchioles was observed (β coefficient per decade −2035, 95% CI −2818 to −1252; R 2=0·55), accompanied by a non-significant increase in alveolar airspace size (6·44, −0·57 to 13·45, R 2=0·10). Moreover, this decrease in terminal bronchioles was associated with the age-related decline of pulmonary function predicted by healthy reference values. Loss of terminal bronchioles is an important structural component of age-related decline in pulmonary function of healthy, non-smoking individuals. Research Foundation—Flanders, KU Leuven, Parker B Francis Foundation, UGent, Canadian Institutes for Health. [ABSTRACT FROM AUTHOR]

Background: Idiopathic pulmonary fibrosis (IPF) is a severe lung disease characterised by extensive pathological changes. The objective for this study was to identify the gene network and regulators underlying disease pathology in IPF and its association with lung function.Methods: Lung Tissue Research Consortium dataset with 262 IPF and control subjects (GSE47460) was randomly divided into two non-overlapping groups for cross-validated differential gene expression analysis. Consensus weighted gene coexpression network analysis identified overlapping coexpressed gene modules between both IPF groups. Modules were correlated with lung function (diffusion capacity, DLCO; forced expiratory volume in 1 s, FEV1; forced vital capacity, FVC) and enrichment analyses used to identify biological function and transcription factors. Module correlation with miRNA data (GSE72967) identified associated regulators. Clinical relevance in IPF was assessed in a peripheral blood gene expression dataset (GSE93606) to identify modules related to survival.Results: Correlation network analysis identified 16 modules in IPF. Upregulated modules were associated with cilia, DNA replication and repair, contractile fibres, B-cell and unfolded protein response, and extracellular matrix. Downregulated modules were associated with blood vessels, T-cell and interferon responses, leucocyte activation and degranulation, surfactant metabolism, and cellular metabolic and catabolic processes. Lung function correlated with nine modules (eight with DLCO, five with FVC). Intermodular network of transcription factors and miRNA showed clustering of fibrosis, immune response and contractile modules. The cilia-associated module was able to predict survival (p=0.0097) in an independent peripheral blood IPF cohort.Conclusions: We identified a correlation gene expression network with associated regulators in IPF that provides novel insight into the pathological process of this disease. [ABSTRACT FROM AUTHOR]

Rationale: Aberrant expression of microRNAs (miRNAs) can have a detrimental role in disease pathogenesis.Objectives: To identify dysregulated miRNAs in lung tissue of patients with chronic obstructive pulmonary disease (COPD).Methods: We performed miRNA and mRNA profiling using high throughput stem-loop reverse-transcriptase quantitative polymerase chain reaction and mRNA microarray, respectively, on lung tissue of 30 patients (screening cohort) encompassing 8 never-smokers, 10 smokers without airflow limitation, and 12 smokers with COPD. Differential expression of miRNA-218-5p (miR-218-5p) was validated by reverse-transcriptase quantitative polymerase chain reaction in an independent cohort of 71 patients, an in vivo murine model of COPD, and primary human bronchial epithelial cells. Localization of miR-218-5p was assessed by in situ hybridization. In vitro and in vivo perturbation of miR-218-5p combined with RNA sequencing and gene set enrichment analysis was used to elucidate its functional role in COPD pathogenesis.Measurements and Main Results: Several miRNAs were differentially expressed among the different patient groups. Interestingly, miR-218-5p was significantly down-regulated in smokers without airflow limitation and in patients with COPD compared with never-smokers. Decreased pulmonary expression of miR-218-5p was validated in an independent validation cohort, in cigarette smoke-exposed mice, and in human bronchial epithelial cells. Importantly, expression of miR-218-5p strongly correlated with airway obstruction. Furthermore, cellular localization of miR-218-5p in human and murine lung revealed highest expression of miR-218-5p in the bronchial airway epithelium. Perturbation experiments with a miR-218-5p mimic or inhibitor demonstrated a protective role of miR-218-5p in cigarette smoke-induced inflammation and COPD.Conclusions: We highlight a role for miR-218-5p in the pathogenesis of COPD. [ABSTRACT FROM AUTHOR]

Bronchiolitis obliterans syndrome (BOS) remains a major complication after lung transplantation, causing significant morbidity and mortality in a majority of recipients. BOS is believed to be the clinical correlate of chronic allograft dysfunction, and is defined as an obstructive pulmonary function defect in the absence of other identifiable causes, mostly not amenable to treatment. Recently, it has become clear that BOS is not the only form of chronic allograft dysfunction and that other clinical phenotypes exist; however, we focus exclusively on BOS. Radiologic findings typically demonstrate air trapping, mosaic attenuation, and hyperinflation. Pathologic examination reveals obliterative bronchiolitis lesions and a pure obliteration of the small airways (< 2 mm), with a relatively normal surrounding parenchyma. In this review, we highlight recent advances in diagnosis, pathologic examination, and risk factors, such as microbes, viruses, and antibodies. Although the pathophysiological mechanisms remain largely unknown, we review the role of the airway epithelium and inflammation and the various experimental animal models. We also clarify the clinical and therapeutic implications of these findings. Although significant progress has been made, the exact pathophysiological mechanisms and adequate therapy for posttransplantation BOS remain unknown, highlighting the need for further research to improve long-term posttransplantation BOS-free and overall survival. [ABSTRACT FROM AUTHOR]

Invasive lung function measurements are useful tools to describe respiratory disease models in mice but only result in one time-point measurements because of tracheostomy. We explored if intubation may overcome the need for tracheostomy thereby allowing invasive lung function monitoring of individual mice over time. Repeated invasive lung function measurements with Scireq© FlexiVent or Buxco© - Forced Pulmonary Maneuvers were performed three times in BALB/c mice with intervals of 10 days. Each lung function assessment following intubation was compared with a similar measurement in age-matched tracheostomized mice, the golden standard in lung function measurements. Tracheostomy and intubation gave similar results for resistance, elastance and compliance of the whole respiratory system as assessed by Flexivent. Likewise, Forced Pulmonary Maneuvers used to measure lung volumes such as total lung capacity, functional residual capacity, forced expiratory volume in 0.1 s and forced vital capacity, resulted in identical outcomes for both airway approaches. No interaction was found between the procedures for any of the pulmonary function variables. The observed changes over time were rather related to animal growth than to repetitive intubation. Eighty percent of the animals survived three consecutive intubations, which were hampered by transient breathing difficulties, weight loss and neutrophilic bronchoalveolar lavage immediately postextubation. Repetitive invasive lung function measurements by intubation are feasible and reproducible in healthy mice and results are comparable to the standard method. This may open new perspectives for longitudinal research in animal models of respiratory diseases. [ABSTRACT FROM AUTHOR]

Primary graft dysfunction (PGD) is the clinical syndrome of acute lung injury after lung transplantation (LTx). However, PGD is an umbrella term that encompasses the ongoing pathophysiological and -biological mechanisms occurring in the lung grafts. Therefore, we aim to provide a focused review on the clinical, physiological, radiological, histological and cellular level of PGD. PGD is graded based on hypoxemia and chest X-ray (CXR) infiltrates. High-grade PGD is associated with inferior outcome after LTx. Lung edema is the main characteristic of PGD and alters pulmonary compliance, gas exchange and circulation. A conventional CXR provides a rough estimate of lung edema, while a chest computed tomography (CT) results in a more in-depth analysis. Macroscopically, interstitial and alveolar edema can be distinguished below the visceral lung surface. On the histological level, PGD correlates to a pattern of diffuse alveolar damage (DAD). At the cellular level, ischemia-reperfusion injury (IRI) is the main trigger for the disruption of the endothelial-epithelial alveolar barrier and inflammatory cascade. The multilevel approach integrating all PGD-related aspects results in a better understanding of acute lung failure after LTx, providing novel insights for future therapies. [ABSTRACT FROM AUTHOR]

Chronic rejection of lung allografts has two major subtypes, bronchiolitis obliterans syndrome (BOS) and restrictive allograft syndrome (RAS), which present radiologically either as air trapping with small airways disease or with persistent pleuroparenchymal opacities. Parametric response mapping (PRM), a computed tomography (CT) methodology, has been demonstrated as an objective readout of BOS and RAS and bears prognostic importance, but has yet to be correlated to biological measures. Using a topological technique, we evaluate the distribution and arrangement of PRM-derived classifications of pulmonary abnormalities from lung transplant recipients undergoing redo-transplantation for end-stage BOS (N = 6) or RAS (N = 6). Topological metrics were determined from each PRM classification and compared to structural and biological markers determined from microCT and histopathology of lung core samples. Whole-lung measurements of PRM-defined functional small airways disease (fSAD), which serves as a readout of BOS, were significantly elevated in BOS versus RAS patients (p = 0.01). At the core-level, PRM-defined parenchymal disease, a potential readout of RAS, was found to correlate to neutrophil and collagen I levels (p < 0.05). We demonstrate the relationship of structural and biological markers to the CT-based distribution and arrangement of PRM-derived readouts of BOS and RAS. [ABSTRACT FROM AUTHOR]

Clinical studies revealed that azithromycin reduces airway neutrophilia during chronic rejection after lung transplantation. Our aim was to investigate the possible effect of azithromycin on ischaemia-reperfusion injury. Azithromycin or water was administered to mice every other day during 2 weeks ( n = 6/group). On the 14th day, the left lung was clamped to induce ischaemia (90 min). In two additional groups, animals underwent the same protocol, followed by 4 h of reperfusion. Two control groups were included with thoracotomy only. Inflammatory parameters and oxidative stress were measured in broncho-alveolar lavage of the left lung. Leukocytes, lymphocytes, neutrophils, 8-isoprostane and IL-1β levels after ischaemia and reperfusion were significantly reduced in mice treated with azithromycin. There was a trend towards lower IL-6 and KC levels. A significant correlation was seen between 8-isoprostanes and neutrophils (Pearson r = 0.72; P = 0.0086), IL-6 (Pearson r = 0.84; P = 0.0006), KC (Pearson r = 0.88; P = 0.0002) and IL-1β (Pearson r = 0.62; P = 0.0326). We conclude (i) that azithromycin reduces inflammation and oxidative stress in our IRI model, and (ii) that oxidative stress is correlated with the number of neutrophils and IL-6, KC and IL-1β levels after ischaemia and reperfusion. Azithromycin should be further investigated as a novel drug to prevent lung ischaemia-reperfusion injury. [ABSTRACT FROM AUTHOR]

Lung transplantation improves the outcome and quality of life of patients with end-stage pulmonary disease. However, the procedure is still hampered by the lack of suitable donors, the complexity of the surgery, and the risk of developing chronic lung allograft dysfunction. Over the past decades, translational experiments in animal models have led to a better understanding of physiology and immunopathology following the lung transplant procedure. Small animal models (e.g., rats and mice) are mostly used in experiments regarding immunology and pathobiology and are preferred over large animal models due to the ethical aspects, the cost–benefit balance, and the high throughput possibility. In this comprehensive review, we summarize the reported surgical techniques for lung transplantation in rodent models and the management of perioperative complications. Furthermore, we propose a guide to help identify the appropriate species for a given experiment and discuss recent experimental findings in small animal lung transplant models. [ABSTRACT FROM AUTHOR]

Bronchiolitis obliterans syndrome (BOS) is considered an airway-centered disease, with bronchiolitis obliterans (BO) as pathologic hallmark. However, the histologic spectrum of pure clinical BOS remains poorly characterized. We provide the first in-depth histopathologic description of well-characterized BOS patients and patients without chronic lung allograft dysfunction (CLAD), defined according to the recent consensus guidelines. Explant lung tissue from 52 clinically-defined BOS and 26 non-CLAD patients (collected 1993–2018) was analyzed for histologic parameters, including but not limited to airway lesions, vasculopathy and fibrosis. In BOS, BO lesions were evident in 38 (73%) patients and varied from concentric sub-epithelial fibrotic BO to inflammatory BO, while 10/14 patients without BO displayed 'vanishing airways', defined by a discordance between arteries and airways. Chronic vascular abnormalities were detected in 22 (42%) patients. Ashcroft fibrosis scores revealed a median of 43% (IQR: 23–69) of normal lung parenchyma per patient; 26% (IQR: 18–37) of minimal alveolar fibrous thickening; and 11% (IQR: 4–18) of moderate alveolar thickening without architectural damage. Patchy areas of definite fibrotic damage to the lung structure (i.e., Ashcroft score ≥5) were present in 28 (54%) patients. Fibrosis was classified as bronchocentric (n = 21/28, 75%), paraseptal (n = 17/28, 61%) and subpleural (n = 15/28, 54%). In non-CLAD patients, BO lesions were absent, chronic vascular abnormalities present in 1 (4%) patient and mean Ashcroft scores were significantly lower compared to BOS (p = 0.0038) with 78% (IQR: 64–88) normally preserved lung parenchyma. BOS explant lungs revealed evidence of various histopathologic findings, including vasculopathy and fibrotic changes, which may contribute to the pathophysiology of BOS. [ABSTRACT FROM AUTHOR]

Chronic lung diseases (CLDs) represent a set of disorders characterized by the progressive loss of proper lung function. Among severe CLDs, the incidence of chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF) has grown over the last decades, mainly in the elderly population. Several studies have highlighted an increased expression of senescence-related markers in the resident progenitor cells in COPD and IPF, possibly undermining epithelial integrity and contributing to the progression and the aggravation of both diseases. Recently, the chronic activation of the canonical Wnt/β-catenin pathway was shown to induce cellular senescence. Here, we investigated the localization and the expression of leucin-rich repeat-containing G-protein-coupled receptor 6 (LGR6), a protein that activates and potentiates the canonical Wnt signalling. Through immunohistochemical analyses, we identified a lesion-associated rise in LGR6 levels in abnormal lung epithelial progenitors in COPD and IPF when compared to histologically normal tissues. Moreover, in areas of aberrant regeneration, chronic damage and fibrosis, LGR6-expressing epithelial progenitors displayed a major increase in the expression of senescence-associated markers. Our study suggests the involvement of LGR6 in the chronic activation of the Wnt/β-catenin pathway, mediating the impairment and exhaustion of epithelial progenitors in COPD and IPF. [ABSTRACT FROM AUTHOR]

Rationale: Patients can change chronic lung allograft dysfunction (CLAD) phenotype, especially from BOS to mixed phenotype. Our aim was to further characterize these patients. Method: Mixed CLAD was defined as a restrictive physiology with persistent CT opacities, after initial bronchiolitis obliterans syndrome (BOS) diagnosis. The incidence, prognosis, pulmonary function, radiology, pathology, and airway inflammation were compared between patients with restrictive allograft syndrome (RAS) and mixed CLAD. Result: A total of 268 (44%) patients developed CLAD of which 47 (18%) were diagnosed with RAS "ab initio," 215 (80%) with BOS, and 6 (2%) an undefined phenotype. Twenty‐five patients developed a mixed CLAD phenotype (24 BOS to mixed and 1 RAS to mixed). Survival after mixed phenotype diagnosis was comparable (P =.39) to RAS. More emphysema patients developed a mixed phenotype (P =.020) compared to RAS ab initio, while mixed CLAD patients had a lower FEV1 (P <.0001) and FEV1/FVC (P =.0002) at diagnosis compared to RAS ab initio. CT scans in patients with the mixed phenotype demonstrated apical predominance of the opacities (P =.0034) with pleuroparenchymal fibroelastosis on histopathology. Conclusion: We further characterized patients with a mixed phenotype of CLAD. Although the survival after diagnosis was comparable to RAS ab initio patients, there was a difference in demography, pulmonary function, radiology, and pathology. [ABSTRACT FROM AUTHOR]

Background: Cystic fibrosis (CF) lung disease is characterised by vigorous airway inflammation eventually resulting in severe lung damage. This study aimed to describe the diversity of the inflammatory pattern in end-stage CF lungs by evaluating and quantifying which components of the innate and adaptive immunity are involved, and by assessing whether this is gender-specific.Methods: CF explant lung tissue (n = 20) collected at time of transplantation and control tissue (n = 22) was sectioned (9 μm) and stained for neutrophils, eosinophils, mast cells, dendritic cells, macrophages, CD4 T cells, cytotoxic T cells and B cells. Quantification with special attention for immune cell location was performed.Results: Neutrophils, mast cells, dendritic cells, macrophages, CD4 T and cytotoxic T cells were significantly increased in CF compared to controls and there was a disproportionate increase of neutrophils around the airways in CF. Large amounts of lymphoid follicles were found in the CF lung and they had a skewed B cell/T cell composition. Upon subdividing the CF patients into a male and female population, eosinophils, mast cells and CD4 T cells were increased specifically in CF females. In this subpopulation, lymphoid follicles had less B cells and more CD8 T cells.Conclusion: These data demonstrate a diverse inflammatory response in the CF lung, reflected by an increase of both myeloid and lymphoid immune cells. Inflammation in the CF lung appeared to be gender-specific in our population, as the significant increase of eosinophils, mast cells and CD4 T cells was especially notable in the female subpopulation. [ABSTRACT FROM AUTHOR]

Chronic lung allograft dysfunction (CLAD) is a major cause of post‒lung transplant mortality, with limited medical treatment options. In this study we assessed the association of montelukast treatment with pulmonary function and outcome in lung transplant recipients with progressive CLAD. We performed a retrospective study of all lung transplant recipients transplanted between July 1991 and December 2016 at our center and who were treated for at least 3 months with montelukast for progressive CLAD, despite at least 3 months of prior azithromycin therapy. Main outcome parameters included evolution of pulmonary function and progression-free and overall survival. A total of 153 patients with CLAD (115 with bronchiolitis obliterans syndrome and 38 with restrictive allograft syndrome) were included, of whom 46% had a forced expiratory volume in 1 second (FEV 1) measure of between 66% and 80%, 31% an FEV 1 between 51% and 65%, and 23% an FEV 1 ≤50% of best post-operative FEV 1 at start of montelukast. Montelukast was associated with attenuation in rate of FEV 1 decline after 3 and 6 months, respectively (both p < 0.0001). Patients in whom FEV 1 improved or stabilized after 3 months of montelukast (81%) had significantly better progression-free (p < 0.0001) and overall (p = 0.0002) survival after CLAD onset, as compared to those with further decline of FEV 1 (hazard ratio [HR] 2.816, 95% confidence interval [CI] 1.450 to 5.467, p = 0.0022 for overall survival after CLAD onset in risk-adjusted multivariate analysis). Montelukast was associated with a significant attenuation in rate of FEV 1 decline in a substantial proportion of patients with established CLAD, which correlated with better outcome. Further study is required regarding use of montelkast. [ABSTRACT FROM AUTHOR]

BACKGROUND Chronic lung allograft dysfunction (CLAD) is the single most important factor limiting long-term survival after lung transplantation (LTx). Azithromycin has been shown to improve CLAD-free and long-term survival, yet the possible impact on early lung allograft function is unclear. METHODS A prospective, randomized, double-blind, placebo-controlled trial of pre-transplant and prompt post-transplant azithromycin treatment was performed at the University Hospitals Leuven. In each arm, 34 patients, transplanted between October 2013 and October 2015, were included for analysis. Study drug was added to standard of care and was administered once before LTx (1,000 mg of azithromycin or placebo) and every other day from Day 1 until Day 31 after LTx (250 mg of azithromycin or placebo). Primary outcome was an anticipated 15% improvement of forced expiratory volume in 1 second (FEV 1 , percent predicted) during the first 3 months post-LTx. Secondary end-points included length of intubation, days on ventilator, duration of intensive care unit and hospital stay, prevalence and severity of primary graft dysfunction, acute rejection, infection, and CLAD-free and overall survival. RESULTS FEV 1 was not significantly different between the 2 groups (p = 0.41). Patients treated with azithromycin demonstrated less airway inflammation, with lower bronchoalveolar lavage (BAL) neutrophilia and BAL interleukin-8 protein levels at Day 30 (p = 0.09 and p = 0.04, respectively) and Day 90 (p = 0.002 and p = 0.08, respectively) after LTx. Other secondary outcomes were not significantly different between placebo and azithromycin groups. CONCLUSIONS Pre-transplant and prompt post-transplant azithromycin treatment was not able to improve early lung allograft function. However, the known anti-inflammatory properties of azithromycin were confirmed (NCT01915082). [ABSTRACT FROM AUTHOR]

BACKGROUND Long-term survival after lung transplantation (LTx) is hampered by chronic lung allograft dysfunction (CLAD). Our study evaluated the prevalence and prognostic importance of obstructive and restrictive CLAD phenotypes, with or without an identifiable underlying cause, to validate the recently proposed classification system for CLAD. METHODS Data for patients who underwent LTx between 2004 and 2015 with a minimal survival of 180 days post-LTx were retrospectively collected. Double LTx patients with CLAD (defined as a persistent forced expiratory volume in 1 second decline of ≥ 20% compared with baseline) were subsequently classified according to obstructive (forced expiratory volume in 1 second /forced vital capacity [FVC] < 70%, total lung capacity > 90%, and FVC > 80%) or restrictive (total lung capacity ≤ 90% or FVC ≤ 80%) pulmonary function and to the presence of an unknown (bronchiolitis obliterans syndrome [BOS]/restrictive allograft syndrome [RAS]) or known (non-BOS/non-RAS) underlying cause. RESULTS After a median of 3.2 years, CLAD developed in 39% of double LTx patients (n = 219), of which 20% (n = 43) had an identifiable cause. Survival was worse in patients with restrictive CLAD (26%) compared with obstructive CLAD (64%; p < 0.0001). Non-BOS patients suffered from inferior survival compared with BOS patients (p = 0.0016), whereas there was no significant difference in survival between RAS and non-RAS (p = 0.17). Patients who evolved from an obstructive (BOS) to a restrictive (RAS) phenotype (10%) experienced better survival than RAS patients and a worse outcome compared with BOS patients (p < 0.0001). CONCLUSIONS Given the differences in outcome, accurate diagnosis of CLAD phenotypes is important, because this helps to inform patients about their prognosis, to reveal underlying pathogenesis, to identify homogenous patient populations for clinical trials, and to guide future therapeutic approaches. [ABSTRACT FROM AUTHOR]

Background: Lungs donated after cardiac death (DCD) may significantly reduce current organ shortage. However, the warm ischemic period following circulatory arrest may enhance ischemia-reperfusion injury (IRI). We investigated the possible therapeutic effect of N-acetyl cysteine (NAC), a potent anti-oxidative agent on IRI in a porcine ex vivo lung reperfusion model. Materials and methods: NAC (50 mg/kg) was nebulized to pigs (n = 6/group) prior to sacrifice (NAC-DCD). In DCD-NAC, animals received NAC 15 min after death. Control animals did not receive an aerosol (DCD). Interleukin (IL)-1β, tumor necrosis factor-alpha, IL-8, lactate dehydrogenase activity and thiobarbituric acid reactive substances were measured and cells were counted in broncho-alveolar lavage from the right lung after a 3-h warm plus 1-h cold ischemic interval. Results: There were no differences in cells between groups, however cell death was lower in NAC-DCD (10.3 ± 1.5%) and DCD-NAC (7.83 ± 1.8%) compared to DCD (18.0 ± 3.8%). IL-1β levels (111.5 ± 28.8 pg/mL and 92.2 ± 51.0 pg/mL versus 250.3 ± 56.6 pg/mL) and lactate dehydrogenase activity (1258.0 ± 440.9 U/L and 1606.0 ± 289.0 U/L versus 2848.0 ± 760.9 U/L) were significantly lower in NAC-DCD and DCD-NAC compared with DCD, respectively. These postischemic inflammatory markers correlated with functional parameters upon reperfusion of the left lung, reported in a previous study. Conclusions: Administration of NAC prior to or shortly after circulatory arrest results in a marked reduction of inflammation during the warm ischemic phase. [Copyright &y& Elsevier]