Title Promoter Affiliations Abstract "Histopathologic evaluation of the different fenotypes of chronic lung allograft dysfunction after lung transplantation" "Geert Verleden" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE), Translational Cell & Tissue Research" "Lung transplantation is a life-saving treatment option for patients with chronic end-stage lung diseases. Worldwide, more than 50,000 lung transplantations have been performed. However, survival remains hampered by the development of chronic rejection (CR), which affects almost 50% of patients within 5 years after lung transplantation. CR is a heterogeneous condition and the clinical differential diagnosis remains troublesome. In this project, we hypothesize that innovative histopathologic research will provide important new mechanistic insights in the development of CR, which could lead to a more accurate differential diagnosis. A systematic and detailed histopathologic analysis of the different clinical phenotypes is highly needed, but currently lacking. Therefore, we will investigate the histopathologic changes in routinely acquired lung biopsies and full explant lungs from patients with CR. In addition, we aim to investigate the pathophysiologic mechanisms of different histopathologic lesions and different clinical phenotypes of CR, using cutting-edge techniques such as laser-assisted microdissection and multiplex imaging. This project aims to identify biomarkers to predict and diagnose CR. This is highly needed given the difficult differential diagnosis of CR and poor survival in patients with CR. By elucidating the pathophysiology of CR, this project forms a first step for the later development of highly needed specific drugs to prevent CR development or progression. " "The BACTERIA-INFLAMMATION-REJECTION AXIS IN CHRONIC LUNG ALLOGRAFT DYSFUNCTION" "Geert Verleden" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "Chronic lung allograft dysfunction (CLAD) remains the major long term complication after lung transplantation with a 5- year incidence of 50%, leading to loss of quality of life and increased mortality. Although azithromycin, a neomacrolide antibiotic, increases CLAD-free survival, a large proportion of patients nevertheless develop CLAD (either bronchiolitis obliterans syndrome, BOS or restrictive allograft syndrome, RAS). Within this project, we will investigate, by next-generation sequencing tools, the association between the microbiome, inflammation and CLAD. Using already collected patients samples from our biobank, we will 1/ investigate the change in microbial content in the post-transplant follow-up (with special consideration for CLAD) and the effect of prophylactic azithromycin on the lung allograft microbiome and the association with inflammation. 2/ Assess the microbiome in explant CLAD specimens (either BOS or RAS) and compare this to unused donor lungs with specific attention to the regional variety within the lung. Using immunohistochemistry, we will relate the microbiome to the host response and determine if certain microbes trigger more pronounced or a specific type of immune response. This knowledge will be crucial to obtain novel insights into the pathophysiology of CLAD post lung transplantation." "Translational research on pre-transplant initiation and promt post-transplant continuation of azithromycin treatment to improve early allograft function and outcome after lung transplantation." "Robin Vos" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "The principal cause of late mortality after lung transplantation (LTx) is chronic rejection, which  histologically presents as obliterative bronchiolitis (OB) (1,2) and clinically as a progressive obstructive spirometry, defined as bronchiolitis obliterans syndrome (BOS) (2,3). We recently demonstrated that azithromycin (AZI), when initiated from discharge after LTx on, improved outcome (BOS-free survival), graft function (FEV1) and attenuated both airway and systemic inflammation (4). These data confirmed the anti-inflammatory and imunomodulatory properties of AZI in LTx (5-15). However, early allograft dysfunction, which may be due to donor lung injury or to ischemiareperfusion  injury, was recently also associated with earlier onset of BOS and a worse long-term outcome (16,17). We previously demonstrated that early administration of AZI reduces airway inflammation and oxidative stress in ischemia-reperfusion injury (18); and also reduced pulmonary fibrosis and improved pulmonary function in bleomycin-induced pulmonary fibrosis (19). Macrolides were shown to attenuate acute and chronic rejection in murine cardiac transplantation (20) and in a rat heterotopic tracheal allograft model of OB (21). Finally, macrolide-use is associated with shorter time of mechanical ventilation and a lower 6 month mortality in patients with acute lung injury (22). Given these findings, we believe AZI should be further investigated as a therapy to improve early allograft function and subsequently outcome after LTx." "A central role for the mesothelium in Restrictive Allograft Syndrome after lung transplantation" "Stijn Verleden" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "Chronic lung allograft dysfunction (CLAD) remains the major long term complication after lung transplantation.  Recently, a new clinical phenotype, restrictive allograft syndrome (RAS), is identified which affects 30% of CLAD patients and which results in a survival after diagnosis limited to 0.5-1 year. In this project, we hypothesize an important role for mesothelial cells and mesothelial mesenchymal transition (MMT) in RAS. Our aim is to investigate: 1/ the role and structural organization of mesothelial cells in RAS 2/ the pathophysiological mechanisms of mesenchymal infiltration with a focus on inflammation and immunosuppression and 3/ the effect of current and new therapeutic agents. Our approach is to work with explant lung tissue, mesothelial cell culture and our mice orthotopic lung transplant model. This study aims to better understand the impact and mechanism of pleural mesenchymal cells in RAS and how current and new treatment options can help to improve the long-term survival." "Transcriptome-based management of transplant-related pulmonary fibrosis in chronic pulmonary rejection after lung or allogeneic stem cell transplantation" "Robin Vos" "Academic Centre for Nursing and Midwifery, Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "BackgroundChronic lung allograft dysfunction (CLAD) and chronic pulmonarygraft-versus-host disease (cGvHD) are life-threatening conditionscommonly encountered respectively after lung transplantation andallogeneic hematopoietic stem cell transplantation.Pathophysiological and molecular pathways associated withtransplant-related pulmonary fibrosis, leading to structural changesand lung failure, in CLAD and pulmonary cGvHD remain unknown.AimsThe primary objective of this translational research project is toinvestigate the pathways and mechanisms involved in transplantrelatedpulmonary fibrosis in obstructive and restrictive phenotypes ofCLAD and pulmonary cGvHD, in order to identify novel diagnosticmarkers and to design improved therapeutics, allowing to bettermanage rejection-mediated fibrotic lung remodeling.MethodsA multi-modal approach, using human lung tissue, bronchoalveolarlavage and blood samples, is used to assess the morphometricstructural changes, immunopathologic organization, and genesignature in CLAD and pulmonary cGvHD.Expected resultsThis project will gain novel, breakthrough insights in thepathophysiology of CLAD and pulmonary cGvHD, lead to noveldiagnostic and better therapeutic tools, allowing for improved,personalized medical management; which will result in better longtermoutcomes in patients undergoing lung transplantation andallogeneic hematopoietic stem cell transplantation." "The role of the B-cell mediated immunuunrespons in phenotypes of chronic rejection after lung transplantation." "Stijn Verleden" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "B-cells are important players in the adaptive immune system. Their principal functions are to make antibodies against antigens, to perform the role of antigen-presenting cells (APCs), and to develop into memory B cells after activation by antigen interaction. B-cells have been shown to play an important role in chronic respiraotry diseases like COPD and IPF. With this research, we want to establish a role for B-cells in chronic lung allograft dysfunction (CLAD), with special attention towards the clinical phenotype of CLAD being Bronchiolitis Obliterans Syndrome (BOS) versus Restrictive Allograft Syndrome (RAS). We want to investigate the role of B-cells and antibodies in early and late disease and confirm our findings in a mouse model.To establish a role for B-cells, we will use human BAL, blood and tissue samples wherein we will investigate the presence of B-cells (immunohistochemistry and gene expression analysis on explanted lung specimens) and antibodies (immunoglobulins in BAL and anti-HLA antibodies in serum), but also a mouse model of heterotopic trachea transplantation to consolidate our findings. Lastly, we will use the mousemodel to assess the potential of pirfenidone and extracorporeal photopheresis to attenuate disease. We believe that this research will improve the understanding of the mechanism of CLAD and might be valuable for further therapeutic treatment of CLAD." "Quantification of leukocyte chemoattractants to identify specific drug targets for chronic rejection in lung transplantation" "Paul Proost" "Molecular Immunology (Rega Institute), Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "Neutrophils are the most abundant leukocytes in the human blood circulation and mediate a fast immune response to infection and tissue damage. Directional leukocyte migration throughout the body is meticulously regulated by chemokines, which are small and structurally conserved chemotactic proteins. Notably, chemokine activity is spatiotemporally controlled at multiple levels, including through post-translational modifications such as proteolysis. Resulting chemokine proteoforms can display a significantly altered biological activity compared to the intact proteins. Excessive neutrophil activation may result in collateral tissue damage during uncontrolled inflammation. In patients with severe coronavirus disease 2019 (COVID-19) or chronic lung allograft dysfunction (CLAD) following lung transplantation, neutrophils, proteases and chemokines are associated with disease severity and pathogenesis. Therefore, the objective of this PhD project is to characterize neutrophils and investigate chemokine proteolysis in the blood and broncho-alveolar lavage (BAL) fluids of COVID-19 and CLAD patients. As such, we aim to improve our understanding of neutrophil phenotypical and functional heterogeneity and their contribution to the pathogenesis of these diseases. Moreover, we aim to identify novel drug targets that could be exploited to reduce excessive neutrophil recruitment and inflammation." "Improving Graft Acceptance in Lung Transplantation by Donor-specific Blood Transfusion" "Laurens Ceulemans" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE), Abdominal Transplantation" "Lung transplantation is the ultimate treatment for patients with end-stage pulmonary disease, the survival of which is hindered by chronic lung allograft dysfunction. The classic immunosuppressive protocol, nonspecifically blocking immune reaction, also causes complications such as nephrotoxicity, infection, and cancer. Donor-specific blood transfusion is an efficient strategy to promote graft acceptance under reduced immunosuppression. The safety and efficacy have been validated in pre-clinical and clinical research models of the heart and intestinal transplantation. However, the specific mechanism of how donor-specific blood transfusion takes its protective effect on recipients has not yet been identified. Furthermore, no animal model was developed for donor-specific blood transfusion experiments in lung transplantation. In this project, we aim firstly to establish a new mice model of donor-specific blood transfusion in lung transplantation, and secondly to clarify its effect and mechanism. We hypothesize that the mice model is suitable for donor-specific blood transfusion research in lung transplantation, and the donor-specific blood transfusion will result in a better acceptance of the lung graft in a setting of minimized immunosuppression." "A study on the loss of small airways and epithelial cell changes in chronic lung diseases" "Ghislaine Gayan-Ramirez" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "The main function of the lung is to ensure optimal gas exchange of oxygen uptake and removal of carbon dioxide during breathing. Air entering the nose and mouth is travelling through a well-organized airway tree consisting of branches like in trees. This starts at the trachea (termed generation 0), splitting at the first bifurcation (generation 1) into two branches, each dividing many times along the tree into smaller airways and ending up in terminal bronchioles (generation 5-16). The bifurcations are not symmetrical dichotomous but most often asymmetrical with major and minor bifurcations (1). The diameter and length of the airways are decreasing with increasing generation in a well-balanced and orderly fashion. The fractal geometry system of the airways and the distribution of flow and pressure along this system is highly relevant to understand lung pathologies but are often neglected. There are currently no comprehensive studies which have explored this aspect on (healthy) human lungs or lungs from patients with respiratory diseases as, up to now techniques allowing analyzing human airway organization were lacking. This will be the first issue addressed in this project.The structure of the airways is adapted to their functions and differs between the conducting zone and the respiratory zone. In the conducting zone (up to generation 16), convection occurs while in the respiratory zone (generation 17-23), gas exchange is taking place by diffusion. While large airways have a thick wall of fibrous connective tissue and cartilage rings, which confers them robustness, small airway (diameter" "The roll of regulatory cells in rejection post lung transplant" "Bart Vanaudenaerde" "Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE)" "Chronic lung allograft dysfunction (CLAD) continues to pose a significant challenge to long-term survival after lung transplantation. CLAD is a complex condition caused by a multifactorial process with alloimmune-dependent and -independent factors. Alloimmune-dependent factors are related to the alloimmune response from the recipient directed against the ‘nonself’ donor, referred to as allograft rejection. The objective of this PhD project was to gain more insights into the alloimmune response following lung transplantation leading to chronic graft failure.Firstly, a retrospective study was performed to investigate the role of eosinophils after human lung transplantation. This study compared CLAD-free and graft survival between patients with high and low blood eosinophils using an 8% threshold. Increased blood eosinophils were correlated with a worse graft and CLAD-free survival. Increased blood (≥8%) and increased eosinophils in bronchoalveolar lavage (BAL) (≥2%) had the worse outcome. However, a specific mechanistic function could not be attributed to the eosinophils in CLAD. Investigating rejection in human lung transplant recipients is challenging, given the variability and confounding factors involved.Secondly, we discussed the different aspects of animal models of lung transplantation. The utilization of animal models is of great interest to facilitate mechanistical research due to the controlled setting they provide. Orthotopic left lung transplantation in mice and rats is the preferred model to investigate post-lung transplant complications. This experimental model offers advantages such as the available immunological read-outs, genetic models, favorable cost–benefit balance, and the high throughput possibility.As third, the murine model of orthotopic left lung transplantation was employed to explore the fundamental immunological elements underlying pulmonary rejection. The model was established using a major genetic mismatch between donor and recipient, with daily immunosuppression. The rejection process followed the basic principles of a classic immune response, including innate but mainly adaptive immune cells, peaking at day seven after lung transplantation.Finally, the structural changes associated with pulmonary rejection following murine lung transplantation were investigated. We demonstrated that pulmonary rejection initiates from the vasculature in the early stage, with subsequent lymphoid follicle formation centered around the blood vessels, progressing to perivascular fibrosis. Notably, CD31+ endothelial cells played a critical role in the rejection process, exhibiting activation and decline at day seven post-transplantation in allografts. However, it is important to acknowledge that the current clinical focus is primarily directed towards the airways, through lung function measurement and pathological obliterated bronchiolitis. Promoting vascular repair and/or protecting the endothelial cells could serve as a vital strategy to improve the outcomes following lung transplantation.Overall, this PhD thesis provided valuable insights into the fundamental principles of pulmonary rejection, utilizing a relevant pre-clinical animal model. The findings highlight the presence of an all-encompassing immune response following murine lung transplantation, with a significant involvement of the vascular compartment during the initial stage."