Document text
Principal Investigator: JOHN GREENLAND
Organization: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
Fiscal Year: 2024
Award: $602,118
Funding agency: National Heart Lung and Blood Institute
The major barrier to long term survival following lung transplantation is a progressive loss of lung function,
termed chronic lung allograft dysfunction (CLAD), for which constrictive fibrosis in small airways is a pathologic
hallmark. CLAD affects over half of lung transplant recipients by 4 years post-transplant and negates much of
the quality of life and functional improvements associated with transplantation. Pitt, Toronto, and UCSF Lung
Transplant programs have refined transcriptional analysis of small airway brushings from lung transplant
recipients as a novel technique to understand the gene expression changes at the anatomical site where CLAD
pathology develops. We have published gene expression changes associated with CLAD validated across our
centers. This proposal leverages this innovative approach to understand mechanisms of CLAD pathogenesis.
Our preliminary data show an early upregulation of hypoxia pathways in airway brushings including genes that
recruit and activate cytotoxic T lymphocytes using both airway epithelial cell culture in hypoxic conditions and
pathway analysis of airway brush transcriptomes. This hypoxia signaling may reflect disordered
microvasculature, absent bronchial circulation, and vascular inflammation associated with lung transplant. In
recruited T lymphocytes, we also observed upregulation of tumor necrosis factor superfamily (TNFSF) genes,
which are major drivers of apoptosis in lymphocyte targets. Our data show preferential apoptosis in airway club
cells, the protectors and progenitors of small airways, in association with upregulated TNF-related apoptosis-
inducing ligand (TRAIL) expression. Our single cell investigations in airway brushings and bronchoalveolar
lavage (BAL) fluid show the segregation of these pathways across epithelial and lymphoid cell types. Based on
these data, we hypothesize that airway hypoxia precedes TNFSF gene expression and T cell-mediated airway
club cell apoptosis that drive CLAD pathogenesis. To test this hypothesis, we will generate parallel cohorts
investigating bulk and single cell transcriptomes of CLAD versus controls across three centers, allowing for
rigorous cross-validation of gene expression signatures. We will complement these studies with cell culture-
based methods to determine mechanisms driving these gene expression changes. In Aim 1, we will quantify
hypoxia-related transcripts in airway brush cells with respect to CLAD and determine how hypoxia can promote
lymphocytic inflammation. In Aim 2, we will determine the cellular sources and kinetics of TNFSF co-stimulatory
molecule expression in CLAD using airway brushes and BAL fluid. In Aim 3, we will investigate whether TRAIL
preferentially induces club cell apoptosis. The synergy of three large lung-transplant translational research
programs with world-class cross-institutional biostatistical infrastructures provides a unique opportunity to
address this hypothesis rigorously. The cell-specific gene expression signatures over the time course of CLAD
development that will be generated through this study are critically needed as surrogate biomarkers to support
clinical trials of targeted therapies and to pioneer a novel approach to CLAD diagnosis.
Terms: <(TNF)-α><APO2><APO2 Ligand><Activated Lymphocyte><Address><Affect><Airway Fibrosis><Airway scar><Alloantigen><Anatomic Sites><Anatomic structures><Anatomy><Apo-2 Ligand><Apoptosis><Apoptosis Pathway><Apoptotic><Automobile Driving><Bio-Informatics><Bioinformatics><Biometrics><Biometry><Biostatistics><Bp50><Bronchioalveolar Lavage><Bronchoalveolar Lavage><Bronchoalveolar Lavage Fluid><Bronchopulmonary Lavage><Brush Cell><CD134><CD154><CD30><CD4 Cells><CD4 Positive T Lymphocytes><CD4 T cells><CD4 helper T cell><CD4 lymphocyte><CD4+ T-Lymphocyte><CD4-Positive Lymphocytes><CD40><CD40L><CD40LG><CD8><CD8B><CD8B1><CD8B1 gene><CDW40><Cachectin><Cell Anoxia><Cell Body><Cell Communication and Signaling><Cell Culture System><Cell Culture Techniques><Cell Death><Cell Hypoxia><Cell Signaling><Cell surface><Cell-Mediated Lympholytic Cells><Cells><Cellular Anoxia><Cellular Hypoxia><Chemotactic Cytokines><Chronic><Cicatrix><Circulation><Clinical Trials><Cohort Studies><Complement><Complement Proteins><Concurrent Studies><Culturing, in vitro Vertebrate, Primary><Cytolysis><Cytolytic T-Cell><Cytometry><Cytotoxic T Cell><Cytotoxic T-Lymphocytes><DR4><Data><Death Receptor 4><Detection><Development><Diagnosis><Disease><Disorder><Distal><Dysfunction><Epithelial Cells><Exposure to><Expression Signature><Fibrosis><Fibrotic lesions in lung><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><Functional disorder><Gene Expression><Gene Expression Profile><Gene Transcription><Genes><Genetic Transcription><Heat shock proteins><Homologous Chemotactic Cytokines><Hypoxia><Hypoxia Pathway><Hypoxia and p53 in the Cardiovascular system><Hypoxic><Image><Immune><Immune response><Immunes><Immunity><Immunobiology><Immunological response><Immunophysiology><In Vitro><Inflammation><Infrastructure><Institution><Intercrines><Intracellular Communication and Signaling><Investigation><KI-1><Kinetics><LYT3><Ligands><Light><Link><Lung><Lung Diseases><Lung Grafting><Lung Lavage><Lung Parenchyma><Lung Respiratory System><Lung Tissue><Lung Transplantation><Lung scar><Lung tissue scar><Lymphatic cell><Lymphocyte><Lymphocytic><Lymphoid Cell><Lysis><MGC9013><MGC9365><Macrophage><Macrophage-Derived TNF><Mediating><Methods><Monocyte-Derived TNF><Mφ><Network Analysis><OX40><Oxygen Deficiency><Pathogenesis><Pathologic><Pathology><Pathway Analysis><Pathway interactions><Phenotype><Photoradiation><Physiopathology><Play><Prevention><Primary Cell Cultures><Progenitor Cells><Programmed Cell Death><Proteins><Publishing><Pulmonary Diseases><Pulmonary Disorder><Pulmonary Graft><Pulmonary Scar><Pulmonary Tissue fibrosis><Pulmonary Transplant><Pulmonary Transplantation><QOL><Quality of life><RNA Expression><RNA Seq><RNA sequencing><RNAseq><Reporting><Respiratory Epithelium><Respiratory fibrosis><Role><SIS cytokines><Scarring at the lung><Scarring in the lung><Scars><Signal Transduction><Signal Transduction Systems><Signaling><Source><Stimulus><Stress><Structure of parenchyma of lung><Structure of respiratory epithelium><Surrogate Markers><T cell response><T-Cells><T-Lymphocyte><T-cell inflamed><T4 Cells><T4 Lymphocytes><TNF><TNF A><TNF Alpha><TNF gene><TNF-Related Apoptosis Inducing Ligand Receptor 1><TNF-Related Apoptosis Inducing Ligand TRAIL><TNF-related apoptosis-inducing ligand><TNF-related apoptosis-inducing ligand receptor 1><TNF-α><TNFA><TNFRSF10A><TNFRSF10A gene><TNFRSF5><TNFRSF5 gene><TNFRSF8><TNFRSF8 gene><TNFSF10 Protein><TNFSF5><TNFSF5 gene><TNFα><TRAIL Protein><TRAIL Receptor 1><TRAIL-R4><TRAILR-1><TRAILR1><TRAP Gene><Techniques><Teff cell><Testing><Time><Transcript><Transcription><Translational Research Enterprise><Transplant Recipients><Transplantation><Tumor Necrosis Factor><Tumor Necrosis Factor Ligand Superfamily Member 10><Tumor Necrosis Factor Receptor Superfamily Member 5 Gene><Tumor Necrosis Factor Receptor Superfamily, Member 10A><Tumor Necrosis Factor-alpha><Up-Regulation><Upregulation><Validation><Vascular blood supply><airway epithelium><airway hypoxia><apoptosis in lymphocytes><apoptotic lymphocytes><biological signal transduction><blood supply><bronchopulmonary lavage therapy><cell culture><cell cultures><cell type><chemoattractant cytokine><chemokine><clinical phenotype><clinical subtypes><cohort><complementation><cytokine><cytotoxic><death receptor-4><developmental><disease of the lung><disorder of the lung><driving><effector T cell><fibrotic lung><flow cytophotometry><functional improvement><gene expression pattern><gene expression signature><global gene expression><global transcription profile><host response><imaging><immune system response><immunoresponse><improve function><improved><improved functional outcomes><innovate><innovation><innovative><insight><killer T cell><lung allograft><lung disorder><lung function><lung transplant><lymph cell><lymphocyte apoptosis><member><multidisciplinary><necrocytosis><new approaches><new marker><novel><novel approaches><novel biomarker><novel marker><novel strategies><novel strategy><p50><pathophysiology><pathway><post-transplant><post-transplantation><posttransplant><posttransplantation><prevent><preventing><progenitor><programs><pulmonary><pulmonary function><receptor TRAIL-4><recruit><respiratory hypoxia><respiratory tract epithelium><response><scRNA-seq><segregation><single cell RNA-seq><single cell RNAseq><single cell expression profiling><single cell transcriptomic profiling><single-cell RNA sequencing><social role><stem cells><stress protein><success><surrogate bio-markers><surrogate biomarkers><synergism><targeted drug trials><targeted pharmaceutical trials><targeted therapy trials><targeted treatment trials><thymus derived lymphocyte><tool><transcriptional profile><transcriptional signature><transcriptome><transcriptome sequencing><transcriptomic sequencing><translation research enterprise><translational research program><transplant><transplant centers><transplant patient><tumor necrosis factor receptor superfamily member 10A><validations><vascular inflammation><vascular supply>