Mortality due to Covid-19 is highly associated with advanced age, owing in large part to severe lower respiratory tract infection. SARS-CoV-2 utilizes the host ACE2 receptor for infection. Whether ...ACE2 abundance in the lung contributes to age-associated vulnerability is currently unknown. We set out to characterize the RNA and protein expression profiles of ACE2 in aging human lung in the context of phenotypic parameters likely to affect lung physiology. Examining publicly available RNA sequencing data, we discovered that mechanical ventilation is a critical variable affecting lung ACE2 levels. Therefore, we investigated ACE2 protein abundance in patients either requiring mechanical ventilation or spontaneously breathing. ACE2 distribution and expression were determined in archival lung samples by immunohistochemistry (IHC). Tissues were selected from the specimen inventory at a large teaching hospital collected between 2010-2020. Twelve samples were chosen from patients receiving mechanical ventilation for acute hypoxic respiratory failure (AHRF). Twenty samples were selected from patients not requiring ventilation. We compared samples across age, ranging from 40-83 years old in the ventilated cohort and 14-80 years old in the non-ventilated cohort. Within the alveolated parenchyma, ACE2 expression is predominantly observed in type II pneumocytes (or alveolar type II / AT2 cells) and alveolar macrophages. All 12 samples from our ventilated cohort showed histologic features of diffuse alveolar damage including reactive, proliferating AT2 cells. In these cases, ACE2 was strongly upregulated with age when normalized to lung area (p = 0.004) or cellularity (p = 0.003), associated with prominent expression in AT2 cells. In non-ventilated individuals, AT2 cell reactive changes were not observed and ACE2 expression did not change with age when normalized to lung area (p = 0.231) or cellularity (p = 0.349). In summary, ACE2 expression increases with age in the setting of alveolar damage observed in patients on mechanical ventilation, providing a potential mechanism for higher Covid-19 mortality in the elderly.
Although single-cell RNA sequencing studies have begun to provide compendia of cell expression profiles
, it has been difficult to systematically identify and localize all molecular cell types in ...individual organs to create a full molecular cell atlas. Here, using droplet- and plate-based single-cell RNA sequencing of approximately 75,000 human cells across all lung tissue compartments and circulating blood, combined with a multi-pronged cell annotation approach, we create an extensive cell atlas of the human lung. We define the gene expression profiles and anatomical locations of 58 cell populations in the human lung, including 41 out of 45 previously known cell types and 14 previously unknown ones. This comprehensive molecular atlas identifies the biochemical functions of lung cells and the transcription factors and markers for making and monitoring them; defines the cell targets of circulating hormones and predicts local signalling interactions and immune cell homing; and identifies cell types that are directly affected by lung disease genes and respiratory viruses. By comparing human and mouse data, we identified 17 molecular cell types that have been gained or lost during lung evolution and others with substantially altered expression profiles, revealing extensive plasticity of cell types and cell-type-specific gene expression during organ evolution including expression switches between cell types. This atlas provides the molecular foundation for investigating how lung cell identities, functions and interactions are achieved in development and tissue engineering and altered in disease and evolution.
Lung cancer is the most prevalent cancer worldwide, and histopathological assessment is indispensable for its diagnosis. However, human evaluation of pathology slides cannot accurately predict ...patients' prognoses. In this study, we obtain 2,186 haematoxylin and eosin stained histopathology whole-slide images of lung adenocarcinoma and squamous cell carcinoma patients from The Cancer Genome Atlas (TCGA), and 294 additional images from Stanford Tissue Microarray (TMA) Database. We extract 9,879 quantitative image features and use regularized machine-learning methods to select the top features and to distinguish shorter-term survivors from longer-term survivors with stage I adenocarcinoma (P<0.003) or squamous cell carcinoma (P=0.023) in the TCGA data set. We validate the survival prediction framework with the TMA cohort (P<0.036 for both tumour types). Our results suggest that automatically derived image features can predict the prognosis of lung cancer patients and thereby contribute to precision oncology. Our methods are extensible to histopathology images of other organs.
Giant cell arteritis (GCA) causes autoimmune inflammation of the aorta and its large branches, resulting in aortic arch syndrome, blindness, and stroke. CD4⁺ T cells and macrophages form organized ...granulomatous lesions in the walls of affected arteries, destroy the tunica media, and induce ischemic organ damage through rapid intimal hyperplasia and luminal occlusion. Pathogenic mechanisms remain insufficiently understood; specifically, it is unknown whether the unopposed activation of the immune system is because of deficiency of immunoinhibitory checkpoints. Transcriptome analysis of GCA-affected temporal arteries revealed low expression of the coinhibitory ligand programmed death ligand-1 (PD-L1) concurrent with enrichment of the programmed death-1 (PD-1) receptor. Tissue-residing and ex vivo-generated dendritic cells (DC) from GCA patients were PD-L1lo, whereas the majority of vasculitic T cells expressed PD-1, suggesting inefficiency of the immunoprotective PD-1/PD-L1 immune checkpoint. DC–PD-L1 expression correlated inversely with clinical disease activity. In human artery-SCID chimeras, PD-1 blockade exacerbated vascular inflammation, enriched for PD-1⁺ effector T cells, and amplified tissue production of multiple T-cell effector cytokines, including IFN-γ, IL-17, and IL-21. Arteries infiltrated by PD-1⁺ effector T cells developed microvascular neoangiogenesis as well as hyperplasia of the intimal layer, implicating T cells in the maladaptive behavior of vessel wall endogenous cells. Thus, in GCA, a breakdown of the tissue-protective PD1/PD-L1 checkpoint unleashes vasculitic immunity and regulates the pathogenic remodeling of the inflamed arterial wall.
After heart transplantation, endomyocardial biopsy (EMBx) is used to monitor for acute rejection (AR). Unfortunately, EMBx is invasive, and its conventional histological interpretation has ...limitations. This is a validation study to assess the performance of a sensitive blood biomarker-percent donor-derived cell-free DNA (%ddcfDNA)-for detection of AR in cardiac transplant recipients.
This multicenter, prospective cohort study recruited heart transplant subjects and collected plasma samples contemporaneously with EMBx for %ddcfDNA measurement by shotgun sequencing. Histopathology data were collected to define AR, its 2 phenotypes (acute cellular rejection ACR and antibody-mediated rejection AMR), and controls without rejection. The primary analysis was to compare %ddcfDNA levels (median and interquartile range IQR) for AR, AMR, and ACR with controls and to determine %ddcfDNA test characteristics using receiver-operator characteristics analysis.
The study included 171 subjects with median posttransplant follow-up of 17.7 months (IQR, 12.1-23.6), with 1392 EMBx, and 1834 %ddcfDNA measures available for analysis. Median %ddcfDNA levels decayed after surgery to 0.13% (IQR, 0.03%-0.21%) by 28 days. Also, %ddcfDNA increased again with AR compared with control values (0.38% IQR, 0.31-0.83%, versus 0.03% IQR, 0.01-0.14%;
<0.001). The rise was detected 0.5 and 3.2 months before histopathologic diagnosis of ACR and AMR. The area under the receiver operator characteristic curve for AR was 0.92. A 0.25%ddcfDNA threshold had a negative predictive value for AR of 99% and would have safely eliminated 81% of EMBx. In addition, %ddcfDNA showed distinctive characteristics comparing AMR with ACR, including 5-fold higher levels (AMR ≥2, 1.68% IQR, 0.49-2.79% versus ACR grade ≥2R, 0.34% IQR, 0.28-0.72%), higher area under the receiver operator characteristic curve (0.95 versus 0.85), higher guanosine-cytosine content, and higher percentage of short ddcfDNA fragments.
We found that %ddcfDNA detected AR with a high area under the receiver operator characteristic curve and negative predictive value. Monitoring with ddcfDNA demonstrated excellent performance characteristics for both ACR and AMR and led to earlier detection than the EMBx-based monitoring. This study supports the use of %ddcfDNA to monitor for AR in patients with heart transplant and paves the way for a clinical utility study. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT02423070.
During the last 25 years, antibody-mediated rejection of the cardiac allograft has evolved from a relatively obscure concept to a recognized clinical complication in the management of heart ...transplant patients. Herein we report the consensus findings from a series of meetings held between 2010-2012 to develop a Working Formulation for the pathologic diagnosis, grading, and reporting of cardiac antibody-mediated rejection. The diagnostic criteria for its morphologic and immunopathologic components are enumerated, illustrated, and described in detail. Numerous challenges and unresolved clinical, immunologic, and pathologic questions remain to which a Working Formulation may facilitate answers.
Antibody-mediated rejection (AMR) is a recognized cause of allograft dysfunction in lung transplant recipients. Unlike AMR in other solid-organ transplant recipients, there are no standardized ...diagnostic criteria or an agreed-upon definition. Hence, a working group was created by the International Society for Heart and Lung Transplantation with the aim of determining criteria for pulmonary AMR and establishing a definition. Diagnostic criteria and a working consensus definition were established. Key diagnostic criteria include the presence of antibodies directed toward donor human leukocyte antigens and characteristic lung histology with or without evidence of complement 4d within the graft. Exclusion of other causes of allograft dysfunction increases confidence in the diagnosis but is not essential. Pulmonary AMR may be clinical (allograft dysfunction which can be asymptomatic) or sub-clinical (normal allograft function). This consensus definition will have clinical, therapeutic and research implications.
Because of their vital function, the wall structures of medium and large arteries are immunoprivileged and protected from inflammatory attack. That vascular immunoprivilege is broken in ...atherosclerosis and in vasculitis, when wall‐invading T cells and macrophages (Mϕ) promote tissue injury and maladaptive repair. Historically, tissue‐residing T cells were studied for their antigen specificity, but recent progress has refocused attention to antigen‐nonspecific regulation, which determines tissue access, persistence, and functional differentiation of T cells. The coinhibitory receptor PD‐1, expressed on T cells, delivers negative signals when engaged by its ligand PD‐L1, expressed on dendritic cells, Mϕ, and endothelial cells to attenuate T cell activation, effector functions, and survival. Through mitigating signals, the PD‐1 immune checkpoint maintains tissue tolerance. In line with this concept, dendritic cells and Mϕs from patients with the vasculitic syndrome giant cell arteritis (GCA) are PD‐L1lo; including vessel‐wall–embedded DCs that guard the vascular immunoprivilege. GCA infiltrates in the arterial walls are filled with PD‐1+ T cells that secrete IFN‐γ, IL‐17, and IL‐21; drive inflammation‐associated angiogenesis; and facilitate intimal hyperplasia. Conversely, chronic tissue inflammation in the atherosclerotic plaque is associated with an overreactive PD‐1 checkpoint. Plaque‐residing Mϕs are PD‐L1hi, a defect induced by their addiction to glucose and glycolytic breakdown. PD‐L1hi Mϕs render patients with coronary artery disease immunocompromised and suppress antiviral immunity, including protective anti–varicella zoster virus T cells. Thus, immunoinhibitory signals affect several domains of vascular inflammation; failing PD‐L1 in vasculitis enables unopposed immunostimulation and opens the flood gates for polyfunctional inflammatory T cells, and excess PD‐L1 in the atherosclerotic plaque disables tissue‐protective T cell immunity.
Like other autoimmune diseases, rheumatoid arthritis (RA) develops in distinct stages, with each phase of disease linked to immune cell dysfunction. HLA class II genes confer the strongest genetic ...risk to develop RA. They encode for molecules essential in the activation and differentiation of T cells, placing T cells upstream in the immunopathology. In Phase 1 of the RA disease process, T cells lose a fundamental function, their ability to be self-tolerant, and provide help for autoantibody-producing B cells. Phase 2 begins many years later, when mis-differentiated T cells gain tissue-invasive effector functions, enter the joint, promote non-resolving inflammation, and give rise to clinically relevant arthritis. In Phase 3 of the RA disease process, abnormal innate immune functions are added to adaptive autoimmunity, converting synovial inflammation into a tissue-destructive process that erodes cartilage and bone. Emerging data have implicated metabolic mis-regulation as a fundamental pathogenic pathway in all phases of RA. Early in their life cycle, RA T cells fail to repair mitochondrial DNA, resulting in a malfunctioning metabolic machinery. Mitochondrial insufficiency is aggravated by the mis-trafficking of the energy sensor AMPK away from the lysosomal surface. The metabolic signature of RA T cells is characterized by the shunting of glucose toward the pentose phosphate pathway and toward biosynthetic activity. During the intermediate and terminal phase of RA-imposed tissue inflammation, tissue-residing macrophages, T cells, B cells and stromal cells are chronically activated and under high metabolic stress, creating a microenvironment poor in oxygen and glucose, but rich in metabolic intermediates, such as lactate. By sensing tissue lactate, synovial T cells lose their mobility and are trapped in the tissue niche. The linkage of defective DNA repair, misbalanced metabolic pathways, autoimmunity, and tissue inflammation in RA encourages metabolic interference as a novel treatment strategy during both the early stages of tolerance breakdown and the late stages of tissue inflammation. Defining and targeting metabolic abnormalities provides a new paradigm to treat, or even prevent, the cellular defects underlying autoimmune disease.
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