Innate Immune Landscape in Early Lung Adenocarcinoma by Paired Single-Cell Analyses.

Little is known about the relative importance of monocyte and tissue-resident macrophages in the development of lung fibrosis. We show that specific genetic deletion of monocyte-derived alveolar macrophages after their recruitment to the lung ameliorated lung fibrosis, whereas tissue-resident alveolar macrophages did not contribute to fibrosis. Using transcriptomic profiling of flow-sorted cells, we found that monocyte to alveolar macrophage differentiation unfolds continuously over the course of fibrosis and its resolution. During the fibrotic phase, monocyte-derived alveolar macrophages differ significantly from tissue-resident alveolar macrophages in their expression of profibrotic genes. A population of monocyte-derived alveolar macrophages persisted in the lung for one year after the resolution of fibrosis, where they became increasingly similar to tissue-resident alveolar macrophages. Human homologues of profibrotic genes expressed by mouse monocyte-derived alveolar macrophages during fibrosis were up-regulated in human alveolar macrophages from fibrotic compared with normal lungs. Our findings suggest that selectively targeting alveolar macrophage differentiation within the lung may ameliorate fibrosis without the adverse consequences associated with global monocyte or tissue-resident alveolar macrophage depletion.

/pdf/monocyte-derived-alveolar-macrophages-drive-lung-fibrosis-3rgqexd4dl.pdf

Monocyte-derived alveolar macrophages drive lung fibrosis and persist in the lung over the life span.

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)

Human monocytes are divided in three major populations; classical (CD14+CD16-), non-classical (CD14dimCD16+), and intermediate (CD14+CD16+). Each of these subsets is distinguished from each other by the expression of distinct surface markers and by their functions in homeostasis and disease. In this review, we discuss the most up-to-date phenotypic classification of human monocytes that has been greatly aided by the application of novel single-cell transcriptomic and mass cytometry technologies. Furthermore, we shed light on the role of these plastic immune cells in already recognized and emerging human chronic diseases, such as obesity, atherosclerosis, chronic obstructive pulmonary disease, lung fibrosis, lung cancer, and Alzheimer's disease. Our aim is to provide an insight into the contribution of human monocytes to the progression of these diseases and highlight their candidacy as potential therapeutic cell targets.

/pdf/human-monocyte-subsets-and-phenotypes-in-major-chronic-250l25x8u3.pdf

Human Monocyte Subsets and Phenotypes in Major Chronic Inflammatory Diseases

Some patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) develop severe pneumonia and acute respiratory distress syndrome1 (ARDS). Distinct clinical features in these patients have led to speculation that the immune response to virus in the SARS-CoV-2-infected alveolus differs from that in other types of pneumonia2. Here we investigate SARS-CoV-2 pathobiology by characterizing the immune response in the alveoli of patients infected with the virus. We collected bronchoalveolar lavage fluid samples from 88 patients with SARS-CoV-2-induced respiratory failure and 211 patients with known or suspected pneumonia from other pathogens, and analysed them using flow cytometry and bulk transcriptomic profiling. We performed single-cell RNA sequencing on 10 bronchoalveolar lavage fluid samples collected from patients with severe coronavirus disease 2019 (COVID-19) within 48 h of intubation. In the majority of patients with SARS-CoV-2 infection, the alveolar space was persistently enriched in T cells and monocytes. Bulk and single-cell transcriptomic profiling suggested that SARS-CoV-2 infects alveolar macrophages, which in turn respond by producing T cell chemoattractants. These T cells produce interferon-γ to induce inflammatory cytokine release from alveolar macrophages and further promote T cell activation. Collectively, our results suggest that SARS-CoV-2 causes a slowly unfolding, spatially limited alveolitis in which alveolar macrophages containing SARS-CoV-2 and T cells form a positive feedback loop that drives persistent alveolar inflammation. Analysis of bronchoalveolar lavage fluid samples from patients with SARS-CoV-2-induced respiratory failure suggests that SARS-CoV-2 infects alveolar macrophages to cause release of T cell chemoattractants, thereby inducing local inflammatory cytokine release and further T cell activation, ultimately resulting in a positive feedback loop that drives alveolar inflammation.

/pdf/circuits-between-infected-macrophages-and-t-cells-in-sars-3dd0qab0tq.pdf

Circuits between infected macrophages and T cells in SARS-CoV-2 pneumonia.

Clear identification of specific cell populations by flow cytometry is important to understand functional roles. A well-defined flow cytometry panel for myeloid cells in human bronchoalveolar lavage (BAL) and lung tissue is currently lacking. The objective of this study was to develop a flow cytometry-based panel for human BAL and lung tissue. We obtained and performed flow cytometry/sorting on human BAL cells and lung tissue. Confocal images were obtained from lung tissue using antibodies for cluster of differentiation (CD)206, CD169, and E cadherin. We defined a multicolor flow panel for human BAL and lung tissue that identifies major leukocyte populations. These include macrophage (CD206(+)) subsets and other CD206(-) leukocytes. The CD206(-) cells include: (1) three monocyte (CD14(+)) subsets, (2) CD11c(+) dendritic cells (CD14(-), CD11c(+), HLA-DR(+)), (3) plasmacytoid dendritic cells (CD14(-), CD11c(-), HLA-DR(+), CD123(+)), and (4) other granulocytes (neutrophils, mast cells, eosinophils, and basophils). Using this panel on human lung tissue, we defined two populations of pulmonary macrophages: CD169(+) and CD169(-) macrophages. In lung tissue, CD169(-) macrophages were a prominent cell type. Using confocal microscopy, CD169(+) macrophages were located in the alveolar space/airway, defining them as alveolar macrophages. In contrast, CD169(-) macrophages were associated with airway/alveolar epithelium, consistent with interstitial-associated macrophages. We defined a flow cytometry panel in human BAL and lung tissue that allows identification of multiple immune cell types and delineates alveolar from interstitial-associated macrophages. This study has important implications for defining myeloid cells in human lung samples.

/pdf/flow-cytometric-analysis-of-myeloid-cells-in-human-blood-42dy1r0epx.pdf

Flow Cytometric Analysis of Myeloid Cells in Human Blood, Bronchoalveolar Lavage, and Lung Tissues.

Rationale: Rigid bronchoscopy is increasingly used by pulmonologists for the management of central airway disorders. However, an assessment tool to evaluate the competency of operators in the performance of this technique has not been developed. We created the Rigid Bronchoscopy Tool for Assessment of Skills and Competence (RIGID-TASC) to serve as an objective, competency-oriented assessment tool of basic rigid bronchoscopic skills, including rigid bronchoscopic intubation and central airway navigation.Objectives: To assess whether RIGID-TASC scores accurately distinguish the basic rigid bronchoscopy skills of novice, intermediate, and expert operators, and to determine whether RIGID-TASC has adequate interrater reliability when used by different independent testers.Methods: At two academic medical centers in the United States, 30 physician volunteers were selected in three categories: 10 novices at rigid bronchoscopy (performed at least 50 flexible, but no rigid, bronchoscopies), 10 operators with interm...

https://www.atsjournals.org/doi/pdf/10.1513/AnnalsATS.201509-593OC

Development of a Tool to Assess Basic Competency in the Performance of Rigid Bronchoscopy.

Epidemiological studies continue to show associations between adverse health effects and outdoor air pollution despite tighter regulation in recent years. These adverse effects occur at levels of pollutants much lower than those encountered in earlier air pollution disasters and, in some studies, at concentrations near or below the national standards. Although the relative risk tends to be low, the population attributable risk is significant due to the large number of people exposed to air pollutants. The adverse health consequences primarily are those related to respiratory and cardiovascular systems. Respiratory effects include decline in lung function, increases in respiratory-related hospital admissions, exacerbation of asthma and COPD, and elevations in the rates of respiratory infection. This chapter reviews the respiratory health effects associated with particulate matter (PM) and gaseous pollutants and briefly discusses clinical approaches to managing air pollution-­associated lung disease, especially in susceptible populations, such as patients with pre-existing cardiopulmonary diseases.

Air Pollution and Lung Diseases

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Flow Cytometric Analysis of Myeloid Cells in Human Blood, Bronchoalveolar Lavage, and Lung Tissues.

Development of a Tool to Assess Basic Competency in the Performance of Rigid Bronchoscopy.

Air Pollution and Lung Diseases