ABSTRACT
The Human BioMolecular Atlas Program (HuBMAP) aims to construct a reference 3D structural, cellular, and molecular atlas of the healthy adult human body. The HuBMAP Data Portal (https://portal.hubmapconsortium.org) serves experimental datasets and supports data processing, search, filtering, and visualization. The Human Reference Atlas (HRA) Portal (https://humanatlas.io) provides open access to atlas data, code, procedures, and instructional materials. Experts from more than 20 consortia are collaborating to construct the HRA's Common Coordinate Framework (CCF), knowledge graphs, and tools that describe the multiscale structure of the human body (from organs and tissues down to cells, genes, and biomarkers) and to use the HRA to understand changes that occur at each of these levels with aging, disease, and other perturbations. The 6th release of the HRA v2.0 covers 36 organs with 4,499 unique anatomical structures, 1,195 cell types, and 2,089 biomarkers (e.g., genes, proteins, lipids) linked to ontologies. In addition, three workflows were developed to map new experimental data into the HRA's CCF. This paper describes the HRA user stories, terminology, data formats, ontology validation, unified analysis workflows, user interfaces, instructional materials, application programming interface (APIs), flexible hybrid cloud infrastructure, and demonstrates first atlas usage applications and previews.
ABSTRACT
The use of single cell/nucleus RNA sequencing (scRNA-seq) technologies that quantitively describe cell transcriptional phenotypes is revolutionizing our understanding of cell biology, leading to new insights in cell type identification, disease mechanisms, and drug development. The tremendous growth in scRNA-seq data has posed new challenges in efficiently characterizing data-driven cell types and identifying quantifiable marker genes for cell type classification. The use of machine learning and explainable artificial intelligence has emerged as an effective approach to study large-scale scRNA-seq data. NS-Forest is a random forest machine learning-based algorithm that aims to provide a scalable data-driven solution to identify minimum combinations of necessary and sufficient marker genes that capture cell type identity with maximum classification accuracy. Here, we describe the latest version, NS-Forest version 4.0 and its companion Python package (https://github.com/JCVenterInstitute/NSForest), with several enhancements to select marker gene combinations that exhibit highly selective expression patterns among closely related cell types and more efficiently perform marker gene selection for large-scale scRNA-seq data atlases with millions of cells. By modularizing the final decision tree step, NS-Forest v4.0 can be used to compare the performance of user-defined marker genes with the NS-Forest computationally-derived marker genes based on the decision tree classifiers. To quantify how well the identified markers exhibit the desired pattern of being exclusively expressed at high levels within their target cell types, we introduce the On-Target Fraction metric that ranges from 0 to 1, with a metric of 1 assigned to markers that are only expressed within their target cell types and not in cells of any other cell types. NS-Forest v4.0 outperforms previous versions on its ability to identify markers with higher On-Target Fraction values for closely related cell types and outperforms other marker gene selection approaches at classification with significantly higher F-beta scores when applied to datasets from three human organs - brain, kidney, and lung.
ABSTRACT
The immunological defects causing susceptibility to severe viral respiratory infections due to early-life dysbiosis remain ill-defined. Here, we show that influenza virus susceptibility in dysbiotic infant mice is caused by CD8+ T cell hyporesponsiveness and diminished persistence as tissue-resident memory cells. We describe a previously unknown role for nuclear factor interleukin 3 (NFIL3) in repression of memory differentiation of CD8+ T cells in dysbiotic mice involving epigenetic regulation of T cell factor 1 (TCF 1) expression. Pulmonary CD8+ T cells from dysbiotic human infants share these transcriptional signatures and functional phenotypes. Mechanistically, intestinal inosine was reduced in dysbiotic human infants and newborn mice, and inosine replacement reversed epigenetic dysregulation of Tcf7 and increased memory differentiation and responsiveness of pulmonary CD8+ T cells. Our data unveils new developmental layers controlling immune cell activation and identifies microbial metabolites that may be used therapeutically in the future to protect at-risk newborns.
ABSTRACT
Postnatal lung development results in an increasingly functional organ prepared for gas exchange and pathogenic challenges. It is achieved through cellular differentiation and migration. Changes in the tissue architecture during this development process are well-documented and increasing cellular diversity associated with it are reported in recent years. Despite recent progress, transcriptomic and molecular pathways associated with human postnatal lung development are yet to be fully understood. In this study, we investigated gene expression patterns associated with healthy pediatric lung development in four major enriched cell populations (epithelial, endothelial, and nonendothelial mesenchymal cells, along with lung leukocytes) from 1-day-old to 8-yr-old organ donors with no known lung disease. For analysis, we considered the donors in four age groups [less than 30 days old neonates, 30 days to < 1 yr old infants, toddlers (1 to < 2 yr), and children 2 yr and older] and assessed differentially expressed genes (DEG). We found increasing age-associated transcriptional changes in all four major cell types in pediatric lung. Transition from neonate to infant stage showed highest number of DEG compared with the number of DEG found during infant to toddler- or toddler to older children-transitions. Profiles of differential gene expression and further pathway enrichment analyses indicate functional epithelial cell maturation and increased capability of antigen presentation and chemokine-mediated communication. Our study provides a comprehensive reference of gene expression patterns during healthy pediatric lung development that will be useful in identifying and understanding aberrant gene expression patterns associated with early life respiratory diseases.NEW & NOTEWORTHY This study presents postnatal transcriptomic changes in major cell populations in human lung, namely endothelial, epithelial, mesenchymal cells, and leukocytes. Although human postnatal lung development continues through early adulthood, our results demonstrate that greatest transcriptional changes occur in first few months of life during neonate to infant transition. These early transcriptional changes in lung parenchyma are particularly notable for functional maturation and activation of alveolar type II cell genes.
Subject(s)
Lung , Transcriptome , Humans , Lung/growth & development , Lung/metabolism , Infant, Newborn , Infant , Child , Child, Preschool , Male , Female , Sequence Analysis, RNA/methods , Epithelial Cells/metabolism , Gene Expression Regulation, Developmental , Gene Expression ProfilingABSTRACT
While animal model studies have extensively defined the mechanisms controlling cell diversity in the developing mammalian lung, there exists a significant knowledge gap with regards to late-stage human lung development. The NHLBI Molecular Atlas of Lung Development Program (LungMAP) seeks to fill this gap by creating a structural, cellular and molecular atlas of the human and mouse lung. Transcriptomic profiling at the single-cell level created a cellular atlas of newborn human lungs. Frozen single-cell isolates obtained from two newborn human lungs from the LungMAP Human Tissue Core Biorepository, were captured, and library preparation was completed on the Chromium 10X system. Data was analyzed in Seurat, and cellular annotation was performed using the ToppGene functional analysis tool. Transcriptional interrogation of 5500 newborn human lung cells identified distinct clusters representing multiple populations of epithelial, endothelial, fibroblasts, pericytes, smooth muscle, immune cells and their gene signatures. Computational integration of data from newborn human cells and with 32,000 cells from postnatal days 1 through 10 mouse lungs generated by the LungMAP Cincinnati Research Center facilitated the identification of distinct cellular lineages among all the major cell types. Integration of the newborn human and mouse cellular transcriptomes also demonstrated cell type-specific differences in maturation states of newborn human lung cells. Specifically, newborn human lung matrix fibroblasts could be separated into those representative of younger cells (n = 393), or older cells (n = 158). Cells with each molecular profile were spatially resolved within newborn human lung tissue. This is the first comprehensive molecular map of the cellular landscape of neonatal human lung, including biomarkers for cells at distinct states of maturity.
Subject(s)
Gene Expression Profiling , Lung , Animals , Humans , Mice , Lung/metabolism , Mammals/genetics , Pericytes , Phenotype , Transcriptome/genetics , Infant, NewbornABSTRACT
Bronchiolitis obliterans (BO) is a fibrotic lung disease characterized by progressive luminal narrowing and obliteration of the small airways. In the nontransplant population, inhalation exposure to certain chemicals is associated with BO; however, the mechanisms contributing to disease induction remain poorly understood. This study's objective was to use single-cell RNA sequencing for the identification of transcriptomic signatures common to primary human airway epithelial cells after chemical exposure to BO-associated chemicals-diacetyl or nitrogen mustard-to help explain BO induction. Primary airway epithelial cells were cultured at air-liquid interface and exposed to diacetyl, nitrogen mustard, or control vapors. Cultures were dissociated and sequenced for single-cell RNA. Differential gene expression and functional pathway analyses were compared across exposures. In total, 75,663 single cells were captured and sequenced from all exposure conditions. Unbiased clustering identified 11 discrete phenotypes, including 5 basal, 2 ciliated, and 2 secretory cell clusters. With chemical exposure, the proportion of cells assigned to keratin 5+ basal cells decreased, whereas the proportion of cells aligned to secretory cell clusters increased compared with control exposures. Functional pathway analysis identified interferon signaling and antigen processing/presentation as pathways commonly upregulated after diacetyl or nitrogen mustard exposure in a ciliated cell cluster. Conversely, the response of airway basal cells differed significantly with upregulation of the unfolded protein response in diacetyl-exposed basal cells, not seen in nitrogen mustard-exposed cultures. These new insights provide early identification of airway epithelial signatures common to BO-associated chemical exposures.NEW & NOTEWORTHY Bronchiolitis obliterans (BO) is a devastating fibrotic lung disease of the small airways, or bronchioles. This original manuscript uses single-cell RNA sequencing for identifying common signatures of chemically exposed airway epithelial cells in BO induction. Chemical exposure reduced the proportion of keratin 5+ basal cells while increasing the proportion of keratin 4+ suprabasal cells. Functional pathways contributory to these shifts differed significantly across exposures. These new results highlight similarities and differences in BO induction across exposures.
Subject(s)
Bronchiolitis Obliterans , Diacetyl , Humans , Keratin-5/metabolism , Diacetyl/metabolism , Mechlorethamine/metabolism , Respiratory Mucosa/metabolism , Bronchiolitis Obliterans/chemically induced , Bronchiolitis Obliterans/metabolism , Epithelial Cells/metabolismABSTRACT
Preterm infants with oxygen supplementation are at high risk for bronchopulmonary dysplasia (BPD), a neonatal chronic lung disease. Inflammation with macrophage activation is central to the pathogenesis of BPD. CXCL10, a chemotactic and pro-inflammatory chemokine, is elevated in the lungs of infants evolving BPD and in hyperoxia-based BPD in mice. Here, we tested if CXCL10 deficiency preserves lung growth after neonatal hyperoxia by preventing macrophage activation. To this end, we exposed Cxcl10 knockout (Cxcl10-/-) and wild-type mice to an experimental model of hyperoxia (85% O2)-induced neonatal lung injury and subsequent regeneration. In addition, cultured primary human macrophages and murine macrophages (J744A.1) were treated with CXCL10 and/or CXCR3 antagonist. Our transcriptomic analysis identified CXCL10 as a central hub in the inflammatory network of neonatal mouse lungs after hyperoxia. Quantitative histomorphometric analysis revealed that Cxcl10-/- mice are in part protected from reduced alveolar. These findings were related to the preserved spatial distribution of elastic fibers, reduced collagen deposition, and protection from macrophage recruitment/infiltration to the lungs in Cxcl10-/- mice during acute injury and regeneration. Complimentary, studies with cultured human and murine macrophages showed that hyperoxia induces Cxcl10 expression that in turn triggers M1-like activation and migration of macrophages through CXCR3. Finally, we demonstrated a temporal increase of macrophage-related CXCL10 in the lungs of infants with BPD. In conclusion, our data demonstrate macrophage-derived CXCL10 in experimental and clinical BPD that drives macrophage chemotaxis through CXCR3, causing pro-fibrotic lung remodeling and arrest of alveolarization. Thus, targeting the CXCL10-CXCR3 axis could offer a new therapeutic avenue for BPD.
ABSTRACT
Multiplexed antibody-based imaging enables the detailed characterization of molecular and cellular organization in tissues. Advances in the field now allow high-parameter data collection (>60 targets); however, considerable expertise and capital are needed to construct the antibody panels employed by these methods. Organ mapping antibody panels are community-validated resources that save time and money, increase reproducibility, accelerate discovery and support the construction of a Human Reference Atlas.
Subject(s)
Antibodies , Community Resources , Humans , Reproducibility of Results , Diagnostic ImagingABSTRACT
Bronchopulmonary dysplasia (BPD) is a disease of prematurity related to the arrest of normal lung development. The objective of this study was to better understand how proteome modulation and cell-type shifts are noted in BPD pathology. Pediatric human donors aged 1-3 yr were classified based on history of prematurity and histopathology consistent with "healed" BPD (hBPD, n = 3) and "established" BPD (eBPD, n = 3) compared with respective full-term born (n = 6) age-matched term controls. Proteins were quantified by tandem mass spectroscopy with selected Western blot validations. Multiplexed immunofluorescence (MxIF) microscopy was performed on lung sections to enumerate cell types. Protein abundances and MxIF cell frequencies were compared among groups using ANOVA. Cell type and ontology enrichment were performed using an in-house tool and/or EnrichR. Proteomics detected 5,746 unique proteins, 186 upregulated and 534 downregulated, in eBPD versus control with fewer proteins differentially abundant in hBPD as compared with age-matched term controls. Cell-type enrichment suggested a loss of alveolar type I, alveolar type II, endothelial/capillary, and lymphatics, and an increase in smooth muscle and fibroblasts consistent with MxIF. Histochemistry and Western analysis also supported predictions of upregulated ferroptosis in eBPD versus control. Finally, several extracellular matrix components mapping to angiogenesis signaling pathways were altered in eBPD. Despite clear parsing by protein abundance, comparative MxIF analysis confirms phenotypic variability in BPD. This work provides the first demonstration of tandem mass spectrometry and multiplexed molecular analysis of human lung tissue for critical elucidation of BPD trajectory-defining factors into early childhood.NEW & NOTEWORTHY We provide new insights into the natural history of bronchopulmonary dysplasia in donor human lungs after the neonatal intensive care unit hospitalization. This study provides new insights into how the proteome and histopathology of BPD changes in early childhood, uncovering novel pathways for future study.
Subject(s)
Bronchopulmonary Dysplasia , Child, Preschool , Infant, Newborn , Humans , Child , Bronchopulmonary Dysplasia/pathology , Immunohistochemistry , Proteome , Proteomics , Lung/metabolismABSTRACT
Accurate cell type identification is a key and rate-limiting step in single-cell data analysis. Single-cell references with comprehensive cell types, reproducible and functionally validated cell identities, and common nomenclatures are much needed by the research community for automated cell type annotation, data integration, and data sharing. Here, we develop a computational pipeline utilizing the LungMAP CellCards as a dictionary to consolidate single-cell transcriptomic datasets of 104 human lungs and 17 mouse lung samples to construct LungMAP single-cell reference (CellRef) for both normal human and mouse lungs. CellRefs define 48 human and 40 mouse lung cell types catalogued from diverse anatomic locations and developmental time points. We demonstrate the accuracy and stability of LungMAP CellRefs and their utility for automated cell type annotation of both normal and diseased lungs using multiple independent methods and testing data. We develop user-friendly web interfaces for easy access and maximal utilization of the LungMAP CellRefs.
Subject(s)
Gene Expression Profiling , Information Dissemination , Animals , Mice , Humans , Single-Cell Analysis , TranscriptomeABSTRACT
At birth, the lung is still immature, heightening susceptibility to injury but enhancing regenerative capacity. Angiogenesis drives postnatal lung development. Therefore, we profiled the transcriptional ontogeny and sensitivity to injury of pulmonary endothelial cells (EC) during early postnatal life. Although subtype speciation was evident at birth, immature lung EC exhibited transcriptomes distinct from mature counterparts, which progressed dynamically over time. Gradual, temporal changes in aerocyte capillary EC (CAP2) contrasted with more marked alterations in general capillary EC (CAP1) phenotype, including distinct CAP1 present only in the early alveolar lung expressing Peg3, a paternally imprinted transcription factor. Hyperoxia, an injury that impairs angiogenesis induced both common and unique endothelial gene signatures, dysregulated capillary EC crosstalk, and suppressed CAP1 proliferation while stimulating venous EC proliferation. These data highlight the diversity, transcriptomic evolution, and pleiotropic responses to injury of immature lung EC, possessing broad implications for lung development and injury across the lifespan.
ABSTRACT
Coronavirus disease (COVID-19) is an infectious disease caused by the SARS coronavirus 2 (SARS-CoV-2) virus. Direct assessment, detection, and quantitative analysis using high throughput methods like single-cell RNA sequencing (scRNAseq) is imperative to understanding the host response to SARS-CoV-2. One barrier to studying SARS-CoV-2 in the laboratory setting is the requirement to process virus-infected cell cultures, and potentially infectious materials derived therefrom, under Biosafety Level 3 (BSL-3) containment. However, there are only 190 BSL3 laboratory facilities registered with the U.S. Federal Select Agent Program, as of 2020, and only a subset of these are outfitted with the equipment needed to perform high-throughput molecular assays. Here, we describe a method for preparing non-hazardous RNA samples from SARS-CoV-2 infected cells, that enables scRNAseq analyses to be conducted safely in a BSL2 facility-thereby making molecular assays of SARS-CoV-2 cells accessible to a much larger community of researchers. Briefly, we infected African green monkey kidney epithelial cells (Vero-E6) with SARS-CoV-2 for 96 hours, trypsin-dissociated the cells, and inactivated them with methanol-acetone in a single-cell suspension. Fixed cells were tested for the presence of infectious SARS-CoV-2 virions using the Tissue Culture Infectious Dose Assay (TCID50), and also tested for viability using flow cytometry. We then tested the dissociation and methanol-acetone inactivation method on primary human lung epithelial cells that had been differentiated on an air-liquid interface. Finally, we performed scRNAseq quality control analysis on the resulting cell populations to evaluate the effects of our virus inactivation and sample preparation protocol on the quality of the cDNA produced. We found that methanol-acetone inactivated SARS-CoV-2, fixed the lung epithelial cells, and could be used to obtain noninfectious, high-quality cDNA libraries. This methodology makes investigating SARS-CoV-2, and related high-containment RNA viruses at a single-cell level more accessible to an expanded community of researchers.
Subject(s)
COVID-19 , SARS-CoV-2 , Animals , Humans , Chlorocebus aethiops , Methanol , Acetone , Single-Cell Gene Expression Analysis , Epithelial CellsABSTRACT
Human disease states are biomolecularly multifaceted and can span across phenotypic states, therefore it is important to understand diseases on all levels, across cell types, and within and across microanatomical tissue compartments. To obtain an accurate and representative view of the molecular landscape within human lungs, this fragile tissue must be inflated and embedded to maintain spatial fidelity of the location of molecules and minimize molecular degradation for molecular imaging experiments. Here, we evaluated agarose inflation and carboxymethyl cellulose embedding media and determined effective tissue preparation protocols for performing bulk and spatial mass spectrometry-based omics measurements. Mass spectrometry imaging methods were optimized to boost the number of annotatable molecules in agarose inflated lung samples. This optimized protocol permitted the observation of unique lipid distributions within several airway regions in the lung tissue block. Laser capture microdissection of these airway regions followed by high-resolution proteomic analysis allowed us to begin linking the lipidome with the proteome in a spatially resolved manner, where we observed proteins with high abundance specifically localized to the airway regions. We also compared our mass spectrometry results to lung tissue samples preserved using two other inflation/embedding media, but we identified several pitfalls with the sample preparation steps using this preservation method. Overall, we demonstrated the versatility of the inflation method, and we can start to reveal how the metabolome, lipidome, and proteome are connected spatially in human lungs and across disease states through a variety of different experiments.
ABSTRACT
An improved understanding of the human lung necessitates advanced systems models informed by an ever-increasing repertoire of molecular omics, cellular, imaging, and pathological datasets. To centralize and standardize information across broad lung research efforts we expanded the LungMAP.net website into a new gateway portal. This portal connects a broad spectrum of research networks, bulk and single-cell multi-omics data and a diverse collection of image data that span mammalian lung development, and disease. The data are standardized across species and technologies using harmonized data and metadata models that leverage recent advances including those from the Human Cell Atlas, diverse ontologies, and the LungMAP CellCards initiative. To cultivate future discoveries, we have aggregated a diverse collection of single-cell atlases for multiple species (human, rhesus, mouse), to enable consistent queries across technologies, cohorts, age, disease, and drug treatment. These atlases are provided as independent and integrated queryable datasets, with an emphasis on dynamic visualization, figure generation, re-analysis, cell-type curation, and automated reference-based classification of user-provided single-cell genomics datasets (Azimuth). As this resource grows, we intend to increase the breadth of available interactive interfaces, supported data types, data portals and datasets from LungMAP and external research efforts.
ABSTRACT
Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease with 30% mortality from heart failure (HF) in the first year of life, but the cause of early HF remains unknown. Induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CM) from patients with HLHS showed that early HF is associated with increased apoptosis, mitochondrial respiration defects, and redox stress from abnormal mitochondrial permeability transition pore (mPTP) opening and failed antioxidant response. In contrast, iPSC-CM from patients without early HF showed normal respiration with elevated antioxidant response. Single-cell transcriptomics confirmed that early HF is associated with mitochondrial dysfunction accompanied with endoplasmic reticulum (ER) stress. These findings indicate that uncompensated oxidative stress underlies early HF in HLHS. Importantly, mitochondrial respiration defects, oxidative stress, and apoptosis were rescued by treatment with sildenafil to inhibit mPTP opening or TUDCA to suppress ER stress. Together these findings point to the potential use of patient iPSC-CM for modeling clinical heart failure and the development of therapeutics.
Subject(s)
Heart Defects, Congenital , Heart Failure , Induced Pluripotent Stem Cells , Antioxidants/metabolism , Heart Defects, Congenital/metabolism , Heart Failure/metabolism , Humans , Mitochondrial Permeability Transition Pore , Myocytes, Cardiac/metabolism , Oxidative StressABSTRACT
Neonatal immune-microbiota co-development is poorly understood, yet age-appropriate recognition of - and response to - pathogens and commensal microbiota is critical to health. In this longitudinal study of 148 preterm and 119 full-term infants from birth through one year of age, we found that postmenstrual age or weeks from conception is a central factor influencing T cell and mucosal microbiota development. Numerous features of the T cell and microbiota functional development remain unexplained; however, by either age metric and are instead shaped by discrete perinatal and postnatal events. Most strikingly, we establish that prenatal antibiotics or infection disrupt the normal T cell population developmental trajectory, influencing subsequent respiratory microbial colonization and predicting respiratory morbidity. In this way, early exposures predict the postnatal immune-microbiota axis trajectory, placing infants at later risk for respiratory morbidity in early childhood.
ABSTRACT
Although increased neuropeptides are often detected in lungs that exhibit respiratory distress, whether they contribute to the condition is unknown. Here, we show in a mouse model of neuroendocrine cell hyperplasia of infancy, a pediatric disease with increased pulmonary neuroendocrine cells (PNECs), excess PNEC-derived neuropeptides are responsible for pulmonary manifestations including hypoxemia. In mouse postnatal lung, prolonged signaling from elevated neuropeptides such as calcitonin gene-related peptide (CGRP) activate receptors enriched on endothelial cells, leading to reduced cellular junction gene expression, increased endothelium permeability, excess lung fluid, and hypoxemia. Excess fluid and hypoxemia were effectively attenuated by either prevention of PNEC formation, inactivation of CGRP gene, endothelium-specific inactivation of CGRP receptor gene, or treatment with CGRP receptor antagonist. Neuropeptides were increased in human lung diseases with excess fluid such as acute respiratory distress syndrome. Our findings suggest that restricting neuropeptide function may limit fluid and improve gas exchange in these conditions.
Subject(s)
Calcitonin Gene-Related Peptide , Neuropeptides , Animals , Calcitonin Gene-Related Peptide/metabolism , Endothelial Cells/metabolism , Humans , Hypoxia/metabolism , Lung/metabolism , Mice , Neuropeptides/metabolismABSTRACT
Oxygen supplementation in preterm infants disrupts alveolar epithelial type 2 (AT2) cell proliferation through poorly understood mechanisms. Here, newborn mice are used to understand how hyperoxia stimulates an early aberrant wave of AT2 cell proliferation that occurs between Postnatal Days (PNDs) 0 and 4. RNA-sequencing analysis of AT2 cells isolated from PND4 mice revealed hyperoxia stimulates expression of mitochondrial-specific methylenetetrahydrofolate dehydrogenase 2 and other genes involved in mitochondrial one-carbon coupled folate metabolism and serine synthesis. The same genes are induced when AT2 cells normally proliferate on PND7 and when they proliferate in response to the mitogen fibroblast growth factor 7. However, hyperoxia selectively stimulated their expression via the stress-responsive activating transcription factor 4 (ATF4). Administration of the mitochondrial superoxide scavenger mitoTEMPO during hyperoxia suppressed ATF4 and thus early AT2 cell proliferation, but it had no effect on normative AT2 cell proliferation seen on PND7. Because ATF4 and methylenetetrahydrofolate dehydrogenase are detected in hyperplastic AT2 cells of preterm infant humans and baboons with bronchopulmonary dysplasia, dampening mitochondrial oxidative stress and ATF4 activation may provide new opportunities for controlling excess AT2 cell proliferation in neonatal lung disease.
Subject(s)
Activating Transcription Factor 4/metabolism , Hyperoxia , Activating Transcription Factor 4/genetics , Animals , Animals, Newborn , Cell Proliferation , Folic Acid/pharmacology , Hyperoxia/metabolism , Infant, Premature , MiceABSTRACT
The human lung plays vital roles in respiration, host defense, and basic physiology. Recent technological advancements such as single-cell RNA sequencing and genetic lineage tracing have revealed novel cell types and enriched functional properties of existing cell types in lung. The time has come to take a new census. Initiated by members of the NHLBI-funded LungMAP Consortium and aided by experts in the lung biology community, we synthesized current data into a comprehensive and practical cellular census of the lung. Identities of cell types in the normal lung are captured in individual cell cards with delineation of function, markers, developmental lineages, heterogeneity, regenerative potential, disease links, and key experimental tools. This publication will serve as the starting point of a live, up-to-date guide for lung research at https://www.lungmap.net/cell-cards/. We hope that Lung CellCards will promote the community-wide effort to establish, maintain, and restore respiratory health.
Subject(s)
Lung/cytology , Lung/physiology , Cell Differentiation/genetics , Databases as Topic , Humans , Lung/metabolism , Regeneration/genetics , Single-Cell Analysis/methodsABSTRACT
Rationale: The current understanding of human lung development derives mostly from animal studies. Although transcript-level studies have analyzed human donor tissue to identify genes expressed during normal human lung development, protein-level analysis that would enable the generation of new hypotheses on the processes involved in pulmonary development are lacking. Objectives: To define the temporal dynamic of protein expression during human lung development. Methods: We performed proteomics analysis of human lungs at 10 distinct times from birth to 8 years to identify the molecular networks mediating postnatal lung maturation. Measurements and Main Results: We identified 8,938 proteins providing a comprehensive view of the developing human lung proteome. The analysis of the data supports the existence of distinct molecular substages of alveolar development and predicted the age of independent human lung samples, and extensive remodeling of the lung proteome occurred during postnatal development. Evidence of post-transcriptional control was identified in early postnatal development. An extensive extracellular matrix remodeling was supported by changes in the proteome during alveologenesis. The concept of maturation of the immune system as an inherent part of normal lung development was substantiated by flow cytometry and transcriptomics. Conclusions: This study provides the first in-depth characterization of the human lung proteome during development, providing a unique proteomic resource freely accessible at Lungmap.net. The data support the extensive remodeling of the lung proteome during development, the existence of molecular substages of alveologenesis, and evidence of post-transcriptional control in early postnatal development.