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Crucial transitions in cancer-including tumor initiation, local expansion, metastasis, and therapeutic resistance-involve complex interactions between cells within the dynamic tumor ecosystem. Transformative single-cell genomics technologies and spatial multiplex in situ methods now provide an opportunity to interrogate this complexity at unprecedented resolution. The Human Tumor Atlas Network (HTAN), part of the National Cancer Institute (NCI) Cancer Moonshot Initiative, will establish a clinical, experimental, computational, and organizational framework to generate informative and accessible three-dimensional atlases of cancer transitions for a diverse set of tumor types. This effort complements both ongoing efforts to map healthy organs and previous large-scale cancer genomics approaches focused on bulk sequencing at a single point in time. Generating single-cell, multiparametric, longitudinal atlases and integrating them with clinical outcomes should help identify novel predictive biomarkers and features as well as therapeutically relevant cell types, cell states, and cellular interactions across transitions. The resulting tumor atlases should have a profound impact on our understanding of cancer biology and have the potential to improve cancer detection, prevention, and therapeutic discovery for better precision-medicine treatments of cancer patients and those at risk for cancer.
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Transformación Celular Neoplásica/metabolismo , Neoplasias/metabolismo , Microambiente Tumoral/fisiología , Atlas como Asunto , Transformación Celular Neoplásica/patología , Genómica/métodos , Humanos , Medicina de Precisión/métodos , Análisis de la Célula Individual/métodosRESUMEN
Lung cancer is closely associated with chronic inflammation, but the causes of inflammation and the specific immune mediators have not been fully elucidated. The lung is a mucosal tissue colonized by a diverse bacterial community, and pulmonary infections commonly present in lung cancer patients are linked to clinical outcomes. Here, we provide evidence that local microbiota provoke inflammation associated with lung adenocarcinoma by activating lung-resident γδ T cells. Germ-free or antibiotic-treated mice were significantly protected from lung cancer development induced by Kras mutation and p53 loss. Mechanistically, commensal bacteria stimulated Myd88-dependent IL-1ß and IL-23 production from myeloid cells, inducing proliferation and activation of Vγ6+Vδ1+ γδ T cells that produced IL-17 and other effector molecules to promote inflammation and tumor cell proliferation. Our findings clearly link local microbiota-immune crosstalk to lung tumor development and thereby define key cellular and molecular mediators that may serve as effective targets in lung cancer intervention.
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Interacciones Microbiota-Huesped/inmunología , Linfocitos Intraepiteliales/inmunología , Neoplasias Pulmonares/inmunología , Animales , Proliferación Celular , Femenino , Interleucina-17/inmunología , Interleucina-1beta/metabolismo , Interleucina-23/metabolismo , Linfocitos Intraepiteliales/metabolismo , Linfocitos Intraepiteliales/fisiología , Pulmón/inmunología , Neoplasias Pulmonares/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microbiota/inmunología , Factor 88 de Diferenciación Mieloide/metabolismo , Neutrófilos/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta , Simbiosis/inmunología , Linfocitos T/inmunologíaRESUMEN
Understanding the dynamic pathogenesis and treatment response in pulmonary diseases requires probing the lung at cellular resolution in real time. Despite advances in intravital imaging, optical imaging of the lung during active respiration and circulation has remained challenging. Here, we introduce the crystal ribcage: a transparent ribcage that allows multiscale optical imaging of the functioning lung from whole-organ to single-cell level. It enables the modulation of lung biophysics and immunity through intravascular, intrapulmonary, intraparenchymal and optogenetic interventions, and it preserves the three-dimensional architecture, air-liquid interface, cellular diversity and respiratory-circulatory functions of the lung. Utilizing these capabilities on murine models of pulmonary pathologies we probed remodeling of respiratory-circulatory functions at the single-alveolus and capillary levels during disease progression. The crystal ribcage and its broad applications presented here will facilitate further studies of nearly any pulmonary disease as well as lead to the identification of new targets for treatment strategies.
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Pulmón , Caja Torácica , Ratones , Animales , Microscopía IntravitalRESUMEN
Bronchial premalignant lesions (PMLs) precede the development of invasive lung squamous cell carcinoma (LUSC), posing a significant challenge in distinguishing those likely to advance to LUSC from those that might regress without intervention. This study followed a novel computational approach, the Graph Perceiver Network, leveraging hematoxylin and eosin-stained whole slide images to stratify endobronchial biopsies of PMLs across a spectrum from normal to tumor lung tissues. The Graph Perceiver Network outperformed existing frameworks in classification accuracy predicting LUSC, lung adenocarcinoma, and nontumor lung tissue on The Cancer Genome Atlas and Clinical Proteomic Tumor Analysis Consortium datasets containing lung resection tissues while efficiently generating pathologist-aligned, class-specific heatmaps. The network was further tested using endobronchial biopsies from two data cohorts, containing normal to carcinoma in situ histology. It demonstrated a unique capability to differentiate carcinoma in situ lung squamous PMLs based on their progression status to invasive carcinoma. The network may have utility in stratifying PMLs for chemoprevention trials or more aggressive follow-up.
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Carcinoma de Células Escamosas , Neoplasias Pulmonares , Lesiones Precancerosas , Humanos , Lesiones Precancerosas/patología , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/genética , Carcinoma de Células Escamosas/patologíaRESUMEN
Molecular events that drive the development of precancerous lesions in the bronchial epithelium, which are precursors of lung squamous cell carcinoma (LUSC), are poorly understood. We demonstrate that disruption of epithelial cellular polarity, via the conditional deletion of the apical determinant Crumbs3 (Crb3), initiates and sustains precancerous airway pathology. The loss of Crb3 in adult luminal airway epithelium promotes the uncontrolled activation of the transcriptional regulators YAP and TAZ, which stimulate intrinsic signals that promote epithelial cell plasticity and paracrine signals that induce basal-like cell growth. We show that aberrant polarity and YAP/TAZ-regulated gene expression associates with human bronchial precancer pathology and disease progression. Analyses of YAP/TAZ-regulated genes further identified the ERBB receptor ligand Neuregulin-1 (NRG1) as a key transcriptional target and therapeutic targeting of ERBB receptors as a means of preventing and treating precancerous cell growth. Our observations offer important molecular insight into the etiology of LUSC and provides directions for potential interception strategies of lung cancer.
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Carcinoma de Células Escamosas/genética , Neoplasias Pulmonares/genética , Glicoproteínas de Membrana/genética , Neurregulina-1/genética , Lesiones Precancerosas/genética , Proteínas Señalizadoras YAP/genética , Carcinoma de Células Escamosas/patología , Polaridad Celular/genética , Células Epiteliales/metabolismo , Células Epiteliales/patología , Epitelio/metabolismo , Epitelio/patología , Receptores ErbB/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Pulmonares/patología , Lesiones Precancerosas/patología , Transducción de Señal/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ/genéticaRESUMEN
Vaping is suggested to be a risk factor for poor wound healing akin to smoking. However, the molecular and histologic mechanisms underlying this postulation remain unknown. Our study sought to compare molecular and histologic changes in cutaneous flap and non-flap tissue between vaping, smoking and control cohorts. Animal study of 15 male Sprague-Dawley rats was randomized to three cohorts: negative control (n = 5), e-cigarette (n = 5) and cigarette (n = 5) and exposed to their respective treatments with serum cotinine monitoring. After 30 days, random pattern flaps were raised and healed for 2 weeks after which skin punch biopsies of flap and non-flap tissues were collected for quantitative-reverse transcription-polymerase chain reaction of three selected wound healing genes (transforming growth factor ß [TGF-ß], vascular endothelial growth factor [VEGF], matrix metalloproteinase-1 [MMP-1]); then, immunohistochemistry for CD68 expression, α-smooth muscle actin looking at microvessel density (MVD) and in situ hybridization to localize VEGF production were undertaken. In flap tissue, vaping (mean[SEM]) (0.61[0.07]) and smoking (0.70[0.04]) were associated with decreased fold change of VEGF expression compared with controls (0.91[0.03]) (p < 0.05, p < 0.05, respectively). In non-flap tissue, only vaping was associated with decreased VEGF expression (mean[SEM]) (0.81[0.07]), compared with controls (1.17[0.10]) (p < 0.05) with expression primarily localized to basal keratinocytes and dermal capillaries. Immunohistochemistry showed decreased MVD in smoking (0.27[0.06]) and vaping (0.26[0.04]) flap tissue compared to matched controls (0.65[0.14]) (p < 0.05, p < 0.05, respectively) and decreased areas of fibrosis compared with controls on gross histology. Vaping and smoking were similarly associated with decreased VEGF expression, MVD and fibrotic changes in flap tissue. The results suggest attenuated angiogenesis via decreased VEGF expression as a mechanism for poor wound healing in vaping-exposed rats.
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Sistemas Electrónicos de Liberación de Nicotina , Vapeo , Animales , Masculino , Densidad Microvascular , Ratas , Ratas Sprague-Dawley , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial Vascular , Cicatrización de HeridasRESUMEN
Lymphatic muscle cells (LMCs) are indispensable for lymphatic vessel contraction and their aberrant recruitment or absence is associated with both primary and secondary lymphedema. Despite their critical role in lymphatic vessel function, the transcriptomic and developmental basis that confer the unique contractile properties to LMCs are largely undefined. In this study, we employed single-cell RNA sequencing (scRNAseq), lineage tracing and in vivo imaging to investigate the basis for the hybrid cardiomyocyte and blood vascular smooth muscle cell (SMC) characteristics that have been described for LMCs. Using scRNAseq, the transcriptomes of LMC and venous SMCs from the murine hindlimb exhibited more similarities than differences, although both were markedly distinct from that of arteriole SMCs in the same tissue. Functionally, both lymphatic vessels and blood vessels in the murine hindlimb displayed pulsatile contractility. However, despite expressing genes that overlap with the venous SMC transcriptome, through lineage tracing we show that LMCs do not originate from Myh11+ SMC progenitors. Previous studies have shown that LMCs express cardiac-related genes, whereas in our study we found that arteriole SMCs, but not LMCs, expressed cardiac-related genes. Through lineage tracing, we demonstrate that a subpopulation of LMCs and SMCs originate from WT1+ mesodermal progenitors, which are known to give rise to SMCs. LMCs, however, do not derive from Nkx2.5+ cardiomyocyte progenitors. Overall, our findings suggest that venous SMCs and LMCs and may derive from a related mesodermal progenitor and adopt a similar gene expression program that enable their contractile properties.
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Microscopic vascular invasion (VI) is predictive of recurrence and benefit from lobectomy in stage I lung adenocarcinoma (LUAD) but is difficult to assess in resection specimens and cannot be accurately predicted prior to surgery. Thus, new biomarkers are needed to identify this aggressive subset of stage I LUAD tumors. To assess molecular and microenvironment features associated with angioinvasive LUAD we profiled 162 resected stage I tumors with and without VI by RNA-seq and explored spatial patterns of gene expression in a subset of 15 samples by high-resolution spatial transcriptomics (stRNA-seq). Despite the small size of invaded blood vessels, we identified a gene expression signature of VI from the bulk RNA-seq discovery cohort (n=103) and found that it was associated with VI foci, desmoplastic stroma, and high-grade patterns in our stRNA-seq data. We observed a stronger association with high-grade patterns from VI+ compared with VI- tumors. Using the discovery cohort, we developed a transcriptomic predictor of VI, that in an independent validation cohort (n=60) was associated with VI (AUROC=0.86; p=5.42×10-6) and predictive of recurrence-free survival (HR=1.98; p=0.024), even in VI- LUAD (HR=2.76; p=0.003). To determine our VI predictor's robustness to intra-tumor heterogeneity we used RNA-seq data from multi-region sampling of stage I LUAD cases in TRACERx, where the predictor scores showed high correlation (R=0.87, p<2.2×10-16) between two randomly sampled regions of the same tumor. Our study suggests that VI-associated gene expression changes are detectable beyond the site of intravasation and can be used to predict the presence of VI. This may enable the prediction of angioinvasive LUAD from biopsy specimens, allowing for more tailored medical and surgical management of stage I LUAD.
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Macrophages are critical for maintenance and repair of mucosal tissues. While functionally distinct subtypes of macrophage are known to have important roles in injury response and repair in the lungs, little is known about macrophages in the proximal conducting airways. Single-cell RNA sequencing and flow cytometry demonstrated murine tracheal macrophages are largely monocyte-derived and are phenotypically distinct from lung macrophages at homeostasis. Following sterile airway injury, monocyte-derived macrophages are recruited to the trachea and activate a pro-regenerative phenotype associated with wound healing. Animals lacking the chemokine receptor CCR2 have reduced numbers of circulating monocytes and tracheal macrophages, deficient pro-regenerative macrophage activation and defective epithelial repair. Together, these studies indicate that recruitment and activation of monocyte-derived tracheal macrophages is CCR2-dependent and is required for normal airway epithelial regeneration.
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Repetitive head impacts (RHI) sustained from contact sports are the largest risk factor for chronic traumatic encephalopathy (CTE). Currently, CTE can only be diagnosed after death and the multicellular cascade of events that trigger initial hyperphosphorylated tau (p-tau) deposition remain unclear. Further, the symptoms endorsed by young individuals with early disease are not fully explained by the extent of p-tau deposition, severely hampering development of therapeutic interventions. Here, we show that RHI exposure associates with a multicellular response in young individuals (<51 years old) prior to the onset of CTE p-tau pathology that correlates with number of years of RHI exposure. Leveraging single nucleus RNA sequencing of tissue from 8 control, 9 RHI-exposed, and 11 low stage CTE individuals, we identify SPP1+ inflammatory microglia, angiogenic and inflamed endothelial cell profiles, reactive astrocytes, and altered synaptic gene expression in excitatory and inhibitory neurons in all individuals with exposure to RHI. Surprisingly, we also observe a significant loss of cortical sulcus layer 2/3 neurons in contact sport athletes compared to controls independent of p-tau pathology. These results provide robust evidence that multiple years of RHI exposure is sufficient to induce lasting cellular alterations that may underlie p-tau deposition and help explain the early clinical symptoms observed in young former contact sport athletes. Furthermore, these data identify specific cellular responses to repetitive head impacts that may direct future identification of diagnostic and therapeutic strategies for CTE.
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Lung cancer, the leading cause of cancer mortality, exhibits diverse histological subtypes and genetic complexities. Numerous preclinical mouse models have been developed to study lung cancer, but data from these models are disparate, siloed, and difficult to compare in a centralized fashion. Here we established the Lung Cancer Mouse Model Database (LCMMDB), an extensive repository of 1,354 samples from 77 transcriptomic datasets covering 974 samples from genetically engineered mouse models (GEMMs), 368 samples from carcinogen-induced models, and 12 samples from a spontaneous model. Meticulous curation and collaboration with data depositors have produced a robust and comprehensive database, enhancing the fidelity of the genetic landscape it depicts. The LCMMDB aligns 859 tumors from GEMMs with human lung cancer mutations, enabling comparative analysis and revealing a pressing need to broaden the diversity of genetic aberrations modeled in GEMMs. Accompanying this resource, we developed a web application that offers researchers intuitive tools for in-depth gene expression analysis. With standardized reprocessing of gene expression data, the LCMMDB serves as a powerful platform for cross-study comparison and lays the groundwork for future research, aiming to bridge the gap between mouse models and human lung cancer for improved translational relevance.
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BACKGROUND: Bronchial premalignant lesions (PMLs) are composed primarily of cells resembling basal epithelial cells of the airways, which through poorly understood mechanisms have the potential to progress to lung squamous cell carcinoma (LUSC). Despite ongoing efforts that have mapped gene expression and cell diversity across bronchial PML pathologies, signaling and transcriptional events driving malignancy are poorly understood. Evidence has suggested key roles for the Hippo pathway effectors YAP and TAZ and associated TEAD and TP63 transcription factor families in bronchial basal cell biology and LUSC. In this study we examine the functional association of YAP/TAZ, TEADs and TP63 in bronchial epithelial cells and PMLs. METHODS: We performed RNA-seq in primary human bronchial epithelial cells following small interfering RNA (siRNA)-mediated depletion of YAP/TAZ, TEADs or TP63, and combined these data with ChIP-seq analysis of these factors. Directly activated or repressed genes were identified and overlapping genes were profiled across gene expression data obtained from progressive or regressive human PMLs and across lung single cell RNA-seq data sets. RESULTS: Analysis of genes regulated by YAP/TAZ, TEADs, and TP63 in human bronchial epithelial cells revealed a converged transcriptional network that is strongly associated with the pathological progression of bronchial PMLs. Our observations suggest that YAP/TAZ-TEAD-TP63 associate to cooperatively promote basal epithelial cell proliferation and repress signals associated with interferon responses and immune cell communication. Directly repressed targets we identified include the MHC Class II transactivator CIITA, which is repressed in progressive PMLs and associates with adaptive immune responses in the lung. Our findings provide molecular insight into the control of gene expression events driving PML progression, including those contributing to immune evasion, offering potential new avenues for lung cancer interception. CONCLUSIONS: Our study identifies important gene regulatory functions for YAP/TAZ-TEAD-TP63 in the early stages of lung cancer development, which notably includes immune-suppressive roles, and suggest that an assessment of the activity of this transcriptional complex may offer a means to identify immune evasive bronchial PMLs and serve as a potential therapeutic target.
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Carcinoma de Células Escamosas , Neoplasias Pulmonares , Lesiones Precancerosas , Humanos , Regulación de la Expresión Génica , Neoplasias Pulmonares/genética , Lesiones Precancerosas/genética , Transducción de Señal , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Proteínas Señalizadoras YAP , Factores de Transcripción de Dominio TEARESUMEN
A greater understanding of molecular, cellular, and immunological changes during the early stages of lung adenocarcinoma development could improve diagnostic and therapeutic approaches in patients with pulmonary nodules at risk for lung cancer. To elucidate the immunopathogenesis of early lung tumorigenesis, we evaluated surgically resected pulmonary nodules representing the spectrum of early lung adenocarcinoma as well as associated normal lung tissues using single-cell RNA sequencing and validated the results by flow cytometry and multiplex immunofluorescence (MIF). Single-cell transcriptomics revealed a significant decrease in gene expression associated with cytolytic activities of tumor-infiltrating natural killer and natural killer T cells. This was accompanied by a reduction in effector T cells and an increase of CD4+ regulatory T cells (Treg) in subsolid nodules. An independent set of resected pulmonary nodules consisting of both adenocarcinomas and associated premalignant lesions corroborated the early increment of Tregs in premalignant lesions compared with the associated normal lung tissues by MIF. Gene expression analysis indicated that cancer-associated alveolar type 2 cells and fibroblasts may contribute to the deregulation of the extracellular matrix, potentially affecting immune infiltration in subsolid nodules through ligand-receptor interactions. These findings suggest that there is a suppression of immune surveillance across the spectrum of early-stage lung adenocarcinoma. SIGNIFICANCE: Analysis of a spectrum of subsolid pulmonary nodules by single-cell RNA sequencing provides insights into the immune regulation and cell-cell interactions in the tumor microenvironment during early lung tumor development.
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Adenocarcinoma del Pulmón , Adenocarcinoma , Neoplasias Pulmonares , Nódulos Pulmonares Múltiples , Humanos , Monitorización Inmunológica , Tomografía Computarizada por Rayos X/métodos , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/patología , Neoplasias Pulmonares/patología , Adenocarcinoma/genética , Adenocarcinoma/patología , Microambiente TumoralRESUMEN
Somatic mutations are a hallmark of tumorigenesis and may be useful for non-invasive diagnosis of cancer. We analyzed whole-genome sequencing data from 2,511 individuals in the Pan-Cancer Analysis of Whole Genomes (PCAWG) study as well as 489 individuals from four prospective cohorts and found distinct regional mutation type-specific frequencies in tissue and cell-free DNA from patients with cancer that were associated with replication timing and other chromatin features. A machine-learning model using genome-wide mutational profiles combined with other features and followed by CT imaging detected >90% of patients with lung cancer, including those with stage I and II disease. The fixed model was validated in an independent cohort, detected patients with cancer earlier than standard approaches and could be used to monitor response to therapy. This approach lays the groundwork for non-invasive cancer detection using genome-wide mutation features that may facilitate cancer screening and monitoring.
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Ácidos Nucleicos Libres de Células , Neoplasias Pulmonares , Neoplasias , Humanos , Estudios Prospectivos , Mutación , Neoplasias/diagnóstico , Neoplasias/genética , Tasa de Mutación , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genéticaRESUMEN
SARS-CoV-2 infection and disease severity are influenced by viral entry (VE) gene expression patterns in the airway epithelium. The similarities and differences of VE gene expression (ACE2, TMPRSS2, and CTSL) across nasal and bronchial compartments have not been fully characterized using matched samples from large cohorts. Gene expression data from 793 nasal and 1673 bronchial brushes obtained from individuals participating in lung cancer screening or diagnostic workup revealed that smoking status (current versus former) was the only clinical factor significantly and reproducibly associated with VE gene expression. The expression of ACE2 and TMPRSS2 was higher in smokers in the bronchus but not in the nose. scRNA-seq of nasal brushings indicated that ACE2 co-expressed genes were highly expressed in club and C15orf48+ secretory cells while TMPRSS2 co-expressed genes were highly expressed in keratinizing epithelial cells. In contrast, these ACE2 and TMPRSS2 modules were highly expressed in goblet cells in scRNA-seq from bronchial brushings. Cell-type deconvolution of the gene expression data confirmed that smoking increased the abundance of several secretory cell populations in the bronchus, but only goblet cells in the nose. The association of ACE2 and TMPRSS2 with smoking in the bronchus is due to their high expression in goblet cells which increase in abundance in current smoker airways. In contrast, in the nose, these genes are not predominantly expressed in cell populations modulated by smoking. In individuals with elevated lung cancer risk, smoking-induced VE gene expression changes in the nose likely have minimal impact on SARS-CoV-2 infection, but in the bronchus, smoking may lead to higher viral loads and more severe disease.
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COVID-19 , Neoplasias Pulmonares , Humanos , SARS-CoV-2/genética , Enzima Convertidora de Angiotensina 2/genética , COVID-19/genética , Detección Precoz del Cáncer , Peptidil-Dipeptidasa A/metabolismo , Neoplasias Pulmonares/metabolismo , Bronquios/metabolismo , Fumar/efectos adversos , Fumar/genéticaRESUMEN
OBJECTIVE: The immune response to invasive carcinoma has been the focus of published work, but little is known about the adaptive immune response to bronchial premalignant lesions (PMLs), precursors of lung squamous cell carcinoma. This study was designed to characterize the T cell receptor (TCR) repertoire in PMLs and its association with clinical, pathological, and molecular features. METHODS: Endobronchial biopsies (n=295) and brushings (n=137) from high-risk subjects (n=50), undergoing lung cancer screening at approximately 1-year intervals via autofluorescence bronchoscopy and CT, were profiled by RNA-seq. We applied the TCR Repertoire Utilities for Solid Tissue/Tumor tool to the RNA-seq data to identify TCR CDR3 sequences across all samples. In the biopsies, we measured the correlation of TCR diversity with previously derived immune-associated PML transcriptional signatures and PML outcome. We also quantified the spatial and temporal distribution of shared and clonally expanded TCRs. Using the biopsies and brushes, the ratio of private (ie, found in one patient only) and public (ie, found in two or more patients) TCRs was quantified, and the CDR3 sequences were compared with those found in curated databases with known antigen specificities. RESULTS: We detected 39,303 unique TCR sequences across all samples. In PML biopsies, TCR diversity was negatively associated with a transcriptional signature of T cell mediated immune activation (p=4e-4) associated with PML outcome. Additionally, in lesions of the proliferative molecular subtype, TCR diversity was decreased in regressive versus progressive/persistent PMLs (p=0.045). Within each patient, TCRs were more likely to be shared between biopsies sampled at the same timepoint than biopsies sampled at the same anatomic location at different times. Clonally expanded TCRs, within a biopsied lesion, were more likely to be expanded at future time points than non-expanded clones. The majority of TCR sequences were found in a single sample, with only 3396 (8.6%) found in more than one sample and 1057 (2.7%) found in two or more patients (ie, public); however, when compared with a public database of CDR3 sequences, 4543 (11.6%) of TCRs were identified as public. TCRs with known antigen specificities were enriched among public TCRs (p<0.001). CONCLUSIONS: Decreased TCR diversity may reflect nascent immune responses that contribute to PML elimination. Further studies are needed to explore the potential for immunoprevention of PMLs.
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Neoplasias Pulmonares/genética , Neoplasias de Células Escamosas/genética , Linfocitos T/inmunología , Progresión de la Enfermedad , Femenino , Humanos , MasculinoRESUMEN
Background : SARS-CoV-2 infection and disease severity are influenced by viral entry (VE) gene expression patterns in airway epithelium. The similarities and differences of VE gene expression (ACE2, TMPRSS2, and CTSL) across nasal and bronchial compartments has not been fully characterized using matched samples from large cohorts. Results : Gene expression data from 793 nasal and 1,673 bronchial brushes obtained from individuals participating in lung cancer screening or diagnostic workup revealed that smoking was the only clinical factor significantly and reproducibly associated with VE gene expression. ACE2 and TMPRSS2 expression were higher in smokers in the bronchus but not in the nose. scRNA-seq of nasal brushings indicated that ACE2 co-expressed genes were highly expressed in club and C15orf48 + secretory cells while TMPRSS2 co-expressed genes were highly expressed in keratinizing epithelial cells. In contrast, these ACE2 and TMPRSS2 modules were highly expressed in goblet cells in scRNA-seq from bronchial brushings. Cell-type deconvolution of the RNA-seq confirmed that smoking increased the abundance of several secretory cell populations in the bronchus, but only goblet cells in the nose. Conclusions : The association of ACE2 and TMPRSS2 with smoking in the bronchus is due to their high expression in goblet cells which increase in abundance in current smoker airways. In contrast, in the nose these genes are not predominantly expressed in cell populations modulated by smoking. Smoking-induced VE gene expression changes in the nose likely has minimal impact on SARS-CoV-2 infection, but in the bronchus, smoking may lead to higher viral loads and more severe disease.
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There is an increasing appreciation for the heterogeneity of myeloid lineages in the lung, but relatively little is known about populations specifically associated with the conducting airways. We use single-cell RNA sequencing, flow cytometry, and immunofluorescence to characterize myeloid cells of the mouse trachea during homeostasis and epithelial injury/repair. We identify submucosal macrophages, similar to lung interstitial macrophages, and intraepithelial macrophages. Following injury, there are early increases in neutrophils and submucosal macrophages, including M2-like macrophages. Intraepithelial macrophages are lost after injury and later restored by CCR2+ monocytes. We show that repair of the tracheal epithelium is impaired in Ccr2-deficient mice. Mast cells and group 2 innate lymphoid cells are sources of interleukin-13 (IL-13) that polarize macrophages and directly influence basal cell behaviors. Their proximity to the airway epithelium establishes these myeloid populations as potential therapeutic targets for airway disease.
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Células Epiteliales/metabolismo , Epitelio/metabolismo , Homeostasis , Macrófagos Alveolares/fisiología , Células Mieloides/fisiología , Receptores CCR2/metabolismo , Tráquea/metabolismo , Animales , Células Cultivadas , Citocinas/metabolismo , Epitelio/lesiones , Femenino , Pulmón/metabolismo , Lesión Pulmonar/inducido químicamente , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Animales , Monocitos/metabolismo , Polidocanol , Receptores CCR2/genética , Regeneración , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Tráquea/lesionesRESUMEN
Bronchial premalignant lesions (PMLs) are precursors of lung squamous cell carcinoma, but have variable outcome, and we lack tools to identify and treat PMLs at risk for progression to cancer. Here we report the identification of four molecular subtypes of PMLs with distinct differences in epithelial and immune processes based on RNA-Seq profiling of endobronchial biopsies from high-risk smokers. The Proliferative subtype is enriched with bronchial dysplasia and exhibits up-regulation of metabolic and cell cycle pathways. A Proliferative subtype-associated gene signature identifies subjects with Proliferative PMLs from normal-appearing uninvolved large airway brushings with high specificity. In progressive/persistent Proliferative lesions expression of interferon signaling and antigen processing/presentation pathways decrease and immunofluorescence indicates a depletion of innate and adaptive immune cells compared with regressive lesions. Molecular biomarkers measured in PMLs or the uninvolved airway can enhance histopathological grading and suggest immunoprevention strategies for intercepting the progression of PMLs to lung cancer.
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Biomarcadores de Tumor/genética , Carcinoma Broncogénico/patología , Regulación Neoplásica de la Expresión Génica/inmunología , Neoplasias Pulmonares/patología , Lesiones Precancerosas/patología , Antineoplásicos Inmunológicos/farmacología , Antineoplásicos Inmunológicos/uso terapéutico , Biomarcadores de Tumor/inmunología , Biopsia , Bronquios/diagnóstico por imagen , Bronquios/inmunología , Bronquios/patología , Broncoscopía , Carcinoma Broncogénico/genética , Carcinoma Broncogénico/inmunología , Carcinoma Broncogénico/prevención & control , Estudios de Cohortes , Conjuntos de Datos como Asunto , Progresión de la Enfermedad , Detección Precoz del Cáncer/métodos , Perfilación de la Expresión Génica , Redes Reguladoras de Genes/genética , Redes Reguladoras de Genes/inmunología , Humanos , Inmunidad Celular/efectos de los fármacos , Inmunidad Celular/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/prevención & control , Tamizaje Masivo/métodos , Persona de Mediana Edad , Lesiones Precancerosas/diagnóstico por imagen , Lesiones Precancerosas/genética , Lesiones Precancerosas/inmunología , ARN Mensajero/genética , Mucosa Respiratoria/citología , Mucosa Respiratoria/diagnóstico por imagen , Mucosa Respiratoria/inmunología , Mucosa Respiratoria/patología , Análisis de Secuencia de ARN , Linfocitos T/inmunología , Tomografía Computarizada por Rayos X , Regulación hacia ArribaRESUMEN
The human bronchial epithelium is composed of multiple distinct cell types that cooperate to defend against environmental insults. While studies have shown that smoking alters bronchial epithelial function and morphology, its precise effects on specific cell types and overall tissue composition are unclear. We used single-cell RNA sequencing to profile bronchial epithelial cells from six never and six current smokers. Unsupervised analyses led to the characterization of a set of toxin metabolism genes that localized to smoker ciliated cells, tissue remodeling associated with a loss of club cells and extensive goblet cell hyperplasia, and a previously unidentified peri-goblet epithelial subpopulation in smokers who expressed a marker of bronchial premalignant lesions. Our data demonstrate that smoke exposure drives a complex landscape of cellular alterations that may prime the human bronchial epithelium for disease.