Molecular taxonomy of human ocular outflow tissues defined by single cell transcriptomics

ABSTRACTThe conventional outflow pathway is a complex tissue responsible for maintaining intraocular pressure (IOP) homeostasis. The coordinated effort of multiple cells with differing responsibilities ensure healthy outflow function and IOP maintenance. Dysfunction of one or more resident cell type results in ocular hypertension and risk for glaucoma, a leading cause of blindness. In this study, single cell RNA sequencing was performed to generate a comprehensive cell atlas of human conventional outflow tissues. We obtained 17757 genes expression profiles from 8758 cells from eight eyes of four donors representing the outflow cell transcriptome. Upon clustering analysis, 12 distinct cell types were identified, and region-specific expression of candidate genes were mapped in human tissues. Significantly, we identified two distinct expression patterns (myofibroblast and fibroblast) from cells located in the trabecular meshwork (TM), the primary structural component of the conventional outflow pathway. We also located neuron and macrophage signatures in the TM. The second primary component structure, Schlemm’s canal displayed a unique combination of lymphatic/blood vascular gene expression. Other expression clusters corresponded to cells from neighboring tissues, predominantly in the ciliary muscle/scleral spur, which together correspond to the uveoscleral outflow path. Importantly, the utility of our atlas was demonstrated by mapping glaucoma-relevant genes to outflow cell clusters. Our study provides a comprehensive molecular and cellular classification of conventional and unconventional outflow pathway structures responsible for IOP homeostasis.Significance statementOcular hypertension is the primary, and only modifiable risk factor for glaucoma, the leading cause of irreversible blindness. Intraocular pressure is regulated homeostatically by resistance to aqueous humor outflow through an architecturally complex tissue, the conventional/trabecular pathway. In this study, we generated a comprehensive cell atlas of the human trabecular meshwork and neighboring tissues using single cell, RNA sequencing. We identified 12 distinct cell types, and mapped region-specific expression of candidate genes. The utility of our atlas was demonstrated by mapping glaucoma-relevant genes to conventional outflow cell clusters. Our study provides a comprehensive molecular and cellular classification of tissue structures responsible for intraocular pressure homeostasis in health, and dysregulation in disease.

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Distinguishing cells from empty droplets in droplet-based single-cell RNA sequencing data

10.1101/234872 ◽

2018 ◽

Cited By ~ 7

Author(s):

Aaron T. L. Lun ◽

Samantha Riesenfeld ◽

Tallulah Andrews ◽

Tomas Gomes ◽

John C. Marioni ◽

...

Keyword(s):

Single Cell ◽

Rna Sequencing ◽

Real Data ◽

Cell Types ◽

Sequencing Data ◽

Minimum Threshold ◽

False Discovery ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

Unique Molecular Identifier

AbstractDroplet-based single-cell RNA sequencing protocols have dramatically increased the throughput and efficiency of single-cell transcriptomics studies. A key computational challenge when processing these data is to distinguish libraries for real cells from empty droplets. Existing methods for cell calling set a minimum threshold on the total unique molecular identifier (UMI) count for each library, which indiscriminately discards cell libraries with low UMI counts. Here, we describe a new statistical method for calling cells from droplet-based data, based on detecting significant deviations from the expression profile of the ambient solution. Using simulations, we demonstrate that our method has greater power than existing approaches for detecting cell libraries with low UMI counts, while controlling the false discovery rate among detected cells. We also apply our method to real data, where we show that the use of our method results in the retention of distinct cell types that would otherwise have been discarded.

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Understanding human gut diseases at single-cell resolution

Human Molecular Genetics ◽

10.1093/hmg/ddaa130 ◽

2020 ◽

Vol 29 (R1) ◽

pp. R51-R58 ◽

Cited By ~ 1

Author(s):

Emilia Bigaeva ◽

Werna T C Uniken Venema ◽

Rinse K Weersma ◽

Eleonora A M Festen

Keyword(s):

Single Cell ◽

Digestive System ◽

Cell Types ◽

Specific Cell ◽

Human Gut ◽

The Past ◽

Distinct Cell ◽

Transcriptional Variation ◽

Single Cell Rna Sequencing ◽

Inflammatory Bowel

Abstract Our understanding of gut functioning and pathophysiology has grown considerably in the past decades, and advancing technologies enable us to deepen this understanding. Single-cell RNA sequencing (scRNA-seq) has opened a new realm of cellular diversity and transcriptional variation in the human gut at a high, single-cell resolution. ScRNA-seq has pushed the science of the digestive system forward by characterizing the function of distinct cell types within complex intestinal cellular environments, by illuminating the heterogeneity within specific cell populations and by identifying novel cell types in the human gut that could contribute to a variety of intestinal diseases. In this review, we highlight recent discoveries made with scRNA-seq that significantly advance our understanding of the human gut both in health and across the spectrum of gut diseases, including inflammatory bowel disease, colorectal carcinoma and celiac disease.

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XYZeq: Spatially resolved single-cell RNA sequencing reveals expression heterogeneity in the tumor microenvironment

Science Advances ◽

10.1126/sciadv.abg4755 ◽

2021 ◽

Vol 7 (17) ◽

pp. eabg4755

Author(s):

Youjin Lee ◽

Derek Bogdanoff ◽

Yutong Wang ◽

George C. Hartoularos ◽

Jonathan M. Woo ◽

...

Keyword(s):

Single Cell ◽

Rna Sequencing ◽

Spatial Organization ◽

Cell Types ◽

Mouse Tumor ◽

Spatially Resolved ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

A Cell ◽

Anatomical Space

Single-cell RNA sequencing (scRNA-seq) of tissues has revealed remarkable heterogeneity of cell types and states but does not provide information on the spatial organization of cells. To better understand how individual cells function within an anatomical space, we developed XYZeq, a workflow that encodes spatial metadata into scRNA-seq libraries. We used XYZeq to profile mouse tumor models to capture spatially barcoded transcriptomes from tens of thousands of cells. Analyses of these data revealed the spatial distribution of distinct cell types and a cell migration-associated transcriptomic program in tumor-associated mesenchymal stem cells (MSCs). Furthermore, we identify localized expression of tumor suppressor genes by MSCs that vary with proximity to the tumor core. We demonstrate that XYZeq can be used to map the transcriptome and spatial localization of individual cells in situ to reveal how cell composition and cell states can be affected by location within complex pathological tissue.

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Single cell RNA sequencing of theStrongylocentrotus purpuratuslarva reveals the blueprint of major cell types and nervous system of a non-chordate deuterostome

10.1101/2021.03.16.435574 ◽

2021 ◽

Author(s):

Periklis Paganos ◽

Danila Voronov ◽

Jacob Musser ◽

Detlev Arendt ◽

Maria I. Arnone

Keyword(s):

Single Cell ◽

Rna Sequencing ◽

Sea Urchin ◽

Regulatory Networks ◽

Neuronal Cell ◽

Cell Types ◽

Molecular Fingerprint ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

Cell Diversity

AbstractIdentifying the molecular fingerprint of organismal cell types is key for understanding their function and evolution. Here, we use single cell RNA sequencing (scRNA-seq) to survey the cell types of the sea urchin early pluteus larva, representing an important developmental transition from non-feeding to feeding larva. We identified 21 distinct cell clusters, representing cells of the digestive, skeletal, immune, and nervous systems. Further subclustering of these revealed a highly detailed portrait of cell diversity across the larva, including the identification of 12 distinct neuronal cell types. Moreover, we corroborated co-expression of key regulatory genes previously shown to drive sea urchin gene regulatory networks, and revealed additional domains in which these regulatory networks are likely to function within the larva. Lastly, we recovered a neuronal cell type co-expressingPdx-1andBrn1/2/4, which had previously been shown to share similar gene expression with vertebrate pancreas. Our results extend this finding, revealing twenty transcription factors shared by this population of neurons in sea urchin and vertebrate pancreatic cells. Using differential expression results from Pdx-1 knockdown experiments, we generate a draft of the Pdx-1 regulatory network in these cells, and hypothesize this network was present in an ancestral deuterostome neuron before being co-opted into the pancreas developmental lineage in vertebrates.

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Single-Cell Regulatory Network Inference and Clustering Identifies Cell-Type Specific Expression Pattern of Transcription Factors in Mouse Sciatic Nerve

Frontiers in Cellular Neuroscience ◽

10.3389/fncel.2021.676515 ◽

2021 ◽

Vol 15 ◽

Author(s):

Mingchao Li ◽

Qing Min ◽

Matthew C. Banton ◽

Xinpeng Dun

Keyword(s):

Transcription Factors ◽

Sciatic Nerve ◽

Single Cell ◽

Cell Types ◽

Cell Type ◽

Specific Expression ◽

Single Cell Rna Sequencing ◽

Cell Type Specific Expression ◽

Cell Type Specific ◽

Different Cell Types

Advances in single-cell RNA sequencing technologies and bioinformatics methods allow for both the identification of cell types in a complex tissue and the large-scale gene expression profiling of various cell types in a mixture. In this report, we analyzed a single-cell RNA sequencing (scRNA-seq) dataset for the intact adult mouse sciatic nerve and examined cell-type specific transcription factor expression and activity during peripheral nerve homeostasis. In total, we identified 238 transcription factors expressed in nine different cell types of intact mouse sciatic nerve. Vascular smooth muscle cells have the lowest number of transcription factors expressed with 17 transcription factors identified. Myelinating Schwann cells (mSCs) have the highest number of transcription factors expressed, with 61 transcription factors identified. We created a cell-type specific expression map for the identified 238 transcription factors. Our results not only provide valuable information about the expression pattern of transcription factors in different cell types of adult peripheral nerves but also facilitate future studies to understand the function of key transcription factors in the peripheral nerve homeostasis and disease.

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Single-cell Transcriptomics Identify Extensive Heterogeneity in the Cellular Composition of Mouse Achilles Tendons

10.1101/801266 ◽

2019 ◽

Cited By ~ 4

Author(s):

Andrea J De Micheli ◽

Jacob B Swanson ◽

Nathaniel P Disser ◽

Leandro M Martinez ◽

Nicholas R Walker ◽

...

Keyword(s):

Single Cell ◽

Cell Types ◽

Tissue Homeostasis ◽

Cellular Heterogeneity ◽

Cell Populations ◽

Cellular Composition ◽

Matrix Assembly ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

Additional Support

AbstractTendon is a connective tissue that transmits forces between muscles and bones. Cellular heterogeneity is increasingly recognized as an important factor in the biological basis of tissue homeostasis and disease, but little is known about the diversity of cells that populate tendon. Our objective was to explore the heterogeneity of cells in mouse Achilles tendons using single-cell RNA sequencing. We assembled a transcriptomic atlas and identified 11 distinct cell types in tendons, including 3 previously undescribed populations of fibroblasts. Using trajectory inference analysis, we provide additional support for the notion that pericytes are progenitor cells for the fibroblasts that compose adult tendons. We also modeled cell-interactions and identified ligand-receptor pairs involved in tendon homeostasis. Our findings highlight notable heterogeneity between and within tendon cell populations, which may contribute to our understanding of tendon extracellular matrix assembly and maintenance, and inform the design of therapies to treat tendinopathies.

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Single-cell RNA-sequencing reveals profibrotic roles of distinct epithelial and mesenchymal lineages in pulmonary fibrosis

10.1101/753806 ◽

2019 ◽

Cited By ~ 16

Author(s):

Arun C. Habermann ◽

Austin J. Gutierrez ◽

Linh T. Bui ◽

Stephanie L. Yahn ◽

Nichelle I. Winters ◽

...

Keyword(s):

Epithelial Cell ◽

Pulmonary Fibrosis ◽

Single Cell ◽

Rna Sequencing ◽

Cell Types ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

Epithelial Remodeling ◽

Discrete Manner ◽

Cell Phenotypes

AbstractPulmonary fibrosis is a form of chronic lung disease characterized by pathologic epithelial remodeling and accumulation of extracellular matrix. In order to comprehensively define the cell types, mechanisms and mediators driving fibrotic remodeling in lungs with pulmonary fibrosis, we performed single-cell RNA-sequencing of single-cell suspensions from 10 non-fibrotic control and 20 PF lungs. Analysis of 114,396 cells identified 31 distinct cell types. We report a remarkable shift in epithelial cell phenotypes occurs in the peripheral lung in PF, and identify several previously unrecognized epithelial cell phenotypes including a KRT5−/KRT17+, pathologic ECM-producing epithelial cell population that was highly enriched in PF lungs. Multiple fibroblast subtypes were observed to contribute to ECM expansion in a spatially-discrete manner. Together these data provide high-resolution insights into the complexity and plasticity of the distal lung epithelium in human disease, and indicate a diversity of epithelial and mesenchymal cells contribute to pathologic lung fibrosis.One Sentence SummarySingle-cell RNA-sequencing provides new insights into pathologic epithelial and mesenchymal remodeling in the human lung.

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Molecular taxonomy of human ocular outflow tissues defined by single-cell transcriptomics

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2001896117 ◽

2020 ◽

Vol 117 (23) ◽

pp. 12856-12867 ◽

Cited By ~ 5

Author(s):

Gaurang Patel ◽

Wen Fury ◽

Hua Yang ◽

Maria Gomez-Caraballo ◽

Yu Bai ◽

...

Keyword(s):

Single Cell ◽

Expression Profiles ◽

Expression Patterns ◽

Molecular Taxonomy ◽

Cell Types ◽

Primary Component ◽

Second Primary ◽

Specific Expression ◽

Component Structure ◽

Outflow Pathway

The conventional outflow pathway is a complex tissue responsible for maintaining intraocular pressure (IOP) homeostasis. The coordinated effort of multiple cells with differing responsibilities ensures healthy outflow function and IOP maintenance. Dysfunction of one or more resident cell types results in ocular hypertension and risk for glaucoma, a leading cause of blindness. In this study, single-cell RNA sequencing was performed to generate a comprehensive cell atlas of human conventional outflow tissues. We obtained expression profiles of 17,757 genes from 8,758 cells from eight eyes of human donors representing the outflow cell transcriptome. Upon clustering analysis, 12 distinct cell types were identified, and region-specific expression of candidate genes was mapped in human tissues. Significantly, we identified two distinct expression patterns (myofibroblast- and fibroblast-like) from cells located in the trabecular meshwork (TM), the primary structural component of the conventional outflow pathway. We also located Schwann cell and macrophage signatures in the TM. The second primary component structure, Schlemm’s canal, displayed a unique combination of lymphatic/blood vascular gene expression. Other expression clusters corresponded to cells from neighboring tissues, predominantly in the ciliary muscle/scleral spur, which together correspond to the uveoscleral outflow pathway. Importantly, the utility of our atlas was demonstrated by mapping glaucoma-relevant genes to outflow cell clusters. Our study provides a comprehensive molecular and cellular classification of conventional and unconventional outflow pathway structures responsible for IOP homeostasis.

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Unbiased classification of mosquito blood cells by single-cell genomics and high-content imaging

10.1101/234492 ◽

2017 ◽

Author(s):

Maiara S. Severo ◽

Jonathan J.M. Landry ◽

Randall L. Lindquist ◽

Christian Goosmann ◽

Volker Brinkmann ◽

...

Keyword(s):

Single Cell ◽

Single Molecule ◽

Blood Cells ◽

Cell Biology ◽

Cell Types ◽

Distinct Cell ◽

Cellular Marker ◽

Active Transfer ◽

Vector Borne

ABSTRACTMosquito blood cells are ancestral immune cells that help control infection by vector-borne pathogens. Despite their importance, little is known about mosquito blood cell biology beyond the ambiguous morphological and functional criteria used for their classification. Here we combined the power of single-cell RNA-sequencing, imaging flow cytometry and single-molecule RNA hybridization to analyze blood cells of the malaria mosquito Anopheles gambiae. By demonstrating that blood cells express nearly half of the mosquito transcriptome, our dataset represents an unprecedented view into their transcriptional machinery. Analyses of differentially expressed genes identified transcriptional signatures of two distinct cell types that challenge the current morphology-based classification of these cells. We further demonstrated an active transfer of a cellular marker between blood cells that confounds their identity. We propose that cell-to-cell exchange is broadly relevant for cell type classification and may account for the remarkable cellular diversity observed in nature.

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Single-cell RNA sequencing reveals profibrotic roles of distinct epithelial and mesenchymal lineages in pulmonary fibrosis

Science Advances ◽

10.1126/sciadv.aba1972 ◽

2020 ◽

Vol 6 (28) ◽

pp. eaba1972 ◽

Cited By ~ 36

Author(s):

Arun C. Habermann ◽

Austin J. Gutierrez ◽

Linh T. Bui ◽

Stephanie L. Yahn ◽

Nichelle I. Winters ◽

...

Keyword(s):

Epithelial Cell ◽

Pulmonary Fibrosis ◽

Single Cell ◽

Rna Sequencing ◽

Cell Types ◽

Distinct Cell ◽

Single Cell Rna Sequencing ◽

Epithelial Remodeling ◽

Discrete Manner ◽

Cell Phenotypes

Pulmonary fibrosis (PF) is a form of chronic lung disease characterized by pathologic epithelial remodeling and accumulation of extracellular matrix (ECM). To comprehensively define the cell types, mechanisms, and mediators driving fibrotic remodeling in lungs with PF, we performed single-cell RNA sequencing of single-cell suspensions from 10 nonfibrotic control and 20 PF lungs. Analysis of 114,396 cells identified 31 distinct cell subsets/states. We report that a remarkable shift in epithelial cell phenotypes occurs in the peripheral lung in PF and identify several previously unrecognized epithelial cell phenotypes, including a KRT5−/KRT17+ pathologic, ECM-producing epithelial cell population that was highly enriched in PF lungs. Multiple fibroblast subtypes were observed to contribute to ECM expansion in a spatially discrete manner. Together, these data provide high-resolution insights into the complexity and plasticity of the distal lung epithelium in human disease and indicate a diversity of epithelial and mesenchymal cells contribute to pathologic lung fibrosis.

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