scholarly journals Single cell transcriptomics identifies a unique adipocyte population that regulates bone marrow environment

2019 ◽  
Author(s):  
Leilei Zhong ◽  
Lutian Yao ◽  
Robert J. Tower ◽  
Yulong Wei ◽  
Zhen Miao ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Single Cell ◽  
De Novo ◽  
Bone Homeostasis ◽  
Genetic Ablation ◽  
Lineage Differentiation ◽  
Vessel Structure ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

AbstractBone marrow mesenchymal lineage cells are a heterogeneous cell population involved in bone homeostasis and diseases such as osteoporosis. While it is long postulated that they originate from mesenchymal stem cells (MSCs), the true identity of MSCs and their in vivo bifurcated differentiation routes into osteoblasts and adipocytes remain poorly understood. Here, by employing single cell transcriptome analysis, we identified MSCs and delineated their bi-lineage differentiation paths in young, adult and aging mice. Among several newly discovered mesenchymal subpopulations, one is a distinct population of adipose-lineage cells that we named marrow environment regulating adipose cells (MERAs). MERAs are non-proliferative, post-progenitor cells that express many mature adipocyte markers but are devoid of lipid droplets. They are abundant in the bone marrow of young mice, acting as pericytes and stromal cells that form numerous connections among themselves and with other cells inside bone, including endothelial cells. Genetic ablation of MERAs disrupts marrow vessel structure, promotes de novo bone formation. Taken together, MERAs represent a unique population of adipose lineage cells that exist only in the bone marrow with critical roles in regulating bone and vessel homeostasis.

scholarly journals Single cell transcriptome analysis reveals an essential role for Gata2b in hematopoietic lineage decisions in zebrafish

2019 ◽  
Author(s):  
Emanuele Gioacchino ◽  
Cansu Koyunlar ◽  
Hans de Looper ◽  
Madelon de Jong ◽  
Tomasz Dobrzycki ◽  
...  
Keyword(s):  
Single Cell ◽  
Transcriptome Analysis ◽  
Progenitor Cell ◽  
Adult Zebrafish ◽  
Hematopoietic Stem ◽  
Lineage Differentiation ◽  
Hematopoietic Lineage ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome ◽  
Monocytic Lineage

AbstractHematopoietic stem cells (HSCs) are tightly controlled to keep a balance between myeloid and lymphoid cell differentiation. Gata2 is a pivotal hematopoietic transcription factor required for HSC generation and maintenance. We generated a zebrafish mutant for the mammalianGata2orthologue,gata2b. We found that in adult zebrafish,gata2bis required for both neutrophilic- and monocytic lineage differentiation. Single cell transcriptome analysis revealed that the myeloid defect present in Gata2b deficient zebrafish arise in the most immature hematopoietic stem and progenitor cell (HSPC) compartment and that this population is instead committed towards the lymphoid and erythroid lineage. Taken together, we find that Gata2b is vital for the fate choice between the myeloid and lymphoid lineages.

scholarly journals Mesenchymal Stromal Cells in the Bone Marrow Niche Consist of Multi-Populations With Distinct Transcriptional and Epigenetic Properties

2021 ◽  
Author(s):  
Sanshiro Kanazawa ◽  
Hironori Hojo ◽  
Shinsuke Ohba ◽  
Junichi Iwata ◽  
Makoto Komura ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Single Cell ◽  
Expression Profiles ◽  
Marker Genes ◽  
Mouse Bone Marrow ◽  
Bone Marrow Niche ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

Abstract Although multiple studies have investigated the mesenchymal stem and progenitor cells (MSCs) that give rise to mature bone marrow, high heterogeneity in their morphologies and properties causes difficulties in molecular separation of their distinct populations. In this study, by taking advantage of the resolution of the single cell transcriptome, we analyzed Sca-1 and PDGFR-α fraction in the mouse bone marrow tissue. The single cell transcriptome enabled us to further classify the population into seven populations according to their gene expression profiles. We then separately obtained the seven populations based on candidate marker genes, and specified their gene expression properties and epigenetic landscape by ATAC-seq. Our findings will enable to elucidate the stem cell niche signal in the bone marrow microenvironment, reconstitute bone marrow in vitro, and shed light on the potentially important role of identified subpopulation in various clinical applications to the treatment of bone- and bone marrow-related diseases.

scholarly journals Droplet-based Single-cell Total RNA-seq Reveals Differential Non-Coding Expression and Splicing Patterns during Mouse Development

2021 ◽  
Author(s):  
Fredrik Salmen ◽  
Joachim De Jonghe ◽  
Tomasz S. Kaminski ◽  
Anna Alemany ◽  
Guillermo Parada ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Single Cell ◽  
Single Cells ◽  
Droplet Microfluidics ◽  
Mammalian Development ◽  
Mouse Development ◽  
Protein Coding ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

In recent years, single-cell transcriptome sequencing has revolutionized biology, allowing for the unbiased characterization of cellular subpopulations. However, most methods amplify the termini of polyadenylated transcripts capturing only a small fraction of the total cellular transcriptome. This precludes the detection of many long non-coding, short non-coding and non-polyadenylated protein-coding transcripts. Additionally, most workflows do not sequence the full transcript hindering the analysis of alternative splicing. We therefore developed VASA- seq to detect the total transcriptome in single cells. VASA-seq is compatible with both plate- based formats and droplet microfluidics. We applied VASA-seq to over 30,000 single cells in the developing mouse embryo during gastrulation and early organogenesis. The dynamics of the total single-cell transcriptome result in the discovery of novel cell type markers many based on non-coding RNA, an in vivo cell cycle analysis and an improved RNA velocity characterization. Moreover, it provides the first comprehensive analysis of alternative splicing during mammalian development.

scholarly journals Single-cell transcriptome identifies molecular subtype of autism spectrum disorder impacted by de novo loss-of-function variants regulating glial cells

2021 ◽  
Vol 15 (1) ◽  
Author(s):  
Nasna Nassir ◽  
Asma Bankapur ◽  
Bisan Samara ◽  
Abdulrahman Ali ◽  
Awab Ahmed ◽  
...  
Keyword(s):  
Single Cell ◽  
Large Scale ◽  
De Novo ◽  
Cell Types ◽  
Brain Regions ◽  
Autism Spectrum ◽  
Loss Of Function ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome ◽  
Variant Genes

Abstract Background In recent years, several hundred autism spectrum disorder (ASD) implicated genes have been discovered impacting a wide range of molecular pathways. However, the molecular underpinning of ASD, particularly from the point of view of ‘brain to behaviour’ pathogenic mechanisms, remains largely unknown. Methods We undertook a study to investigate patterns of spatiotemporal and cell type expression of ASD-implicated genes by integrating large-scale brain single-cell transcriptomes (> million cells) and de novo loss-of-function (LOF) ASD variants (impacting 852 genes from 40,122 cases). Results We identified multiple single-cell clusters from three distinct developmental human brain regions (anterior cingulate cortex, middle temporal gyrus and primary visual cortex) that evidenced high evolutionary constraint through enrichment for brain critical exons and high pLI genes. These clusters also showed significant enrichment with ASD loss-of-function variant genes (p < 5.23 × 10–11) that are transcriptionally highly active in prenatal brain regions (visual cortex and dorsolateral prefrontal cortex). Mapping ASD de novo LOF variant genes into large-scale human and mouse brain single-cell transcriptome analysis demonstrate enrichment of such genes into neuronal subtypes and are also enriched for subtype of non-neuronal glial cell types (astrocyte, p < 6.40 × 10–11, oligodendrocyte, p < 1.31 × 10–09). Conclusion Among the ASD genes enriched with pathogenic de novo LOF variants (i.e. KANK1, PLXNB1), a subgroup has restricted transcriptional regulation in non-neuronal cell types that are evolutionarily conserved. This association strongly suggests the involvement of subtype of non-neuronal glial cells in the pathogenesis of ASD and the need to explore other biological pathways for this disorder.

scholarly journals De Novo Prediction of Stem Cell Identity using Single-Cell Transcriptome Data

2016 ◽  
Vol 19 (2) ◽  
pp. 266-277 ◽  
Author(s):  
Dominic Grün ◽  
Mauro J. Muraro ◽  
Jean-Charles Boisset ◽  
Kay Wiebrands ◽  
Anna Lyubimova ◽  
...  
Keyword(s):  
Stem Cell ◽  
Single Cell ◽  
De Novo ◽  
Transcriptome Data ◽  
Cell Identity ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

scholarly journals Strategies to Identify Mesenchymal Stromal Cells in Minimally Manipulated Human Bone Marrow Aspirate Concentrate Lack Consensus

2021 ◽  
pp. 036354652199378
Author(s):  
Severin Ruoss ◽  
J. Todd Walker ◽  
Chanond A. Nasamran ◽  
Kathleen M. Fisch ◽  
Conner J. Paez ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Single Cell ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Bone Marrow Aspirate ◽  
Transcriptional Profile ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

Background: There is a need to identify and quantify mesenchymal stromal cells (MSCs) in human bone marrow aspirate concentrate (BMAC) source tissues, but current methods to do so were established in cultured cell populations. Given that surface marker and gene expression change in cultured cells, it is doubtful that these strategies are valid to quantify MSCs in fresh BMAC. Purpose: To establish the presence, quantity, and heterogeneity of BMAC-derived MSCs in minimally manipulated BMAC using currently available strategies. Study Design: Descriptive laboratory study. Methods: Five published strategies to identify MSCs were compared for suitability and efficiency to quantify clinical-grade BMAC-MSCs and cultured MSCs at the single cell transcriptome level on BMAC samples being used clinically from 15 orthopaedic patients and on 1 cultured MSC sample. Strategies included (1) the guidelines by the International Society for Cellular Therapy (ISCT), (2) CD271 expression, (3) the Ghazanfari et al transcriptional profile, (4) the Jia et al transcriptional profile, and (5) the Silva et al transcriptional profile. Results: ISCT guidelines did not identify any MSCs in BMAC at the transcriptional level and only 1 in 9 million cells at the protein level. Of 12,850 BMAC cells, 9 expressed the CD271 gene. Only 116 of 396 Ghazanfari genes were detected in BMAC, whereas no cells expressed all of them. No cells expressed all Jia genes, but 25 cells expressed at least 13 of 22. No cells expressed all Silva genes, but 19 cells expressed at least 8 of 23. Most importantly, the liberalized strategies tended to identify different cells and most of them clustered with immune cells. Conclusion: Currently available methods need to be liberalized to identify any MSCs in fresh human BMAC and lack consensus at the single cell transcriptome and protein expression levels. These different cells should be isolated and challenged to establish phenotypic differences. Clinical Relevance: This study demonstrated that improved strategies to quantify MSC concentrations in BMAC for clinical applications are urgently needed. Until then, injected minimally manipulated MSC doses should be reported as rough estimates or as unknown.

scholarly journals Single-cell RNA sequencing deconvolutes the in vivo heterogeneity of human bone marrow-derived mesenchymal stem cells

2020 ◽  
Author(s):  
Zun Wang ◽  
Xiaohua Li ◽  
Junxiao Yang ◽  
Yun Gong ◽  
Huixi Zhang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Single Cell ◽  
Rna Sequencing ◽  
Critical Role ◽  
Cellular Heterogeneity ◽  
Bone Homeostasis ◽  
Single Cell Rna Sequencing

AbstractBone marrow-derived mesenchymal stem cells (BM-MSCs) are multipotent stromal cells, which have a critical role in the maintenance of skeletal tissues such as bone, cartilage, and the fat found in bone marrow. In addition to providing microenvironmental support for hematopoietic processes, BM-MSCs can differentiate into various mesodermal lineages including osteoblast/osteocyte, chondrocyte, and adipocyte cells that are crucial for bone metabolism. While BM-MSCs have high cell-to-cell heterogeneity in gene expression, the cell subtypes that contribute to this heterogeneity in vivo in humans have not been characterized. To investigate the transcriptional diversity of BM-MSCs, we applied single-cell RNA sequencing (scRNA-seq) on freshly isolated CD271+ BM-derived mononuclear cells (BM-MNCs) from two human subjects. We successfully identified LEPRhiCD45low BM-MSCs within the CD271+ BM-MNC population, and further codified the BM-MSCs into distinct subpopulations corresponding to the osteogenic, chondrogenic, and adipogenic differentiation trajectories, as well as terminal-stage quiescent cells. Biological functional annotations of transcriptomes suggest that osteoblast precursors may induce angiogenesis coupled with osteogenesis, and chondrocyte precursors may have the potential to differentiate into myocytes. We discovered transcripts for several cluster of differentiation (CD) markers that were highly expressed (e.g., CD167b, CD91, CD130 and CD118) or absent (e.g., CD74, CD217, CD148 and CD68) in BM-MSCs and could be novel markers for human BM-MSC purification. This study is the first systematic in vivo dissection of human BM-MSCs cell subtypes at the single-cell resolution, revealing insight into the extent of their cellular heterogeneity and bone homeostasis.

scholarly journals Functional In vivo Single-cell Transcriptome (FIST) Analysis Reveals Molecular Properties of Light-Sensitive Neurons in Mouse V1

2018 ◽  
Author(s):  
Jianwei Liu ◽  
Na Pan ◽  
Le Sun ◽  
Mengdi Wang ◽  
Junjing Zhang ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Single Cell ◽  
Three Dimensional ◽  
Whole Cell ◽  
Mrna Sequencing ◽  
Layer 2 ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome ◽  
Live Mouse

ABSTRACTVision formation is classically based on projections from the retinal ganglion cells (RGC) to the lateral geniculate nucleus (LGN) and the primary visual cortex (V1). Although the cellular information of the retina and the LGN has been widely studied, the transcriptome profiles of single neurons with specific functions in V1 still remain unknown. Some neurons in mouse V1 are tuned to light stimulus. To determine the molecular properties of light-stimulated neurons in layer 2/3 of V1, we developed a method of functional in vivo single-cell transcriptome (FIST) analysis that integrates sensory evoked calcium imaging, whole-cell electrophysiological patch-clamp recordings, single-cell mRNA sequencing and three-dimensional morphological characterization in a live mouse, based on a two-photon microscope system. In our study, 58 individual cells from layer 2/3 of V1 were identified as either light-sensitive (LS) or non-light-sensitive (NS) by single-cell light-evoked calcium evaluation and action potential spiking. The contents of every single cell after individual functional tests were aspirated through the patch-clamp pipette for mRNA sequencing. Furthermore, the three-dimensional (3-D) morphological characterizations of the neurons were reconstructed in the live mouse after the whole-cell recordings. Our sequencing results indicated that V1 neurons with high expression of genes related to transmission regulation, such as Rtn4r, Nr4a1, and genes involved in membrane transport, such as Na+/K+ ATPase, NMDA-type glutamatergic receptor, preferentially respond to light stimulation. Our findings demonstrate the ability of FIST analysis to characterize the functional, morphological and transcriptomic properties of a single cell in alive animal, thereby providing precise neuronal information and predicting its network contribution in the brain.

scholarly journals Single-cell transcriptome reveals the redifferentiation trajectories of the early stage of de novo shoot regeneration in Arabidopsis thaliana

2022 ◽  
Author(s):  
Guangyu Liu ◽  
Jie Li ◽  
Jiming Li ◽  
Zhiyong Chen ◽  
Peisi Yuan ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Shoot Regeneration ◽  
De Novo ◽  
Developmental Process ◽  
Early Stage ◽  
Cell Populations ◽  
Cell Heterogeneity ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

De novo shoot regeneration from a callus plays a crucial role in both plant biotechnology and the fundamental research of plant cell totipotency. Recent studies have revealed many regulatory factors involved in this developmental process. However. our knowledge of the cell heterogeneity and cell fate transition during de novo shoot regeneration is still limited. Here, we performed time-series single-cell transcriptome experiments to reveal the cell heterogeneity and redifferentiation trajectories during the early stage of de novo shoot regeneration. Based on the single-cell transcriptome data of 35,669 cells at five-time points, we successfully determined seven major cell populations in this developmental process and reconstructed the redifferentiation trajectories. We found that all cell populations resembled root identities and undergone gradual cell-fate transitions. In detail, the totipotent callus cells differentiated into pluripotent QC-like cells and then gradually developed into less differentiated cells that have multiple root-like cell identities, such as pericycle-like cells. According to the reconstructed redifferentiation trajectories, we discovered that the canonical regeneration-related genes were dynamically expressed at certain stages of the redifferentiation process. Moreover, we also explored potential transcription factors and regulatory networks that might be involved in this process. The transcription factors detected at the initial stage, QC-like cells, and the end stage provided a valuable resource for future functional verifications. Overall, this dataset offers a unique glimpse into the early stages of de novo shoot regeneration, providing a foundation for a comprehensive analysis of the mechanism of de novo shoot regeneration.

Sign in / Sign up

Export Citation Format

Share Document