single cell rna sequence scrna seq analysis  (10X Genomics)

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Complement C3d enables cell-mediated immunity capable of distinguishing spontaneously transformed from nontransformed cells.

doi: 10.1073/pnas.2405824121

Figure Lengend Snippet: Fig. 3. C3d decreases malignant plasma cells sparing normal plasma cells. Vk*myc mice were injected with 20 microg C3d peptide (N = 6) or with PBS (N = 5) intratibia 2 mo before analysis. Malignancy and nonmalignancy associated gene expression was analyzed in 3,995 individual plasma cells obtained from the bone marrow. (A) UMAP plots of sc RNA gene expression depicting the different cell populations analyzed. (B) Graph depicts the differential expression (DE) of a set of malignancy associated or nonmalignancy associated genes in the upper and lower percentile 20 for each category and in each group of mice. Each horizontal line represents a gene, each vertical line represents a cell. (C) Graphs show PCA of the expression of selected genes in the upper and lower percentile 20 associated with malignancy or nonmalignancy in all the plasma cells analyzed from both C3d-treated and untreated mice. The genes chosen contributed most of the difference between malignant and nonmalignant cells. (D) Distribution of clones according to a compound score obtained by subtracting the malignant score value from the nonmalignant score value for each cell. Upper graph depicts a histogram with bins separated by 0.2 compound score units. Frequencies (Y-axis) are shown as a percent of the clones (obtained from C3d treated or controls) included in the analysis. Lower graph is a scatter plot of clones according to their compound score. Statistical analysis by the Mann–Whitney test yielded P < 0/0001, (****). (E) Two-dimensional plots depicting malignant (X- axis) and nonmalignant (Y-axis) gene expression scores from treated and untreated mice. The size and color of the circles represent the number of copies of each clonotype defined according to the CDR3 sequence, VH and JH identity.

Article Snippet: To probe C3d-induced CMI on normal and malignant plasma cells we used single cell RNA sequencing (scRNA seq) analysis to compare bone marrow cells from Vκ*-MYC mice treated with C3d with bone marrow cells from Vκ*-MYC mice that had not received C3d, using 10X genomics technologies.

Techniques: Clinical Proteomics, Injection, Gene Expression, Quantitative Proteomics, Expressing, Clone Assay, MANN-WHITNEY, Sequencing

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A-D . Anatomy of the heart and pericardium in zebrafish embryos and early larvae. A,B . Lateral confocal imaging of transgenic hand2:EGFP;myl7:DsRed zebrafish, anterior to the left. Embryo showing hand2:EGFP and myl7:DsRed (myocardium marker) co-expression in the heart tube and hand2:EGFP- expressing cell populations in the pericardium, posterior mesothelium, pectoral fin, and pharyngeal arches at 48 hpf ( A , 10x; B , 20x). C,D . Ventral confocal imaging ( C 72 hpf single Z-slice, D 96 hpf max projection, anterior to the top) of hand2:EGFP;myl7:DsRed embryo showing reporter co-expression in the atrium and ventricle myocardium and hand2:EGFP- expressing cells in the pharyngeal arches and pericardial sac surrounding the heart ( C ), with the pericardium acquiring a mesh-like squamous epithelial structure at 96 hpf ( D ). E . SPIM-based Mercator projection of double-transgenic hand2:EGFP;drl:mCherry embryo at 14 hpf, anterior to the top, showing the anterior-to-posterior extent of lateral-most hand2:EGFP -expressing LPM cells fated as mesothelial progenitors (mp), with the anterior-most extent of the bilateral stripes indicated by white arrowheads. Dotted box outlines region of interest for subsequent imaging panels. F-H . Dorsal confocal imaging of transgenic reporter combinations at the level of the heart field, anterior to the top, asterisks showing the midline where the heart field converges, arrowheads at lateral mesh-like cells. F . hand2:EGFP;drl:mCherry embryo at 16 hpf prior to medial heart field migration, showing hand2:EGFP -expressing cell populations comprising lateral-most LPM. G ) hand2:EGFP;nkx2 . 5:ZsYellow embryo with dual-marked emerging cardiac disk. H . hand2:EGFP;tbx1:mCerulean embryo with similar co-expression across bilateral mesh-like cells and the cardiac disk. I . SPIM-based live imaging stills from Movie 1 depicting a hand2:EGFP;tbx1:mCerulean embryo from 19-24 hpf, anterior to the top, showing hand2:EGFP- expressing lateral-most cells migrating away from the midline towards the front (migration front, yellow dashed line), while cardiac precursors form the heart tube towards the midline (white dashed line). White arrows for migration direction. pa, pharyngeal arches; pc, pericardium; ht, heart; v, ventricle; a, atrium; hg, hatching gland. Scale bars A-D 100 μm, E-H 200 μm, I 50 μm.

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques: Imaging, Transgenic Assay, Marker, Expressing, Migration

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A . Representative confocal max projection of drl:mCherry ; hand2:EGFP double-transgenic embryo at 10 hpf as used for FACS-based isolation of post-gastrulation LPM for 10xGenomics-based single cell transcriptomics; anterior-posterior axis as indicated. B . UMAP plot of single cell transcriptomes of mCherry-sorted 10 hpf drl:mCherry ; hand2:EGFP zebrafish embryo cells showing 18 significant cell clusters, colored by identified subpopulation. C,D . Whole-mount mRNA in situ hybridization of representative pericardial/mesothelial genes sfrp5 ( C ) and jam2b ( D ), white dashed outlines to emphasize mesh-like spread of the expression domains lateral to the embryo axis; anterior to the top. E,F . UMAP plots of key myocardial ( E ) and pericardial genes ( F ) expressed across identified cluster identities. Cell representations are colored by scaled expression values using lower and upper 2%-quantiles as boundaries. G . Clustering analysis of bulk mRNA-sequenced left ventricle myocardium (Myo) and pericardium (Per) from neonatal rats for genes defining myocardium versus pericardium as identified from the zebrafish-based scRNA-seq analysis in B . Heatmap bins colored by row-scaled log2-normalized counts; columns (samples) split by tissue type; rows and columns ordered by hierarchical clustering (scaled expression values), sex of sample indicated on top. Scale bar A,B 200 μm.

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques: Transgenic Assay, Isolation, Single-cell Transcriptomics, In Situ Hybridization, Expressing

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A . Dotplot including key cell fate marker genes to annotate broad mesothelial, pericardial, and myocardial clusters, respectively. Dots colored by column-scaled mean expression (log-transformed library-size-normalized counts) and sized by expression frequency (fraction of cells with non-zero counts). B . Dorsal confocal imaging of representative dual-transgenic hand2:EGFP;nkx2 . 5:ZsYellow zebrafish embryo at 16 hpf showing marker co-expression and heterogeneity in prospective cardiac and pericardial progenitor cells around the heart field; anterior to the top. C-E Slingshot-based trajectory inference analysis of early LPM cells assigned using key marker genes to the public Zebrahub data set of single cell transcriptomes throughout zebrafish development. Inferred end points for myocardium and pericardium indicated as color-coded clusters ( C ). PCA plots of key myocardial ( D ) and pericardial genes ( E ) expressed across identities over time. Cells are colored by scaled expression values using lower and upper 2%-quantiles as boundaries. Scale bar B 200 μm.

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques: Marker, Expressing, Transformation Assay, Imaging, Transgenic Assay

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A . UMAP projection of zebrafish cells in Zebrahub dataset, colored by anatomical ontology classes, highlighting the diversity of cell types within the embryo. B . UMAP projection of zebrafish embryonic cells with pericardium cells highlighted in red, showing their distribution across different clusters. C . UMAP projections highlighting specific cell types within the zebrafish embryo: pericardium cells, pectoral fin cells, floor plate cells, pharyngeal arch 1 cells, pharyngeal arch 2 cells, and myocardium cells, each marked in red to show their spatial organization within the clusters. D . UMAP projections of zebrafish embryonic cells, with cells colored by cluster (left) and by time point (right), demonstrating the developmental stages and the organization of clusters over time. E . UMAP projection of a subset of clusters identified in zebrafish embryonic cells, highlighting the specific lineage groups: undifferentiated lateral plate mesoderm (LPM) at different hours post-fertilization (hpf), pericardium, and myocardium cells. F . Principal component analysis (PCA) of zebrafish embryonic cells, showing the distribution of cells over time (time point). G . PCA projection of the subset clusters identified in F , demonstrating the developmental trajectory of these specific lineages over time. See also .

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques:

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A . UMAP projections of representative genes found among LPM, myocardial, and pericardial clusters from zebrafish embryonic cells across time as key cell fate markers used to annotate clusters. Cells are colored by scaled expression values using top and bottom 2%-quantiles as boundaries. B . PCA projections of LPM, myocardial, and pericardial clusters from zebrafish embryonic cells across time showing key cell fate markers used to annotate clusters. Cells are colored by scaled expression values using top and bottom 2%-quantiles as boundaries. See also .

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques: Expressing

Journal: bioRxiv

Article Title: The pericardium forms as a distinct structure during heart formation

doi: 10.1101/2024.09.18.613484

Figure Lengend Snippet: A,B . Metascape analysis of top-50 cluster-defining genes from early LPM scRNA-seq; gene ontology terms enriched in myocardial cluster ( A ), and pericardial cluster ( B ); note regulation of canonical Wnt signaling as significant in pericardial cells. C-E . Confocal max projections of representative transgene expression for hand2:EGFP (green) and 7xTCF:mCherry (magenta, broadly reading out canonical Wnt signaling activity) with colocalization (white overlay), embryonic axis as indicated. At 10 hpf, canonical Wnt signaling shows a graded activity from the posterior towards the anterior ( C , lateral view, asterisk). At the start of heart field convergence, various anterior LPM cells show heterogeneous canonical Wnt activity, with lateral-most putative pericardial progenitors strongly expressing the 7xTCF:mCherry reporter ( D , dorsal view, arrowheads). This pattern continues throughout to 24 hpf ( E , lateral view, white arrowheads). F,G . Expression of canonical Wnt signaling-associated genes across myocardial and pericardial/mesothelial cells at tail bud stage in zebrafish. Cropped UMAP plot to depict cell clusters of interest ( F ) and individual Wnt signaling-associated genes, with expressing cells colored by scaled expression values using lower and upper 2%-quantiles as boundaries ( G ). Scale bar C-E 200 μm.

Article Snippet: We conducted a 10xGenomics-based single-cell RNA-sequencing (scRNA-seq) analysis of zebrafish LPM cells at 10 hpf (tailbud stage, end of gastrulation) when the LPM has coalesced into its bilateral stripe architecture ( ).

Techniques: Expressing, Activity Assay