ABSTRACT: To investigate spatial heterogeneities in the axolotl forebrain, a coronal section of it was obtained for spatial transcriptomics using Visium V1.
Project description:Induced pluripotent stem cell (iPSC) derived organoid systems provide models to study human organ development. Single-cell transcriptome sequencing enables highly-resolved descriptions of cell state heterogeneity within these systems and computational methods can reconstruct developmental trajectories. However, new approaches are needed to directly measure lineage relationships in these systems. Here we establish an inducible dual channel lineage recorder, iTracer, that couples reporter barcodes, inducible CRISPR/Cas9 scarring, and single-cell transcriptomics to analyze state and lineage relationships in iPSC-derived systems. This data set include the spatial iTracer data of three slices of one cerebral organoid measured by 10x Visium.
Project description:B cell-interacting reticular cells (BRC) form transcriptionally and topologically stable immune-interacting microenvironments that direct efficient humoral immunity. While several immune niche factors have been elucidated, the cues sustaining BRC function and topology across activation states remain unclear. Here, we employed spatial transcriptome analysis of murine ingunal and mesenteric lymph nodes to examine co-localization of distinct BRC subsets and immune cells complementing BRC-immune cell interaction analysis. Spatial analysis supported predicted feedforward BRC-immune cell circuits that sustain topologically-organized, functional niches across inflammatory states, lymphoid organs and species.
Project description:Aging is a major, yet unmodifiable, risk factor for cardiovascular disease, leading to vascular alterations, increased cardiac fibrosis, and inflammation, all of which contribute to impaired cardiac function. To investigate the spatial impact of aging, we applied an integrative approach combining single-nucleus RNA sequencing and spatial transcriptomics in 12-week-old and 18-month-old mice. We systematically mapped the aging heart and identified larger vessel-associated niches as key hotspots for activated macrophages and fibroblasts in aged hearts. These niches, surrounding arteries, were also enriched in senescent cells. Our findings suggest that the microenvironment around the vasculature is particularly susceptible to age-related changes and serves as a primary site for inflammation-driven aging so called \"inflammaging\". This study provides new insights into how aging reshapes cardiac cellular architecture, highlighting vessel-associated niches as potential therapeutic targets for age-related cardiac dysfunction.
Project description:We used Visium technology (10X Genomics) to infer cell-to-cell communication in ovarian and uterine tissue based on spatial proximity. Organs from 3-month mice in diestrus and 18-month old mice were collected and frozen in OCT. 10 µm thick tissue slices were placed on Visium Spatial Gene Expression Slides (10X Genomics) and stained with Hematoxylin and Eosin (H&E). Libraries were prepared by manufacturer’s recommendations and sequenced on NovaSeq6000. For samples that were sequenced in two runs, both sequencing runs were merged when running spaceranger (10X Genomics). Original nd2 microscopy images and results of scRNA-seq (linked datasets) and spatial transcriptomics analysis are available at Biostudies (S-BIAD482 and S-BSST852).
Project description:spatial transcriptomics by means of 10x Visium of first and second trimester human reproductive tract (including female and male internal and external genitalia)
Project description:Anorectal malformation (ARM), a common congenital anomaly of the digestive tract, is a result of insufficient elongation of the urorectal septum. The cytoplasmic protein Receptor of Activated C-Kinase 1 (Rack1) is involved in embryonic neural development; however, its role in embryonic digestive tract development and ARM formation is unexplored. Our study explored the hindgut development and cell death mechanisms in ARM-affected rats using spatial transcriptome analysis. We induced ARM in rats by administering ethylenethiourea via gavage on gestational day (GD) 10. On GDs 14-16, embryos from both normal and ARM groups underwent spatial transcriptome sequencing, which identified key genes and signalling pathways. Rack1 exhibited significant interactions among differentially expressed genes on GDs 15 and 16. Reduced Rack1 expression in the ARM-affected hindgut, verified by Rack1 silencing in intestinal epithelial cells, led to increased P38 phosphorylation and activation of the MAPK signalling pathway. The suppression of this pathway downregulated Nqo1 and Gpx4 expression, resulting in elevated intracellular levels of ferrous ions, reactive oxygen species (ROS) and lipid peroxides. Downregulation of Gpx4 expression in the ARM hindgut, coupled with Rack1 co-localisation and consistent mitochondrial morphology, indicated ferroptosis. In summary, Rack1, acting as a hub gene, modulates ferrous ions, lipid peroxides, and ROS via the P38-MAPK/Nqo1/Gpx4 axis. This modulation induces ferroptosis in intestinal epithelial cells, potentially influencing hindgut development during ARM onset.
Project description:Using Multiome and previously published sc/snRNA-seq data, we studied eight anatomical regions of the human heart including left and right ventricular free walls (LV and RV), left and right atria (LA and RA), left ventricular apex (AX), interventricular septum (SP), sino-atrial node (SAN) and atrioventricular node (AVN). For the first time, we profile the cells of the human cardiac conduction system, revealing their distinctive repertoire of ion channels, G-protein coupled receptors and cell-cell interactions. We map the identified cells to spatial transcriptomic data to discover cellular niches within the eight regions of the heart.
Project description:To study the spatial localisations of the cell populations in an early haematopoietic tissue and lymphoid organs critical for T and B cell development, we profiled fetal liver, thymus and spleen from 3 donors at 18 PCW with sequencing-based spatial transcriptomics (10x Genomics Visium).
Project description:Because of their central role in inflammatory processes pathological alterations in lymph nodes can affect innate and adaptive immunity. However, interpretation of lymph node pathology in humans is hampered by the lack of a reference under homeostatic conditions and a limited understanding of the molecular processes that occur during inflammatory activation. Using a combination of confocal microscopy, flow cytometry, and single cell and spatial transcriptome analysis, our study provides an in-depth characterization of the immunoanatomy of clinically inflammed and non-inflamed human lymph nodes.
Project description:We developed cell2location, a principled and versatile Bayesian model that is designed to resolve fine-grained cell types in spatial transcriptomic data and create comprehensive cellular maps of diverse tissues. To validate cell2location in real tissue, we applied the model to data from the mouse brain, which features diverse neural cell types organised in a well characterised spatial architecture across brain areas, thus presenting a canonical use case to test spatial genomics. We generated matched single nucleus (sn, this submission) and Visium spatial RNA-seq (10X Genomics) profiles of adjacent mouse brain sections that contain multiple regions from the telencephalon and diencephalon. To assess the biological and intra-organ technical variation in spatial mapping, we assayed two mouse brains and serial tissue sections from each brain (total of 3 and 2 matched sections from two animals, respectively, and an extra section for snRNA-seq), creating a rich multi-modal and replicated transcriptomic dataset. Tissue processing. Brains of wild-type adult C57BL/6 mice (postnatal day 56, 1 female and 1 male) were dissected, snap frozen, embedded in optimal cutting temperature compound (Tissue-Tek) and stored at -80oC. Brain hemispheres were cryosectioned at -20oC using a cryostat (Leica, CM3050S). To assess tissue quality, RNA was extracted from test tissue sections using the RNeasy Pico Kit (Qiagen) and yielded high RIN values (9.6 and 9.7) on an Agilent Bioanalyser, indicating high RNA quality. For matched single nuclei and Visium RNA-seq experiments, brain hemispheres were cryosectioned to adjacent thick (200 µm) and thin (10 µm) coronal sections, respectively, and processed the same day. In total, four consecutive sets of thick and thin tissue sections were collected from each brain. Five sets of tissue sections yielded both good quality single nuclei and Visium data (three adjacent sections from mouse 1 and two sections from mouse 2) while one additional section from mouse 2 yielded good single nuclei; these were considered for analysis in this study. Visium spatial transcriptomics. Thin (10 µm) mouse brain sections were cryosectioned and mounted directly onto separate capture areas on 10X Visium Spatial Gene Expression slides (beta product version). Processing was done per manufacturer’s protocols. Briefly, sections were methanol-fixed, hematoxylin and eosin (H&E)-stained, and imaged on a NanoZoomer 2.0 slide scanner (Hamamatsu). Sections were then permeabilized and further processed to obtain cDNA libraries that were quality controlled using the Agilent Bioanalyser. The cDNA libraries were sequenced on the Illumina HiSeq 4000 system, aiming at 300 million raw reads per section with read lengths 28cy R1, 8cy i7 index, 0cy i5 index, 91cy read 2. 10X Visium spatial sequencing data was aligned to mouse pre-mRNA genome reference version mm10 using 10X SpaceRanger and mRNA count matrices were generated by adding intronic and exonic reads for each gene in each location. The paired histology H&E images were processed using 10X SpaceRanger to select locations covered by tissue by aligning pre-recorded spot locations with fiducial border spots in the histology image. This allows evaluating the correspondence between cell maps produced using our method and the known brain anatomy. This also allows identifying the number of nuclei in each spot using nuclear segmentation as described in Suppl. Methods and reported in Fig S8A-D. The histology image was used to manually annotate cortical layers in the primary somatosensory cortex (SSp) region using the lasso tool in the 10X Loupe browser.