| Experiment Id | GSE172127 | Name | Spatial transcriptome profiling by MERFISH reveals fetal liver hematopoietic stem cell niche architecture [scRNA-seq] |
| Experiment Type | RNA-Seq | Study Type | Baseline |
| Source | GEO | Curation Date | 2024-02-09 |
| description | The hematopoietic stem cell (HSC) niche has been extensively studied in bone marrow, yet a more systematic investigation into the microenvironment regulation of hematopoiesis in fetal liver is necessary. Here we investigate the spatial organization and transcriptional profile of individual cells in both wild type and Tet2-/- fetal livers, by multiplexed error robust fluorescence in situ hybridization (MERFISH). We find that specific pairs of fetal liver cell types are preferentially positioned next to each other. Ligand-receptor singling molecule pairs such as Kitl and Kit are enriched in neighboring cell types. The majority of HSCs are directly in contact with endothelial cells (ECs) in both wild type and Tet2-/- fetal livers. Loss of Tet2 increases the number of HSCs, and upregulates Wnt and Notch signaling genes in the HSC niche. Two subtypes of ECs -- arterial ECs and sinusoidal ECs -- and other cell types contribute distinct signaling molecules to the HSC niche. Collectively, this study provides a comprehensive picture and bioinformatic foundation for HSC spatial regulation in fetal liver. Here we performed single-cell RNA sequencing of E14.5 whole fetal liver cells and E14.5 fetal liver HSCs by 10x Genomics platform. For whole fetal liver single cell RNA sequencing, fetal liver tissues were isolated from E14.5 C57BL/6 pregnant mice and digested by dissociation enzyme (Accutase) for 10 min at room temperature. Red blood cells were removed using 1xRBC lysis buffer at 4°C. Then, cells were passed through a 40 um strainer and re-suspended in cold DPBS with 0.04% (wt/vol) BSA. For sorted HSC single cell RNA sequencing, fetal liver tissues were isolated from E14.5 C57BL/6 pregnant mice and in cold 1xPBS dissociated into single cell suspension by pipetting up and down through 15 mL pipette. Red blood cells were removed using 1xRBC lysis buffer at room temperature for 5 min. Cells were incubated for 5 minutes at room temperature and mixed constantly by inverting the tube. After that, cells were centrifuged for 5 minutes then the supernatant was carefully removed without disturbing the pellet cells. Cells were resuspended in cold 1xPBS with 0.5% BSA, passed through a 40 um strainer. To sorting HSCs, we added Biotin Lineage depletion cocktail (anti-Mac1, anti-CD3e, anti-B220, anti-Gr-1, anti-CD11b and anti-Ter119) and streptavidin Particles Plus-DM befor sorting. Lineage positive cells were depleted by transferring cells to a magnetic stand. Enriched lineage negative cells were further stained with antibodies (LSK CD150+CD34-CD48-Flt3-). Then HSCs were sorted with FACS AriaII (BD Biosciences). Finally, we took 8,000 cells for 10x library preparation using Chromium Single Cell 3' Reagent Kits (v2) for each sample according to the manufacturer's description. |
