A lymphatic-stem cell interactome regulates intestinal stem cell activity

Single-cell RNA-sequencing of murine small intestinal organoid cultured with and without lymphatic endothelial cells: Purpose: To identify differences in cell composition in murine organoids upon coculture with lymphatic endothelial cells Results: We recovered 818 cells with a median of 37750 UMIs per cell for the organoid alone and 1233 cells with a median of 79354 UMIs per cells for the cocultured organoid condition. Conclusions: scRNASeq reveals different cell composition in organoids cocultured with lymphatic endothelial cells compared to control Single-cell RNA-Sequencing of murine small and large intestinal tissue: Purpose: To generate a reference dataset of all major cell types present in the small and large intestine of the mouse Results: For the small intestine we recovered 8842 cells with a median of 4088 UMIs per cell compared to 8646 cells with a median of 10009 UMIs per cell for the large intestine. Conclusions: scRNASeq recovers all of the major epithelial, immune and stromal cell types in the intestine. Spatial transcriptomic profiling of murine small and large intestine: Purpose: To spatially map the major cell types in the mouse intestine and infer cell:cell communication from neighboring cells Conclusions: Spatial transcriptomic profiling of the mouse small and large intestine reveals all major cell types and allows for cell:cell signaling analyses upon integration with our matching scRNA-Seq data Overall design: 16 wells each of organoids alone and cocultured organoids were pooled to sort 100k epithelial cells from each. Ctrl and coculture sample were pooled and submitted for 10X single-cell RNA-Sequencing Bulk intestinal tissue was digested and we sorted 50k alive cells, enriching for LGR5+ intestinal stem cells (50k) and CD31+ PDPN+ LYVE1+ lymphatic endothelial cells (20-25k). Tissue processing and 10X-based single-cell RNA-sequencing was done separately for the small and large intestine. C57BL/6 and Lgr5-eGFP-ires-CreERT2 (C57BL/6 background) mice were used for sorting experiments. 10X-Visium-based spatial transcriptomics was done on mouse small and large intestine (technical duplicates for each). We used the immunofluorescence-based protocol on 10 µm cryosections of small and large intestinal tubes.