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Abstract: FR-OR041

High Throughput Single Cell RNA Sequencing Reveals a Roadmap to Recreate the Kidney

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 501 Development, Stem Cells, and Regenerative Medicine: Basic


  • Combes, Alexander N., University of Melbourne, Parkville, Victoria, Australia
  • Phipson, Belinda, Murdoch Children''s Research Institute, Parkville, New South Wales, Australia
  • Lawlor, Kynan T., Murdoch Children's Research Institute, Melbourne, Victoria, Australia
  • Zappia, Luke, Murdoch Children''s Research Institute, Parkville, New South Wales, Australia
  • Oshlack, Alicia, Murdoch Children's Research Institute, Melbourne, Victoria, Australia
  • Little, Melissa H., Murdoch Children's Research Institute, Melbourne, Victoria, Australia

The developing mouse kidney represents a valuable model to understand the formation and maturation of renal cell types. High throughput single cell RNA sequencing offers new opportunities to understand the formation of complex tissues.


We performed single cell RNA-seq on over 6000 cells (median 2896 genes per cell) from E18.5 kidneys on the Chromium 10x platform. Seurat was used for normalisation and clustering of cells, edgeR to test for differentially expressed genes. Findings were investigated using immunofluorescence and lineage tracing.


We used single cell profiling to define cell type-specific profiles and revisit the molecular regulation of kidney development in mouse. A detailed analysis of cellular heterogeneity in the nephron lineage identified multiple Six2+ subpopulations including a nephron progenitor ‘ground state’, a putative committing state with evidence of FGF suppression and Notch activation, and cells expressing markers of both nephron progenitor and stroma. In vivo lineage analysis from E12.5 with inducible Six2CreERT2 and PdgfraCreER lines supported a rare transition from nephron progenitor to stroma, but not the other way. Several markers thought to be specific to one population were expressed more broadly. For example Hoxb7 was expressed in collecting duct, distal tubule, and in some endothelial cells. Likewise, most markers considered specific to the ureteric epithelium were also expressed in the distal tubule or connecting segment. Comparative analysis between these populations yielded unique markers for each, which should be useful for distinguishing between these cell types in kidney organoids. An analysis of signaling pathway component expression was performed for each lineage, identifying known and previously unassociated ligands and receptors with progenitor states and stages of maturation in each lineage.


This study offers new insight into the mechanisms of progenitor maintenance and differentiation in the stroma, the ureteric epithelium, and the nephron lineages and therefore provides a roadmap of the signals that regulate differentiation in vivo. This information can be used to refine strategies to direct differentiation of mature renal cell types in vitro.


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