WT vs TNFAIP8 knockout enterocyte single cell RNA-Seq
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE143916 | GEO | 2020/04/20
REPOSITORIES: GEO
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Project description:The molecular mechanisms controlling stem cell renewal and lineage commitment are still poorly understood due to lack of reliable markers. In the adult small intestine, an example of high rate self-renewing tissue, four different epithelial cell lineages (enterocytes, Goblet, enteroendocrine and Paneth cells) are generated from a pool of stem cells localised at the bottom of the crypts of Lieberkühn. Recently, the orphan Leucine-rich repeat G protein-coupled receptor 5 (GPR49), a target of Wnt signalling, has been proposed as a marker for such cells. We investigated the role of LGR5 during intestinal development by using LGR5 null/LacZ-NeoR knock-in mice. We show that LGR5 deficiency leads to premature Paneth cell differentiation. X-gal staining on E18.5 intestine revealed that LGR5 expression is restricted to a few cells clustered within the intervillus region known to contain progenitor cells. In LGR5-null mice, expression from the LGR5 promoter was found upregulated, suggesting a loss of negative feedback control. However, neither epithelial cell proliferation nor cell migration appeared significantly impaired by LGR5 deficiency. Finally, transcriptional profiling of ileal mutant mice suggests that LGR5 plays a role in regulating the Wnt and Notch signalling pathways controlling progenitor renewal and differentiation.
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Project description:WT vs TNFAIP8 knockout enterocyte single cell RNA-Seq
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Project description:Although Lgr5+ intestinal stem cells have been expanded in vitro as organoids, homogeneous culture of these cells has not been possible so far. Here we show that two small molecules, CHIR99021 and valproic acid, synergistically maintain self-renewal of mouse Lgr5+ intestinal stem cells, resulting in nearly homogeneous cultures. The colony-forming efficiency of cells from these cultures is ~100-fold greater than that of cells cultured in the absence of CHIR99021 and valproic acid, and multilineage differentiation ability is preserved. We made use of these homogeneous cultures to identify conditions employing simultaneous modulation of Wnt and Notch signaling to direct lineage differentiation into mature enterocytes, goblet cells and Paneth cells. Expansion in these culture conditions may be feasible for Lgr5+ cells from the mouse stomach and colon and from the human small intestine. These methods provide new tools for the study and application of multiple intestinal epithelial cell types.
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