Induced Quiescence of Lgr5+ Stem Cells in Intestinal Organoids Enables Differentiation of Hormone-Producing Enteroendocrine Cells

Cell Stem Cell. 2017 Feb 2;20(2):177-190.e4. doi: 10.1016/j.stem.2016.11.001. Epub 2016 Dec 8.


Lgr5+ adult intestinal stem cells are highly proliferative throughout life. Single Lgr5+ stem cells can be cultured into three-dimensional organoids containing all intestinal epithelial cell types at near-normal ratios. Conditions to generate the main cell types (enterocyte, goblet cells, Paneth cells, and M cells) are well established, but signals to induce the spectrum of hormone-producing enteroendocrine cells (EECs) have remained elusive. Here, we induce Lgr5+ stem cell quiescence in vitro by blocking epidermal growth factor receptor (EGFR) or mitogen-associated protein kinase (MAPK) signaling pathways in organoids and show that their quiescent state is readily reverted. Quiescent Lgr5+ stem cells acquire a distinct molecular signature biased toward EEC differentiation. Indeed, combined inhibition of Wnt, Notch, and MAPK pathways efficiently generates a diversity of EEC hormone-expressing subtypes in vitro. Our observations uncouple Wnt-dependent stem cell maintenance from EGF-dependent proliferation and provide an approach for the study of the elusive EECs in a defined environment.

Keywords: EGFR signaling; Lgr5; enteroendocrine cell; intestinal stem cells; organoids; quiescence.

MeSH terms

  • Animals
  • Cell Cycle*
  • Cell Differentiation*
  • Cell Lineage
  • Cell Proliferation
  • Cells, Cultured
  • Enteroendocrine Cells / cytology*
  • ErbB Receptors / antagonists & inhibitors
  • ErbB Receptors / metabolism
  • Gene Expression Profiling
  • Hormones / biosynthesis*
  • Intestines / cytology*
  • MAP Kinase Signaling System
  • Mice
  • Organoids / cytology*
  • Receptors, G-Protein-Coupled / metabolism*
  • Receptors, Notch / metabolism
  • Sequence Analysis, RNA
  • Signal Transduction
  • Single-Cell Analysis
  • Stem Cells / cytology*
  • Wnt Proteins / metabolism


  • Hormones
  • Lgr5 protein, mouse
  • Receptors, G-Protein-Coupled
  • Receptors, Notch
  • Wnt Proteins
  • ErbB Receptors