Organoid Culture Model for the Study of Adult Slow-Cycling Putative Intestinal Stem Cells

The study of intestinal stem cells is crucial for developing therapies for gut regeneration. Using Tet-Op histone 2 B (H2B)-green fluorescent protein (GFP) fusion protein transgenic mouse, we have developed a three-dimensional small intestinal (SI) organoid culture model in which cells with different cell cycle durations can be distinguished, allowing real-time visualization of the kinetics of slow-cycling H2B-GFP-retaining putative SI epithelial stem cells. The model allows real-time monitoring of the behavior of H2B-GFP-retaining putative SI stem cells under basal conditions, of their response to manipulations aimed at studying their regulation, and further functional characterization after isolation from SI organoids.