Three-Dimensional Gastrointestinal Organoid Culture in Combination with Nerves or Fibroblasts: A Method to Characterize the Gastrointestinal Stem Cell Niche

Figure 7

Reconstruction of stem cell niche in the intestine in vitro and proposed workflow. Intestinal crypts and stromal cells (neurons and/or myofibroblasts) are isolated and combined together in three-dimensional system such as organoid culture. Role of stromal cells as stem cell niche can be analyzed, for example, by the measurement of organoid diameter, cell viability assay, clonogenicity assay, gene expression analysis, and different types of staining. Identification of stem cell markers and individual cell types can be analyzed by FACS. In addition, genetic manipulations in organoids can be performed by lentiviral transfection and CRISPR-CASP9 system.