Analysis of Cell Death Induction in Intestinal Organoids In Vitro.
The intestinal epithelium has an important function in the absorption of nutrients contained in the food. Furthermore, it also has an important barrier function, preventing luminal pathogens from entering the bloodstream. This single cell layer epithelium is quite sensitive to various cell death-promoting triggers, including drugs, irradiation, and TNF family members, leading to loss of barrier integrity, epithelial erosion, inflammation, malabsorption, and diarrhea. In order to assess the intestinal epithelium-damaging potential of treatments and substances specific test systems are required. As intestinal tumor cell lines are a poor substitute for primary intestinal epithelial cells, and in vivo experiments in mice are costly and often unethical, the use of intestinal organoids cultured from intestinal crypts provide an ideal tool to study cell death induction and mechanisms in primary intestinal epithelial cells. This protocol describes the isolation and culture of intestinal organoids from murine small intestinal crypts, and the quantitative assessment of cell death induction in these organoids.
Biochemical Pharmacology, University of Konstanz, Box 660, Universitätsstrasse 10, 78457, Konstanz, Germany.
Thomas
T
Grabinger
Biochemical Pharmacology, University of Konstanz, Box 660, Universitätsstrasse 10, 78457, Konstanz, Germany.
Eugenia
E
Delgado
Biochemical Pharmacology, University of Konstanz, Box 660, Universitätsstrasse 10, 78457, Konstanz, Germany. thomas.brunner@uni-konstanz.de.
Thomas
T
Brunner
D000072198:ADAM17 Protein / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D066298:In Vitro Techniques; D007413:Intestinal Mucosa / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D009940:Organoids / Q000473:pathology*
D000072198:ADAM17 Protein
Apoptosis; Chemotherapy; Crypts; Enteroids; Intestinal epithelial cells; Irradiation; MTT; Organoids; TNFα