Rapid establishment of murine gastrointestinal organoids using mechanical isolation method

Gastrointestinal (GI) diseases, including pathological dysplasia, inflammation, neoplasia and injury, suffer millions of patients globally per year. Organoids, three-dimensional cell mass structures supported by biomaterials in dishes, are currently used as a research model for diseases of the small intestine. However, the traditional enzymatic-digestion method for establishing small-intestinal organoids (EnzyOs) is time consuming and often loses the bulk of crypts, a more efficient and reliable method needs to be developed. In this study, using mouse GI organoids as a model, we formulated a rapid mechanical isolation method that could efficiently isolate and culture villi-crypts into small intestinal organoids (MechOs). Primary duodenum organoids generated by MechOs displayed three types of morphology: spheroid, semi-budding and budding, while EnzyOs produced much less budding. Moreover, primary duodenum organoids from MechOs could be subcultured and presented similar gene expression profiles of small intestine specific markers as that from EnzyOs. Importantly, the MechOs method could also be used to generate other types of organoids derived from the stomach, jejunum-ileum, sigmoid-rectum and bile cysts. Taken together, the results show that MechOs could efficiently and economically generate digestive system organoids, providing a potential basis of epithelial organoids for the clinical treatment of gastroenterological diseases.