Conclusions
We established efficient rat colonic stem cell cultures in vitro. This success will contribute to the study of rat intestinal-disease models.
Methods
Using various tissue homogenates, we investigated the colonic organoid forming capacity under the TMDU protocol immediately adjacent to Ootani's 3D culture assembly in the same culture dish. Next, we examined whether the supernatant from the colon could be replaced by a colon homogenate. Finally, we identified the bioactive substances that were indispensable for efficient organoid culture using protein purification by three-step column chromatography and proteomic analysis with a quantitative nanoflow liquid chromatography-tandem mass spectrometry (LC-MS/MS).
Results
By combining Ootani's method with the TMDU protocol, we established a refined culture method for Lewis rat colon organoids, which we refer to as the modified TMDU protocol. Furthermore, we confirmed that PGE2 and galection-4 promoted rat colonic organoid formation. Conclusions: We established efficient rat colonic stem cell cultures in vitro. This success will contribute to the study of rat intestinal-disease models.