Establishment of a refined culture method for rat colon organoids
Abstract
Background Methods for the artificial three-dimensional (3D) culture of mouse and human small-intestinal and large-intestinal stem cells have been established with CD24+ or Paneth cell niches. In contrast, no studies have established stable 3D culture for rat colon stem cells. In this study, we established an advanced method for efficient rat colonic stem cell culture.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 PGE 2 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.
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Two factors affect the differentiation of progenitor cells into organoids in collagen-based matrices: the source of seeded cells, and the spatial arrangement of collagen types around the cells.
“Two factors have been shown to affect the differentiation of progenitor cells into organoids in collagen-based matrices: the source of seeded cells, and the spatial arrangement of collagen types around the cells.”
From Towards organoid culture without Matrigel (2021) · cites this paper as [91]
The choice between the TMDU and Ootani methods should be governed by the source of cells used to generate the intestinal organoid.
“Isshiki and colleagues reported that the choice between the TMDU and Ootani methods should be governed by the source of cells used to generate the intestinal organoid.”
From Towards organoid culture without Matrigel (2021) · cites this paper as [91]
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Towards organoid culture without Matrigel
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- Authors
- Isshiki, H., Arimura, Y., Nagaishi, K., Kawakami, K., Onodera, K., Yamashita, K., Naishiro, Y., Fujimiya, M., Imai, K., & Shinomura, Y.
- Affiliations
- Sapporo Medical University (JP)
- Institute of Medical Science, The University of Tokyo (JP)
- Corresponding author
- Arimura, Yoshiaki
- Venue
- Biochemical and Biophysical Research Communications
- Publisher
- Elsevier BV
- Published
- 2017-07
- Volume
- 489
- Issue
- 3
- Pages
- 305-311
- ISSN
- 0006-291X (ISSN-L)
- OpenAlex
- W2619363316
- Cited by
- 5
- References
- 22
- Citations by year
- 2025: 1 · 2024: 1 · 2021: 1 · 2020: 1 · 2018: 1
- FWCI
- 0.24
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Biochemical and Biophysical Research Communications (publishedVersion)
- PubMed (publishedVersion)
- Funders
- JSPS KAKENHI (15K08972)
- Topics
- Health Sciences › Medicine › Oncology › Cancer Cells and Metastasis
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Digestive system and related health
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Wnt/β-catenin signaling in development and cancer
- Keywords
- colon organoids, 3D cell culture, organoid formation, proteomics, prostaglandin E2, galectin-4, Lewis rats
- MeSH terms
- Animals, Cells, Cultured, Colon, Organ Culture Techniques, Organoids, Stem Cells, Rats
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