Establishment of a long-term three-dimensional primary culture of mouse glandular stomach epithelial cells within the stem cell niche

• Published in: Biochemical and biophysical research communications Vol. 432; no. 4; pp. 558 - 563
• Main Authors: Katano, Takahito, Ootani, Akifumi, Mizoshita, Tsutomu, Tanida, Satoshi, Tsukamoto, Hironobu, Ozeki, Keiji, Ebi, Masahide, Mori, Yoshinori, Kataoka, Hiromi, Kamiya, Takeshi, Toda, Shuji, Joh, Takashi
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 22.03.2013
• Subjects: 60 APPLIED LIFE SCIENCES; Animals; CELL CULTURES; Cell Differentiation; COLLAGEN; CYTOPLASM; Epithelial Cells - cytology; Epithelial Cells - metabolism; Gastric Mucosa - cytology; Gastric Mucosa - metabolism; GLYCOGEN; IN VITRO; IN VIVO; LARGE INTESTINE; MICE; Mucin 5AC - metabolism; Primary Cell Culture; SMALL INTESTINE; Stem cell; Stem Cell Niche; STEM CELLS; STOMACH; Three-dimensional primary culture; Stomach; Three-dimensional primary culture; Stem cell
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2013.02.051

► We established a 3D culture system to allow long-term culture of stomach cells. ► In this culture system, gastric epithelial cells grew for about 3months. ► The cultured cells differentiated into multi-units of the stomach. ► This culture method should be useful for elucidating the cause of gastric diseases. Compared to the small intestine and colon, little is known about stem cells in the stomach because of a lack of specific stem cell markers and an in vitro system that allows long-term culture. Here we describe a long-term three-dimensional (3D) primary gastric culture system within the stem cell niche. Glandular stomach cells from neonatal mice cultured in collagen gel yielded expanding sphere-like structures for 3months. The wall of the gastrospheres consisted of a highly polarized epithelial monolayer with an outer lining of myofibroblasts. The epithelial cells showed a tall columnar cell shape, basal round nuclei, and mucus-filled cytoplasm as well as expression of MUC5AC, indicating differentiation into gastric surface mucous cells. These cells demonstrated the features of fully differentiated gastric surface mucous cells such as microvilli, junctional complexes, and glycogen and secretory granules. Fewer than 1% of cultured epithelial cells differentiated into enteroendocrine cells. Active proliferation of the epithelial cells and many apoptotic cells in the inner lumen revealed the rapid cell turnover in gastrospheres in vitro. This method enables us to investigate the role of signaling between cell–cell and epithelial–mesenchymal interactions in an environment that is extremely similar to the in vivo environment.