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Leucine-rich repeat-containing G-protein coupled receptor 5 enriched organoids under chemically-defined growth conditions

Hahn, Soojung, Kim, Mi Sun, Choi, Seon Young, Jeong, Sukin, Jee, JooHyun, Kim, Han Kyung, Jeong, Sang Yun, Shin, Hyunsoo, Kim, Hyoung Sun, Park, Joon Seong, Yoo, Jongman
Biochemical and biophysical research communications 2019 v.508 no.2 pp. 430-439
G-protein coupled receptors, G-proteins, culture media, epidermal growth factor, epithelial cells, epithelium, intestines, mice, stem cells, transgenic animals, valproic acid
An organoid is a complex, multi-cell three-dimensional (3D) structure that contains tissue-specific cells. Epithelial stem cells, which are marked by leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5), have the potential for self-renewal and expansion as organoids. However, in the case of intestinal organoids from Lgr5-EGFP-IRES-CreERT2 transgenic mice, in vitro expansion of the Lgr5 expression is limited in a culture condition supplemented with essential proteins, such as epidermal growth factor (E), noggin (N), and R-spondin 1 (R). In this study, we hypothesized that self-renewal of Lgr5+ stem cells in a 3D culture system can be stimulated by defined compounds (CHIR99021, Valproic acid, Y-27632, and A83-01). Our results demonstrated that dissociated single cells from organoids were organized into a 3D structure in the four compounds containing the ENR culture medium in a 3D and two-dimensional (2D) culture system. Moreover, the Lgr5 expression level of organoids from the ENR- and compound-containing media increased. Furthermore, the conversion of cultured Lgr5+ stem cells from 2D to 3D was confirmed. Therefore, defined compounds promote the expansion of Lgr5+ stem cells in organoids.