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In vitro maturation of human pluripotent stem cell-derived intestinal organoids

ABSTRACT: Human intestinal organoids (hIOs) derived from human pluripotent stem cells (hPSCs) have immense potential as a source of intestines. However, due to problems obtaining mature adult-like cells from hPSCs, effective strategies for the maturation of hIOs with improved functionalities must be developed. We established conditions that enable the in vitro maturation of hIOs derived from hPSCs, recapitulating the essential features of intestinal tissue. The maturity of the hPSC-derived hIOs was enhanced by the in vitro maturation in our co-culture system, evidenced by transcriptional changes observed in our microarray data. Global transcription of in vitro-matured hIOs is highly similar to that of adult human small intestine (hSI). Compared to control hIOs, in vitro-matured hIOs exhibited significant increases in the expression of intestinal maturation markers, major transporters, and key enzymes for drug absorption and first-pass metabolism.

ORGANISM(S): Homo sapiens

PROVIDER: GSE116738 | GEO | 2018/08/27

REPOSITORIES: GEO

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Project description:Genetically engineered human pluripotent stem cells (hPSCs) have been proposed as a source for transplantation therapies and are rapidly becoming valuable tools for human disease modeling. However, many of the potential applications are still limited by the lack of robust differentiation paradigms that allow for the isolation of defined functional tissues. These challenges could be overcome by the use of adult tissue stem cells derived from hPSCs, as their restricted potential could limit the differentiation towards other undesired linages, and allow in vitro expansion and long- term propagation of fully differentiated tissue. To isolate adult stem cells from hPSCs, we applied genome-editing to generate an LGR5-GFP reporter system and subsequently developed a differentiation protocol for human intestinal tissue comprising an adult stem cell niche and all major cell types of the adult intestine. This novel derivation protocol is highly robust and even permits the isolation of intestinal organoids without the LGR5 reporter. Transcriptional profiling, electron microscopy and functional analysis revealed that such human organoid cultures could be derived with high purity, and a composition and morphology similar to that of cultures obtained from human biopsies. Importantly, hPSC-derived organoids responded to the canonical signaling pathways that control self-renewal and differentiation in the adult human intestinal stem cell compartment. With our ability to genetically engineer hPSCs using site-specific nucleases, this adult stem cell system provides a novel platform by which to study human intestinal disease in vitro. RNA from primary organoid samples was isolated from organoid lines that were both cultured for 1-6 months and derived from duodenum, ileum, or rectum biopsies of human subjects as described previously (Sato et al., Gastroenterology 2011) grown in media called WENR+inhibitors. RNA was also isolated from various steps in the culturing and differentiation protocol.

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In vitro maturation of human pluripotent stem cell-derived intestinal organoids

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