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Cell Stem Cell. 2018 Dec 6;23(6):787-793.e6. doi: 10.1016/j.stem.2018.11.016.

Human Intestinal Organoids Maintain Self-Renewal Capacity and Cellular Diversity in Niche-Inspired Culture Condition.

Author information

1
Department of Gastroenterology, Keio University School of Medicine, Tokyo 160-8582, Japan; Department of Organoid Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan; Department of Surgical Oncology, The University of Tokyo, Tokyo 113-8655, Japan.
2
Department of Gastroenterology, Keio University School of Medicine, Tokyo 160-8582, Japan; Department of Organoid Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan.
3
Department of Gastroenterology, Keio University School of Medicine, Tokyo 160-8582, Japan.
4
Department of Gastroenterology, Keio University School of Medicine, Tokyo 160-8582, Japan; Department of Organoid Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan; Fujii Memorial Research Institute, Otsuka Pharmaceutical Company, Ltd., Shiga 520-0106, Japan.
5
Department of Gastroenterology, Keio University School of Medicine, Tokyo 160-8582, Japan; Department of Organoid Medicine, Keio University School of Medicine, Tokyo 160-8582, Japan. Electronic address: t.sato@keio.jp.

Abstract

Cellular diversity that shapes tissue architecture and function is governed by multiple niche signals. Nonetheless, maintaining cellular diversity in human intestinal organoids has been challenging. Based on niche ligands present in the natural stem cell milieu, we establish a refined organoid culture condition for intestinal epithelia that allows human intestinal organoids to concurrently undergo multi-differentiation and self-renewal. High-throughput screening reveals that the combination of insulin-like growth factor 1 (IGF-1) and fibroblast growth factor 2 (FGF-2) enhances the clonogenic capacity and CRISPR-genome engineering efficiency of human intestinal stem cells. The combination equally enables long-term culture of a range of intestinal organoids, including rat small intestinal organoids. Droplet-based single-cell RNA sequencing further illustrates the conservation of the native cellular diversity in human small intestinal organoids cultured with the refined condition. The modified culture protocol outperforms the conventional method and offers a viable strategy for modeling human intestinal tissues and diseases in an in vivo relevant context.

KEYWORDS:

CRISPR-Cas9; LGR5; M cells; Paneth cells; diffusion pseudotime; enteroendocrine cells; intestinal stem cells; p38 signaling; tuft cells; ulcerative colitis

PMID:
30526881
DOI:
10.1016/j.stem.2018.11.016

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