Stem Cell Reports. 2017 Dec 1. pii: S2213-6711(17)30490-3. doi: 10.1016/j.stemcr.2017.11.004. [Epub ahead of print]

A Refined Culture System for Human Induced Pluripotent Stem Cell-Derived Intestinal Epithelial Organoids.

Takahashi Y¹, Sato S², Kurashima Y³, Yamamoto T⁴, Kurokawa S³, Yuki Y³, Takemura N⁵, Uematsu S⁵, Lai CY⁶, Otsu M⁶, Matsuno H⁷, Osawa H⁷, Mizushima T⁷, Nishimura J⁷, Hayashi M⁸, Yamaguchi T⁸, Kiyono H³.

Abstract

Gut epithelial organoids are routinely used to investigate intestinal biology; however, current culture methods are not amenable to genetic manipulation, and it is difficult to generate sufficient numbers for high-throughput studies. Here, we present an improved culture system of human induced pluripotent stem cell (iPSC)-derived intestinal organoids involving four methodological advances. (1) We adopted a lentiviral vector to readily establish and optimize conditioned medium for human intestinal organoid culture. (2) We obtained intestinal organoids from human iPSCs more efficiently by supplementing WNT3A and fibroblast growth factor 2 to induce differentiation into definitive endoderm. (3) Using 2D culture, followed by re-establishment of organoids, we achieved an efficient transduction of exogenous genes in organoids. (4) We investigated suspension organoid culture without scaffolds for easier harvesting and assays. These techniques enable us to develop, maintain, and expand intestinal organoids readily and quickly at low cost, facilitating high-throughput screening of pathogenic factors and candidate treatments for gastrointestinal diseases.