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

Takahashi, Yu; Sato, Shintaro; Kurashima, Yosuke; Yamamoto, Tomohisa; Kurokawa, Shiho; Yuki, Yoshikazu; Takemura, Nobuyuki; Uematsu, Satoshi; Lai, Chen-Yi; Otsu, Makoto; Matsuo, Hiroshi; Osawa, Hideki; Mizushima, Tsunekazu; Nishimura, Junichi; Hayashi, Mikio; Yamaguchi, Takayuki; Kiyono, Hiroshi

doi:10.1016/j.stemcr.2017.11.004

Cited by 86 publications

(98 citation statements)

References 47 publications

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“…Finally, we attempt to establish a maintenance culture procedure for a prolonged period. A long-term culture procedure is useful for drug screening 31 . When hiPSC-IECs were embedded in Matrigel supplemented with media for 8 weeks, they proliferated, formed organoids and expressed the Villin gene.…”

Section: Discussionmentioning

confidence: 99%

Generation of intestinal organoids derived from human pluripotent stem cells for drug testing

Yoshida

Miwa

Kawachi

et al. 2020

Sci Rep

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Drug absorption via the intestinal tissue is modulated by membrane permeability and metabolism in intestinal epithelial cells (IECs). In drug discovery research, using human IECs to evaluate membrane permeability and metabolic stability can offer very useful information when exploring for drug candidate compounds that have good bioavailability and when trying to predict the fraction absorbed and intestinal availability in humans. Here, we evaluated the pharmacokinetic functions of human IECs differentiated from human induced pluripotent stem cells (hiPSCs) in 3D cultures. As human IECs differentiated in 3D cultures form intestinal organoids and spheroids (herein termed organoids), their morphology makes it difficult to evaluate their pharmacokinetic functions. Therefore, we dissociated intestinal organoids into single cells and attempted to purify human IECs. We found that hiPSC-derived IECs (hiPSC-IECs) expressed the epithelial cell adhesion molecule (EpCAM) and could be highly purified by sorting EpCAM+ cells. The hiPSC-IEC monolayer showed a high TEER value (approximately 350 Ω × cm 2). In addition, hiPSC-IECs oxidatively metabolized terfenadine (CYP3A and CYP2J2 substrate) and midazolam (CYP3A substrate). These results indicated that hiPSC-IECs form tight-junction and have cytochrome P450 enzymatic activities. In conclusion, we developed a novel application of hiPSCderived intestinal organoids for drug testing.

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Section: Discussionmentioning

confidence: 99%

Generation of intestinal organoids derived from human pluripotent stem cells for drug testing

Yoshida

Miwa

Kawachi

et al. 2020

Sci Rep

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“…Intestinal epithelial stem cells can be obtained from intestinal biopsies or resection and, once isolated, can be maintained in vitro under defined culture conditions where they undergo functional differentiation and derived crypt-villus organoid structures [22, 23]. Moreover, intestinal epithelial cells can also be derived from induced pluripotent stem cells (iPSC) [24]. …”

Section: Discussionmentioning

confidence: 99%

Isolation and characterisation of mouse intestinal mesoangioblasts

et al. 2018

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Aims and objectivesChildren suffering from intestinal failure (IF) endure considerable morbidity and overall have poor survival rates, complicated by the shortage of organs available for transplantation. Therefore, new therapeutic approaches are pivotal if outcomes are to be improved. Over the past years, tissue engineering (TE) has emerged as a possible alternative treatment for many congenital and acquired conditions. TE aims at creating bioengineered organs by means of combining scaffolds with appropriate cell types, which in the intestine are organised within a multilayer structure. In order to generate functional intestine, this cellular diversity and organisation will need to be recreated. While the cells for the epithelial, neural and vascular compartments have been well defined, so far, less attention has been put on the muscular compartment. More recently, mesoangioblasts (MABs) have been identified as a novel source for tissue regeneration since they are able to give rise to vascular and other mesodermal derivatives. To date MABs have not been successfully isolated from intestinal tissue. Therefore, our aim was to demonstrate the possibility of isolating MABs from adult mouse small intestine.Materials and methodsAll experiments were carried out using small intestinal tissues from C57BL/6J mice. We applied an established protocol for MAB isolation from the isolated neuromuscular layer of the small intestine. Cultured cells were stained for Ki67 to assess proliferation rates as well as for a panel of pericyte markers to determine their phenotype.ResultsCells were successfully isolated from gut biopsies. Cultured cells showed good proliferative capacity and positivity for at least three pericytes markers found in vessels of the gut neuromuscular wall: neuron-glial antigen 2, alkaline phosphatase and platelet-derived growth factor β.ConclusionThis proof-of-principle study lays the foundation for further characterization of MABs as a possible cell source for intestinal smooth muscle regeneration and TE.

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“…The use of intestinal stem cells has been more recently explored in the development of intestinal organoids and cell structures which are able to recapitulate their 3D organization naturally occurring in vivo . However, when culturing organoids, the use of an extracellular matrix (e.g., Matrigel or collagen) is fundamental for their adhesion and growth.…”

Section: In Vitro Cell Culture Modelsmentioning

confidence: 99%

Models of the Gut for Analyzing the Impact of Food and Drugs

Fois

Schindeler

et al. 2019

Adv Healthcare Materials

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The human intestine is the primary organ responsible for the uptake of nutrients and water, and this is facilitated by its complex structure that features a large surface area. With an average total length of around seven meters, including both small and large intestine, it connects the stomach to the rectum while enabling absorption in a specialized manner. At a cellular level, the gut epithelium is critical in selective transport to the bloodstream. The finger-like projections of the intestinal epithelium Models of the human gastrointestinal tract (GIT) can be powerful tools for examining the biological interactions of food products and pharmaceuticals. This can be done under normal healthy conditions or using models of disease-many of which have no curative therapy. This report outlines the field of gastrointestinal modeling, with a particular focus on the intestine. Traditional in vivo animal models are compared to a range of in vitro models. In vitro systems are elaborated over time, recently culminating with microfluidic intestines-on-chips (IsOC) and 3D bioengineered models. Macroscale models are also reviewed for their important contribution in the microbiota studies. Lastly, it is discussed how in silico approaches may have utility in predicting and interpreting experimental data. The various advantages and limitations of the different systems are contrasted. It is posited that only through complementary use of these models will salient research questions be able to be addressed.

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

Cited by 86 publications

References 47 publications

Generation of intestinal organoids derived from human pluripotent stem cells for drug testing

Generation of intestinal organoids derived from human pluripotent stem cells for drug testing

Isolation and characterisation of mouse intestinal mesoangioblasts

Models of the Gut for Analyzing the Impact of Food and Drugs

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