Generation of small intestinal organoids for experimental intestinal physiology

Capeling, Meghan M.; Mulero‐Russe, Adriana; Cieza, Roberto J.; Tsai, Yu-Hwai; Garcı́a, Andrés J.; Hill, David R.

doi:10.1016/bs.mcb.2020.03.007

Cited by 19 publications

(19 citation statements)

References 53 publications

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“…Generation and culture of in vitro human intestinal organoids hESCs were differentiated into human intestinal organoids (HIOs) based on the previously described protocol (Capeling et al, 2020;Spence et al, 2011;Tsai et al, 2017). Briefly, hESCs were patterned into definitive endoderm (DE) by culturing in the presence of Activin A (100 ng/mL) in RPMI-1640 media for 3 days with increasing concentrations of HyClone FBS respectively (0%, 0.2%, 2%).…”

Section: Methods Detailsmentioning

confidence: 99%

“…The media was changed every 4-5 days. The HIOs were passaged when they outgrew matrigel and/or accumulated excessive internal debris (Capeling et al, 2020). The mini gut basal media is composed of the following components: Advanced DMEM/F-12 (Life Technologies, 12634), 1x B27 supplement (Life Technologies, 17504044), 2 mM L-Glutamine (Life Technologies, 25030), 15 mM HEPES (Life Technologies, 15630080).…”

Section: Methods Detailsmentioning

confidence: 99%

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Charting human development using a multi-endodermal organ atlas and organoid models

Kilik

Holloway

et al. 2021

Cell

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Section: Methods Detailsmentioning

confidence: 99%

Section: Methods Detailsmentioning

confidence: 99%

Charting human development using a multi-endodermal organ atlas and organoid models

Kilik

Holloway

et al. 2021

Cell

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“…Small intestinal crypt isolation and culture were performed as previously described (14). Brie y, remove the mesentery, fat and blood vessels attached to the surface of the intestine, and the feces inside the intestine, cut the intestinal cavity after the longitudinal axis and cut the tissue into small pieces, then incubated in 5 mM EDTA in PBS, on ice for 30 min.…”

Section: Crypt Isolation Organoid Culture and Quanti Cationmentioning

confidence: 99%

Interleukin-10 expands transit-amplifying cells while depleting Lgr5+ stem cells via inhibition of Wnt and notch signaling

Deng

Yang

et al. 2020

Biochemical and Biophysical Research Communications

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Background & Aims: Epithelial regeneration is essential for homeostasis and mucosal barrier repair. In infectious and immune-mediated intestinal diseases, interleukin (IL)-10 is thought to enhance these processes. We aimed to de ne the mechanism by which IL-10 played in mucosal healing or injury. Methods: Intestinal stem cells (ISCs) cultures and mice were treated with recombinant mice IL-10 (rmIL-10). The level of cell proliferation, differentiation, death and related signaling pathways for self-renewal of ISCs were measured in vitro and in vivo. Results: It was uncovered that rmIL-10 increased the size and death, but reduced the total number of organoids. In addition, rmIL-10 depleted Lgr5 + ISCs and reduced epithelial proliferation, but enhanced the differentiation of epithelial cells and expanded numbers of transit-amplifying (TA) cells. These changes are related to the decrease of Wnt and Notch signals in vivo and in vitro. Meanwhile, increased expression of Paneth cells and decreased expression of enteroendocrine cells and goblet cells were induced by rmIL-10. Conclusions: IL-10 reduces the survival of Lgr5 + ISCs and proliferation of epithelial cells by inhibiting Notch and Wnt signaling, but promotes enhanced the differentiation of epithelial cells and expanded numbers of TA cells. Therefore, IL-10 acts as an anti-in ammatory factor, but may damage intestinal mucosa repair and maybe a potential target for the treatment of intestinal injury.

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“…For example, by understanding that WNT signaling is important for maintaining intestinal stem cell (ISC) homeostasis (Muncan et al, 2006;Pinto et al, 2003;Sansom et al, 2004), that blockade of Bone Morphogenetic Protein (BMP) signaling by NOGGIN (NOG) promotes ectopic crypt formation (Haramis et al, 2004), and that Epidermal Growth Factor (EGF) is a potent stimulator of proliferation (Goodlad et al, 1987;Ulshen et al, 1986), it was determined that WNTs, RSPON-DINs (RSPOs), NOG, and EGF can be utilized to expand and main-tain ISCs in culture as three-dimensional intestinal organoids (Ootani et al, 2009;Sato et al, 2009Sato et al, , 2011b. This information has been leveraged to expand and culture human pluripotent stem cell-derived intestinal organoids in vitro (Capeling et al, 2020;Finkbeiner et al, 2015;Spence et al, 2011;Wells and Spence, 2014).…”

Section: Introductionmentioning

confidence: 99%