Transplantation of intestinal organoids into a mouse model of colitis

Watanabe, Satoshi; Kobayashi, Shinzo; Ogasawara, Nobuhiko; Okamoto, Ryuichi; Nakamura, Toshio; Watanabe, Mamoru; Jensen, Kim B.; Yui, Satoru

doi:10.1038/s41596-021-00658-3

Cited by 57 publications

(56 citation statements)

References 47 publications

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“…The establishment of organoids was performed as previously described [ 11 ]. Harvested crypts were suspended in 30μl Matrigel (Corning, #356231) or cellmatrix type IA (Collagen Type I, Nitta Gelatin, #631-00651) and plated onto a 24-well plate (Flat bottom; Corning, #3524).…”

Section: Methodsmentioning

confidence: 99%

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Collagen type I-mediated mechanotransduction controls epithelial cell fate conversion during intestinal inflammation

et al. 2022

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Background The emerging concepts of fetal-like reprogramming following tissue injury have been well recognized as an important cue for resolving regenerative mechanisms of intestinal epithelium during inflammation. We previously revealed that the remodeling of mesenchyme with collagen fibril induces YAP/TAZ-dependent fate conversion of intestinal/colonic epithelial cells covering the wound bed towards fetal-like progenitors. To fully elucidate the mechanisms underlying the link between extracellular matrix (ECM) remodeling of mesenchyme and fetal-like reprogramming of epithelial cells, it is critical to understand how collagen type I influence the phenotype of epithelial cells. In this study, we utilize collagen sphere, which is the epithelial organoids cultured in purified collagen type I, to understand the mechanisms of the inflammatory associated reprogramming. Resolving the entire landscape of regulatory networks of the collagen sphere is useful to dissect the reprogrammed signature of the intestinal epithelium. Methods We performed microarray, RNA-seq, and ATAC-seq analyses of the murine collagen sphere in comparison with Matrigel organoid and fetal enterosphere (FEnS). We subsequently cultured human colon epithelium in collagen type I and performed RNA-seq analysis. The enriched genes were validated by gene expression comparison between published gene sets and immunofluorescence in pathological specimens of ulcerative colitis (UC). Results The murine collagen sphere was confirmed to have inflammatory and regenerative signatures from RNA-seq analysis. ATAC-seq analysis confirmed that the YAP/TAZ-TEAD axis plays a central role in the induction of the distinctive signature. Among them, TAZ has implied its relevant role in the process of reprogramming and the ATAC-based motif analysis demonstrated not only Tead proteins, but also Fra1 and Runx2, which are highly enriched in the collagen sphere. Additionally, the human collagen sphere also showed a highly significant enrichment of both inflammatory and fetal-like signatures. Immunofluorescence staining confirmed that the representative genes in the human collagen sphere were highly expressed in the inflammatory region of ulcerative colitis. Conclusions Collagen type I showed a significant influence in the acquisition of the reprogrammed inflammatory signature in both mice and humans. Dissection of the cell fate conversion and its mechanisms shown in this study can enhance our understanding of how the epithelial signature of inflammation is influenced by the ECM niche.

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Section: Methodsmentioning

confidence: 99%

“…The establishment of organoids was performed as previously described [11]. Harvested crypts were suspended in 30μl Matrigel (Corning, #356231) or cellmatrix type 1).…”

Section: Primary Cultures Of Adult Murine Small Intestinal Epithelium...mentioning

confidence: 99%

Collagen type I-mediated mechanotransduction controls epithelial cell fate conversion during intestinal inflammation

et al. 2022

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“…Because ASC-derived organoids can be grown from most epithelial tissues, there could be similar benefits in transplanting CRISPR engineered organoids to patients. Moreover, organoids from most epithelial tissues can be maintained indefinitely and can be split and propagated rapidly to yield the biomass needed for successful transplantation 156 . Current efforts to transplant ASC-derived organoids have shown some success in mouse intestine 124,125,157 .…”

Section: Crispr From Bench To Bedsidementioning

confidence: 99%

CRISPR engineering in organoids for gene repair and disease modelling

Geurts

2023

Nat Rev Bioeng

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Organoids bridge the gap between 2D cell lines and in vivo studies. With their 3D organization and cellular heterogeneity, adult stem cell-derived organoids closely resemble their tissue of origin. The development of CRISPR-mediated genome engineering and the recent additions of base and prime editing to the CRISPR toolbox have greatly simplified the generation of exact, isogenic models for Mendelian diseases. Here, we review recent developments in CRISPR-mediated genome engineering and its application in human adult-stem-cell-derived organoids in the construction of isogenic disease models. These models allow accurate qualification of the impact of allelic disease variants observed in patients. Furthermore, we discuss the use of organoids as models for safety and efficacy of CRISPR for gene repair. Although transplantation Key points• CRISPR-Cas9-mediated genome engineering acts by introducing double-stranded DNA breaks into the genome. The damage repair process can be used for gene knockout or precise targeted introduction of exogenous DNA.• Next-generation CRISPR tools, including base and prime editing, allow for induction of precise base changes and small insertions and deletions, bypassing potentially deleterious double-stranded DNA breaks.• Owing to their 3D organization, adult-stem-cell-derived organoids closely resemble the tissue of origin and are therefore a good model system to study human health and disease.• CRISPR-Cas9-mediated genome engineering can be used to create isogenic models to investigate the onset, cause and treatment of human diseases.• CRISPR tools can be benchmarked for efficiency and safety by studying gene repair ex vivo in adult-stem-cell-derived organoids, facilitating CRISPR-Cas9 clinical translation.• Ex vivo repaired adult-stem-cell-derived organoids can potentially be transplanted into patients to relieve disease phenotypes.Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author selfarchiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

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“…Alternative models include injecting colon cancer cells subcutaneously or in the cecum; however, that does not recapitulate the natural microenvironment for colon cancer cells. While several publications have recently described the transplantation of normal organoids into injured colonic mucosa, 1 , 2 here we describe a protocol that enables the generation of primary colonic tumors that can metastasize into the liver. This rapid and reproducible protocol also significantly reduces mouse mortality due to the procedure.…”

Section: Before You Beginmentioning

confidence: 99%