Fluorescent labelling of intestinal epithelial cells reveals independent long-lived intestinal stem cells in a crypt

Weaning is a stressful event associated with gastrointestinal disorders and increased disease susceptibility. Many studies have reported the changes that happened in the gut of various mammals such as pigs and rats after weaning. These findings suggest that the development of intestinal tract mainly is affected at the time of weaning through interfering in the differentiation and proliferation of intestinal stem cells. Weaning stress stimulates the rapid differentiation and proliferation of intestinal stem cells in order to adjust to changes caused by weaning, which are mainly manifested as deeper crypt depth and decreased intestine villus height. However, the accelerated cellular process may lead to an increase in the proportion of immature intestinal epithelial cells and goblet cells, which affect intestinal permeability and reduce the gut-barrier function against toxins and pathogens. This review briefly describes the effects coforticotrophin-releasing factor (CRF), epidermal growth factor (EGF) and polyamines on the differentiation and proliferation of intestinal stem cells after weaning and discusses its possible underlying regulatory mechanisms. Firstly, weaning stress activates CRF to binds its receptors, which induces proinflammatory responses and promote rapid differentiation and proliferation of intestinal stem cells to a larger fraction of immature intestinal epithelial cells and goblet cells. Secondly, the lack of EGF after weaning inhibits the expression of goblet cell maturation factors and makes it difficult for goblet cells and intestinal epithelial cells to mature. Finally, diet and endogenous synthesis lead to excessive polyamines in the intestine, which promote the proliferation of intestinal stem cells by regulating the expression of human antigen R (HuR) and other related genes at the time of weaning.

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Human Small Intestinal Epithelial Cells Differentiated from Adult Intestinal Stem Cells as a Novel System for Predicting Oral Drug Absorption in Humans

Drug Metabolism and Disposition ◽

10.1124/dmd.114.059493 ◽

2014 ◽

Vol 42 (11) ◽

pp. 1947-1954 ◽

Cited By ~ 41

Author(s):

Toru Takenaka ◽

Naomoto Harada ◽

Jiro Kuze ◽

Masato Chiba ◽

Takahiro Iwao ◽

...

Keyword(s):

Stem Cells ◽

Epithelial Cells ◽

Drug Absorption ◽

Intestinal Epithelial Cells ◽

Intestinal Stem Cells ◽

Oral Drug ◽

Intestinal Epithelial ◽

Oral Drug Absorption ◽

Small Intestinal

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Arid1a is essential for intestinal stem cells through Sox9 regulation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1804858116 ◽

2019 ◽

Vol 116 (5) ◽

pp. 1704-1713 ◽

Cited By ~ 5

Author(s):

Yukiko Hiramatsu ◽

Akihisa Fukuda ◽

Satoshi Ogawa ◽

Norihiro Goto ◽

Kozo Ikuta ◽

...

Keyword(s):

Stem Cells ◽

Epithelial Cells ◽

Functional Role ◽

Molecular Mechanisms ◽

Intestinal Epithelial Cells ◽

Lineage Tracing ◽

Intestinal Stem Cells ◽

Intestinal Epithelial ◽

Intestinal Homeostasis ◽

Self Renewal

Inactivating mutations of Arid1a, a subunit of the Switch/sucrose nonfermentable chromatin remodeling complex, have been reported in multiple human cancers. Intestinal deletion of Arid1a has been reported to induce colorectal cancer in mice; however, its functional role in intestinal homeostasis remains unclear. We investigated the functional role of Arid1a in intestinal homeostasis in mice. We found that intestinal deletion of Arid1a results in loss of intestinal stem cells (ISCs), decreased Paneth and goblet cells, disorganized crypt-villous structures, and increased apoptosis in adult mice. Spheroids did not develop from intestinal epithelial cells deficient for Arid1a. Lineage-tracing experiments revealed that Arid1a deletion in Lgr5+ ISCs leads to impaired self-renewal of Lgr5+ ISCs but does not perturb intestinal homeostasis. The Wnt signaling pathway, including Wnt agonists, receptors, and target genes, was strikingly down-regulated in Arid1a-deficient intestines. We found that Arid1a directly binds to the Sox9 promoter to support its expression. Remarkably, overexpression of Sox9 in intestinal epithelial cells abrogated the above phenotypes, although Sox9 overexpression in intestinal epithelial cells did not restore the expression levels of Wnt agonist and receptor genes. Furthermore, Sox9 overexpression permitted development of spheroids from Arid1a-deficient intestinal epithelial cells. In addition, deletion of Arid1a concomitant with Sox9 overexpression in Lgr5+ ISCs restores self-renewal in Arid1a-deleted Lgr5+ ISCs. These results indicate that Arid1a is indispensable for the maintenance of ISCs and intestinal homeostasis in mice. Mechanistically, this is mainly mediated by Sox9. Our data provide insights into the molecular mechanisms underlying maintenance of ISCs and intestinal homeostasis.

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Human Primary Intestinal Epithelial Cells as an ImprovedIn VitroModel for Cryptosporidium parvum Infection

Infection and Immunity ◽

10.1128/iai.01131-12 ◽

2013 ◽

Vol 81 (6) ◽

pp. 1996-2001 ◽

Cited By ~ 44

Author(s):

Alejandro Castellanos-Gonzalez ◽

Miguel M. Cabada ◽

Joan Nichols ◽

Guillermo Gomez ◽

A. Clinton White

Keyword(s):

Stem Cells ◽

Epithelial Cells ◽

Intestinal Epithelial Cells ◽

Intestinal Stem Cells ◽

Intestinal Epithelial ◽

Content Type ◽

Host Parasite Interactions ◽

Intestinal Pathogens ◽

Novel Method ◽

Host Parasite

ABSTRACTThe study of human intestinal pathogens has been limited by the lack of methods for the long-term culture of primary human intestinal epithelial cells (PECs). The development of infection models with PECs would allow a better understanding of host-parasite interactions. The objective of this study was to develop a novel method for prolongedin vitrocultivation of PECs that can be used to studyCryptosporidiuminfection. We isolated intact crypts from human intestines removed during weight loss surgery. The fragments of intestinal layers were cultivated with culture medium supplemented with growth factors and antiapoptotic molecules. After 7 days, the PECs formed self-regenerating cell clusters, forming villi that resemble intestinal epithelium. The PECs proliferated and remained viable for at least 60 days. The cells expressed markers for intestinal stem cells, epithelial cells, and mature enterocytes. The PECs were infected withCryptosporidium. In contrast to older models in which parasite numbers decay, the burden of parasites increased for >120 h. In summary, we describe here a novel method for the cultivation of self-regenerating human epithelial cells from small intestinal crypts, which contain both intestinal stem cells and mature villus cells. We present data that suggest these cells supportCryptosporidiumbetter than existing cell lines. PECs should provide an improved tool for studying host-parasite interactions involvingCryptosporidiumand other intestinal pathogens.

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Application of Intestinal Epithelial Cells Differentiated from Human Induced Pluripotent Stem Cells for Studies of Prodrug Hydrolysis and Drug Absorption in the Small Intestine

Drug Metabolism and Disposition ◽

10.1124/dmd.118.083246 ◽

2018 ◽

Vol 46 (11) ◽

pp. 1497-1506 ◽

Cited By ~ 13

Author(s):

Takanori Akazawa ◽

Shinpei Yoshida ◽

Shuichi Ohnishi ◽

Takushi Kanazu ◽

Makoto Kawai ◽

...

Keyword(s):

Stem Cells ◽

Small Intestine ◽

Epithelial Cells ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

Drug Absorption ◽

Intestinal Epithelial Cells ◽

Intestinal Epithelial ◽

Induced Pluripotent ◽

Prodrug Hydrolysis

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The Hippo Pathway Effector YAP1 Regulates Intestinal Epithelial Cell Differentiation

Cells ◽

10.3390/cells9081895 ◽

2020 ◽

Vol 9 (8) ◽

pp. 1895 ◽

Cited By ~ 1

Author(s):

Sepideh Fallah ◽

Jean-François Beaulieu

Keyword(s):

Stem Cells ◽

Cell Differentiation ◽

Epithelial Cells ◽

Cell Line ◽

Intestinal Epithelial Cells ◽

Hippo Pathway ◽

Intestinal Cell ◽

Intestinal Epithelial ◽

The Hippo Pathway

The human intestine is covered by epithelium, which is continuously replaced by new cells provided by stem cells located at the bottom of the glands. The maintenance of intestinal stem cells is supported by a niche which is composed of several signaling proteins including the Hippo pathway effectors YAP1/TAZ. The role of YAP1/TAZ in cell proliferation and regeneration is well documented but their involvement on the differentiation of intestinal epithelial cells is unclear. In the present study, the role of YAP1/TAZ on the differentiation of intestinal epithelial cells was investigated using the HT29 cell line, the only multipotent intestinal cell line available, with a combination of knockdown approaches. The expression of intestinal differentiation cell markers was tested by qPCR, Western blot, indirect immunofluorescence and electron microscopy analyses. The results show that TAZ is not expressed while the abolition of YAP1 expression led to a sharp increase in goblet and absorptive cell differentiation and reduction of some stem cell markers. Further studies using double knockdown experiments revealed that most of these effects resulting from YAP1 abolition are mediated by CDX2, a key intestinal cell transcription factor. In conclusion, our results indicate that YAP1/TAZ negatively regulate the differentiation of intestinal epithelial cells through the inhibition of CDX2 expression.

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T helper cells modulate intestinal stem cell renewal and differentiation

10.1101/217133 ◽

2017 ◽

Cited By ~ 2

Author(s):

Moshe Biton ◽

Adam L. Haber ◽

Semir Beyaz ◽

Noga Rogel ◽

Christopher Smillie ◽

...

Keyword(s):

Stem Cells ◽

T Cells ◽

Epithelial Cells ◽

Immune Cell ◽

Intestinal Epithelial Cells ◽

Cell Types ◽

Intestinal Epithelial ◽

T Helper ◽

Self Renewal

AbstractIn the small intestine, a cellular niche of diverse accessory cell types supports the rapid generation of mature epithelial cell types through self-renewal, proliferation, and differentiation of intestinal stem cells (ISCs). However, not much is known about interactions between immune cells and ISCs, and it is unclear if and how immune cell dynamics affect eventual ISC fate or the balance between self-renewal and differentiation. Here, we used single-cell RNA-seq (scRNA-Seq) of intestinal epithelial cells (IECs) to identify new mechanisms for ISC–immune cell interactions. Surprisingly, MHC class II (MHCII) is enriched in two distinct subsets of Lgr5+ crypt base columnar ISCs, which are also distinguished by higher proliferation rates. Using co-culture of T cells with intestinal organoids, cytokine stimulations, and in vivo mouse models, we confirm that CD4+ T helper (Th) cells communicate with ISCs and affect their differentiation, in a manner specific to the Th subtypes and their signature cytokines and dependent on MHCII expression by ISCs. Specific inducible knockout of MHCII in intestinal epithelial cells in mice in vivo results in expansion of the ISC pool. Mice lacking T cells have expanded ISC pools, whereas specific depletion of Treg cells in vivo results in substantial reduction of ISC numbers. Our findings show that interactions between Th cells and ISCs mediated via MHCII expressed in intestinal epithelial stem cells help orchestrate tissue-wide responses to external signals.

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