R-spondin 3 promotes stem cell recovery and epithelial regeneration in the colon

Abstract The colonic epithelial turnover is driven by crypt-base stem cells that express the R-spondin receptor Lgr5. Signals that regulate epithelial regeneration upon stem cell injury are largely unknown. Here, we explore the dynamics of Wnt signaling in the colon. We identify two populations of cells with active Wnt signaling: highly proliferative Lgr5+/Axin2+ cells, as well as secretory Lgr5−/Axin2+ cells. Upon Lgr5+ cell depletion, these cells are recruited to contribute to crypt regeneration. Chemical injury induced by DSS leads to a loss of both Lgr5+ cells and Axin2+ cells and epithelial regeneration is driven by Axin2− cells, including differentiated Krt20+ surface enterocytes. Regeneration requires stromal Rspo3, which is present at increased levels upon injury and reprograms Lgr5− but Lgr4+ differentiated cells. In contrast, depletion of stromal Rspo3 impairs crypt regeneration, even upon mild injury. We demonstrate that Rspo3 is essential for epithelial repair via induction of Wnt signaling in differentiated cells.

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Clostridioides difficileinfection damages colonic stem cells via TcdB, impairing epithelial repair and recovery from disease

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1915255117 ◽

2020 ◽

Vol 117 (14) ◽

pp. 8064-8073 ◽

Cited By ~ 6

Author(s):

Steven J. Mileto ◽

Thierry Jardé ◽

Kevin O. Childress ◽

Jaime L. Jensen ◽

Ashleigh P. Rogers ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Disease Recurrence ◽

Gastrointestinal Infections ◽

Cell Compartment ◽

Epithelial Repair ◽

Epithelial Regeneration ◽

Clostridioides Difficile ◽

Stem Cell Compartment ◽

Colonic Stem Cells

Gastrointestinal infections often induce epithelial damage that must be repaired for optimal gut function. While intestinal stem cells are critical for this regeneration process [R. C. van der Wath, B. S. Gardiner, A. W. Burgess, D. W. Smith,PLoS One8, e73204 (2013); S. Kozaret al.,Cell Stem Cell13, 626–633 (2013)], how they are impacted by enteric infections remains poorly defined. Here, we investigate infection-mediated damage to the colonic stem cell compartment and how this affects epithelial repair and recovery from infection. Using the pathogenClostridioides difficile,we show that infection disrupts murine intestinal cellular organization and integrity deep into the epithelium, to expose the otherwise protected stem cell compartment, in a TcdB-mediated process. Exposure and susceptibility of colonic stem cells to intoxication compromises their function during infection, which diminishes their ability to repair the injured epithelium, shown by altered stem cell signaling and a reduction in the growth of colonic organoids from stem cells isolated from infected mice. We also show, using both mouse and human colonic organoids, that TcdB from epidemic ribotype 027 strains does not require Frizzled 1/2/7 binding to elicit this dysfunctional stem cell state. This stem cell dysfunction induces a significant delay in recovery and repair of the intestinal epithelium of up to 2 wk post the infection peak. Our results uncover a mechanism by which an enteric pathogen subverts repair processes by targeting stem cells during infection and preventing epithelial regeneration, which prolongs epithelial barrier impairment and creates an environment in which disease recurrence is likely.

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A29 HOPX LABELS A COLONIC STEM CELL THAT CONTRIBUTES TO COLONIC REGENERATION BUT NOT COLONIC TUMORS

Journal of the Canadian Association of Gastroenterology ◽

10.1093/jcag/gwz047.028 ◽

2020 ◽

Vol 3 (Supplement_1) ◽

pp. 35-37

Author(s):

V Iablokov ◽

H J Good ◽

A E Shin ◽

E N Fazio ◽

J W Loggie ◽

...

Keyword(s):

Stem Cells ◽

Colon Cancer ◽

Stem Cell ◽

Colorectal Adenomas ◽

Stem Cell Population ◽

Genetic Lineage ◽

Cellular Origin ◽

Epithelial Regeneration ◽

Histological Damage ◽

Crypt Base

Abstract Background Colorectal cancer is the 2nd leading cause of cancer death in Canada. In rapidly dividing tissues such as the intestine or colon, only long-lived, multipotent, self-renewing tissue stem cells have longevity to accumulate mutations and serve as the cellular origin of cancer. In the small intestine, genetic fate mapping studies have demonstrated that there are at least two principal stem cell pools: actively cycling, crypt base cells expressing Lgr5, and quiescent cells situated above the crypt base. Clevers and colleagues have previously shown that Lgr5-expressing cells can give rise to cancer upon mutation. Interestingly, when Lgr5+ stem cells are selectively “killed”, intestinal integrity remains intact and other stem cells restore homeostasis. To determine whether another stem cell population can give rise to cancer in the colon, we examined whether the atypical homeobox protein Hopx, marks stem cells in the colon and whether these cells can give rise to colon cancer. Aims In the present study, we aim to determine whether Hopx-expressing cells are colonic stem cells that contribute to gut healing and can give rise to colonic tumours following the loss of APC. Methods To determine whether Hopx expressing cells show stemness, we crossed Hopx-CreERT mice to R26-TdTomato reporter mice. We then conducted genetic lineage tracing studies in the colon during homeostasis and following dextran sodium sulphate (DSS)-induced colitis. To test the function of Hopx expressing cells, Hopx-CreERT mice were also crossed to R26DTR mice and treated with diphtheria toxin (DT) following tamoxifen. These mice were then exposed to either normal drinking water or DSS to determine the role of Hopx+ cells in colonic regeneration. To test whether Hopx expressing cells can serve as a cellular origin for colon cancer, Hopx-CreERT mice were crossed to Apcf/f (floxed) mice. Results Consistent with the labeling of a stem cell, following tamoxifen, Hopx+ cells expressing tdTomato expanded to lineage trace full colonic crypts within 7 days, and labelling was persistent for greater than 6 months. Interestingly, ablation of Hopx+ cells with DT did not alter weight, histological damage or survival during normal homeostasis, however, Hopx+ cell ablation in mice treated with DSS resulted in increased histological damage. Surprisingly, loss of APC in Hopx-expressing cells did not induce colonic adenomas even after 8 months following tamoxifen administration. Conclusions These findings prove that Hopx expressing cells identify a novel colonic stem cell pool that is redundant in homeostasis, but in the context of injury, are essential for epithelial regeneration. Interestingly, Hopx+ cells do not have the capacity to give rise to colorectal adenomas upon loss of the APC gene. Funding Agencies CIHR

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Axin2 marks quiescent hair follicle bulge stem cells that are maintained by autocrine Wnt/β-catenin signaling

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1601599113 ◽

2016 ◽

Vol 113 (11) ◽

pp. E1498-E1505 ◽

Cited By ~ 52

Author(s):

Xinhong Lim ◽

Si Hui Tan ◽

Ka Lou Yu ◽

Sophia Beng Hui Lim ◽

Roeland Nusse

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Wnt Signaling ◽

Hair Follicle ◽

Extended Period ◽

Lineage Tracing ◽

Hair Cycle ◽

Cell Potential ◽

Differentiated Cells ◽

Hair Follicle Stem Cells

How stem cells maintain their identity and potency as tissues change during growth is not well understood. In mammalian hair, it is unclear how hair follicle stem cells can enter an extended period of quiescence during the resting phase but retain stem cell potential and be subsequently activated for growth. Here, we use lineage tracing and gene expression mapping to show that the Wnt target gene Axin2 is constantly expressed throughout the hair cycle quiescent phase in outer bulge stem cells that produce their own Wnt signals. Ablating Wnt signaling in the bulge cells causes them to lose their stem cell potency to contribute to hair growth and undergo premature differentiation instead. Bulge cells express secreted Wnt inhibitors, including Dickkopf (Dkk) and secreted frizzled-related protein 1 (Sfrp1). However, the Dickkopf 3 (Dkk3) protein becomes localized to the Wnt-inactive inner bulge that contains differentiated cells. We find that Axin2 expression remains confined to the outer bulge, whereas Dkk3 continues to be localized to the inner bulge during the hair cycle growth phase. Our data suggest that autocrine Wnt signaling in the outer bulge maintains stem cell potency throughout hair cycle quiescence and growth, whereas paracrine Wnt inhibition of inner bulge cells reinforces differentiation.

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Plasticity of Intestinal Epithelium: Stem Cell Niches and Regulatory Signals

International Journal of Molecular Sciences ◽

10.3390/ijms22010357 ◽

2020 ◽

Vol 22 (1) ◽

pp. 357

Author(s):

Ken Kurokawa ◽

Yoku Hayakawa ◽

Kazuhiko Koike

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Inflammatory Cytokines ◽

Intestinal Epithelium ◽

Intestinal Stem Cells ◽

Signal Pathways ◽

Epithelial Regeneration ◽

Cell Functions ◽

Differentiated Cells ◽

Stem Cell Niches

The discovery of Lgr5+ intestinal stem cells (ISCs) triggered a breakthrough in the field of ISC research. Lgr5+ ISCs maintain the homeostasis of the intestinal epithelium in the steady state, while these cells are susceptible to epithelial damage induced by chemicals, pathogens, or irradiation. During the regeneration process of the intestinal epithelium, more quiescent +4 stem cells and short-lived transit-amplifying (TA) progenitor cells residing above Lgr5+ ISCs undergo dedifferentiation and act as stem-like cells. In addition, several recent reports have shown that a subset of terminally differentiated cells, including Paneth cells, tuft cells, or enteroendocrine cells, may also have some degree of plasticity in specific situations. The function of ISCs is maintained by the neighboring stem cell niches, which strictly regulate the key signal pathways in ISCs. In addition, various inflammatory cytokines play critical roles in intestinal regeneration and stem cell functions following epithelial injury. Here, we summarize the current understanding of ISCs and their niches, review recent findings regarding cellular plasticity and its regulatory mechanism, and discuss how inflammatory cytokines contribute to epithelial regeneration.

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GSK3β, a Master Kinase in the Regulation of Adult Stem Cell Behavior

Cells ◽

10.3390/cells10020225 ◽

2021 ◽

Vol 10 (2) ◽

pp. 225

Author(s):

Claire Racaud-Sultan ◽

Nathalie Vergnolle

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Adult Stem Cells ◽

Glycogen Synthase ◽

Inflammatory Diseases ◽

Adult Stem Cell ◽

Leukemic Cells ◽

Cell Phenotype ◽

Epithelial Regeneration ◽

Cell Functions

In adult stem cells, Glycogen Synthase Kinase 3β (GSK3β) is at the crossroad of signaling pathways controlling survival, proliferation, adhesion and differentiation. The microenvironment plays a key role in the regulation of these cell functions and we have demonstrated that the GSK3β activity is strongly dependent on the engagement of integrins and protease-activated receptors (PARs). Downstream of the integrin α5β1 or PAR2 activation, a molecular complex is organized around the scaffolding proteins RACK1 and β-arrestin-2 respectively, containing the phosphatase PP2A responsible for GSK3β activation. As a consequence, a quiescent stem cell phenotype is established with high capacities to face apoptotic and metabolic stresses. A protective role of GSK3β has been found for hematopoietic and intestinal stem cells. Latters survived to de-adhesion through PAR2 activation, whereas formers were protected from cytotoxicity through α5β1 engagement. However, a prolonged activation of GSK3β promoted a defect in epithelial regeneration and a resistance to chemotherapy of leukemic cells, paving the way to chronic inflammatory diseases and to cancer resurgence, respectively. In both cases, a sexual dimorphism was measured in GSK3β-dependent cellular functions. GSK3β activity is a key marker for inflammatory and cancer diseases allowing adjusted therapy to sex, age and metabolic status of patients.

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Immunostaining of Lgr5, an Intestinal Stem Cell Marker, in Normal and Premalignant Human Gastrointestinal Tissue

The Scientific World JOURNAL ◽

10.1100/tsw.2008.148 ◽

2008 ◽

Vol 8 ◽

pp. 1168-1176 ◽

Cited By ~ 76

Author(s):

Laren Becker ◽

Qin Huang ◽

Hiroshi Mashimo

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Stem Cell Niche ◽

Intestinal Stem Cells ◽

Stem Cell Marker ◽

Tumor Initiating Cells ◽

Colonic Adenomas ◽

Potential Marker ◽

Cell Niche ◽

Crypt Base

Lgr5 has recently been identified as a murine marker of intestinal stem cells. Its expression has not been well characterized in human gastrointestinal tissues, but has been reported in certain cancers. With the increasing appreciation for the role of cancer stem cells or tumor-initiating cells in certain tumors, we sought to explore the expression of Lgr5 in normal and premalignant human gastrointestinal tissues. Using standard immunostaining, we compared expression of Lgr5 in normal colon and small intestine vs. small intestinal and colonic adenomas and Barrett's esophagus. In the normal tissue, Lgr5 was expressed in the expected stem cell niche, at the base of crypts, as seen in mice. However, in premalignant lesions, Lgr5+cells were not restricted to the crypt base. Additionally, their overall numbers were increased. In colonic adenomas, Lgr5+cells were commonly found clustered at the luminal surface and rarely at the crypt base. Finally, we compared immunostaining of Lgr5 with that of CD133, a previously characterized marker for tumor-initiating cells in colon cancer, and found that they identified distinct subpopulations of cells that were in close proximity, but did not costain. Our findings suggest that (1) Lgr5 is a potential marker of intestinal stem cells in humans and (2) loss of restriction to the stem cell niche is an early event in the premalignant transformation of stem cells and may play a role in carcinogenesis.

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me31B regulates stem cell homeostasis by preventing excess dedifferentiation in the Drosophila male germline

Journal of Cell Science ◽

10.1242/jcs.258757 ◽

2021 ◽

Author(s):

Lindy Jensen ◽

Zsolt G. Venkei ◽

George J. Watase ◽

Bitarka Bisai ◽

Scott Pletcher ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Critical Role ◽

Stem Cell Population ◽

Germline Stem Cells ◽

Cell Identity ◽

Stem Cell Maintenance ◽

Differentiated Cells ◽

Male Germline ◽

Elevated Frequency

Tissue-specific stem cells maintain tissue homeostasis by providing a continuous supply of differentiated cells throughout the life of organisms. Differentiated/differentiating cells can revert back to a stem cell identity via dedifferentiation to help maintain the stem cell pool beyond the lifetime of individual stem cells. Although dedifferentiation is important to maintain the stem cell population, it is speculated to underlie tumorigenesis. Therefore, this process must be tightly controlled. Here we show that a translational regulator me31B plays a critical role in preventing excess dedifferentiation in the Drosophila male germline: in the absence of me31B, spermatogonia (SGs) dedifferentiate into germline stem cells (GSCs) at a dramatically elevated frequency. Our results show that the excess dedifferentiation is likely due to misregulation of nos, a key regulator of germ cell identity and GSC maintenance. Taken together, our data reveal negative regulation of dedifferentiation to balance stem cell maintenance with differentiation.

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Silencing of long noncoding RNA HOXA11-AS inhibits the Wnt signaling pathway via the upregulation of HOXA11 and thereby inhibits the proliferation, invasion, and self-renewal of hepatocellular carcinoma stem cells

Experimental & Molecular Medicine ◽

10.1038/s12276-019-0328-x ◽

2019 ◽

Vol 51 (11) ◽

pp. 1-20 ◽

Cited By ~ 2

Author(s):

Jun-Cheng Guo ◽

Yi-Jun Yang ◽

Jin-Fang Zheng ◽

Jian-Quan Zhang ◽

Min Guo ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Stem Cells ◽

Stem Cell ◽

Wnt Signaling ◽

Signaling Pathway ◽

Methylation Level ◽

Wnt Signaling Pathway ◽

The Self ◽

Self Renewal

AbstractHepatocellular carcinoma (HCC) is a major cause of cancer-related deaths, but its molecular mechanisms are not yet well characterized. Long noncoding RNAs (lncRNAs) play crucial roles in tumorigenesis, including that of HCC. However, the role of homeobox A11 antisense (HOXA11-AS) in determining HCC stem cell characteristics remains to be explained; hence, this study aimed to investigate the effects of HOXA11-AS on HCC stem cell characteristics. Initially, the expression patterns of HOXA11-AS and HOXA11 in HCC tissues, cells, and stem cells were determined. HCC stem cells, successfully sorted from Hep3B and Huh7 cells, were transfected with short hairpin or overexpression plasmids for HOXA11-AS or HOXA11 overexpression and depletion, with an aim to study the influences of these mediators on the self-renewal, proliferation, migration, and tumorigenicity of HCC stem cells in vivo. Additionally, the potential relationship and the regulatory mechanisms that link HOXA11-AS, HOXA11, and the Wnt signaling pathway were explored through treatment with Dickkopf-1 (a Wnt signaling pathway inhibitor). HCC stem cells showed high expression of HOXA11-AS and low expression of HOXA11. Both HOXA11-AS silencing and HOXA11 overexpression suppressed the self-renewal, proliferation, migration, and tumorigenicity of HCC stem cells in vivo, as evidenced by the decreased expression of cancer stem cell surface markers (CD133 and CD44) and stemness-related transcription factors (Nanog, Sox2, and Oct4). Moreover, silencing HOXA11-AS inactivated the Wnt signaling pathway by decreasing the methylation level of the HOXA11 promoter, thereby inhibiting HCC stem cell characteristics. Collectively, this study suggested that HOXA11-AS silencing exerts an antitumor effect, suppressing HCC development via Wnt signaling pathway inactivation by decreasing the methylation level of the HOXA11 promoter.

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The Hippo–YAP Signaling as Guardian in the Pool of Intestinal Stem Cells

Biomedicines ◽

10.3390/biomedicines8120560 ◽

2020 ◽

Vol 8 (12) ◽

pp. 560

Author(s):

Yoojin Seo ◽

So-Yeon Park ◽

Hyung-Sik Kim ◽

Jeong-Seok Nam

Keyword(s):

Stem Cells ◽

Intestinal Epithelium ◽

Fine Tuning ◽

Intestinal Stem Cells ◽

Dynamic Regulation ◽

Intestinal Integrity ◽

Differentiated Cells ◽

Quiescent Stem Cells ◽

G Protein Coupled ◽

Crypt Base

Despite endogenous insults such as mechanical stress and danger signals derived from the microbiome, the intestine can maintain its homeostatic condition through continuous self-renewal of the crypt–villus axis. This extraordinarily rapid turnover of intestinal epithelium, known to be 3 to 5 days, can be achieved by dynamic regulation of intestinal stem cells (ISCs). The crypt base-located leucine-rich repeat-containing G-protein-coupled receptor 5-positive (Lgr5+) ISCs maintain intestinal integrity in the steady state. Under severe damage leading to the loss of conventional ISCs, quiescent stem cells and even differentiated cells can be reactivated into stem-cell-like cells with multi-potency and contribute to the reconstruction of the intestinal epithelium. This process requires fine-tuning of the various signaling pathways, including the Hippo–YAP system. In this review, we summarize recent advances in understanding the correlation between Hippo–YAP signaling and intestinal homeostasis, repair, and tumorigenesis, focusing specifically on ISC regulation.

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Loss of foxo rescues stem cell aging in Drosophila germ line

eLife ◽

10.7554/elife.27842 ◽

2017 ◽

Vol 6 ◽

Cited By ~ 8

Author(s):

Filippo Artoni ◽

Rebecca E Kreipke ◽

Ondina Palmeira ◽

Connor Dixon ◽

Zachary Goldberg ◽

...

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Stem Cell ◽

Ionizing Radiation ◽

Cell Injury ◽

Germ Line ◽

Adult Stem Cell ◽

Cell Aging ◽

Germline Stem Cells ◽

Cell Cycle Reentry

Aging stem cells lose the capacity to properly respond to injury and regenerate their residing tissues. Here, we utilized the ability of Drosophila melanogaster germline stem cells (GSCs) to survive exposure to low doses of ionizing radiation (IR) as a model of adult stem cell injury and identified a regeneration defect in aging GSCs: while aging GSCs survive exposure to IR, they fail to reenter the cell cycle and regenerate the germline in a timely manner. Mechanistically, we identify foxo and mTOR homologue, Tor as important regulators of GSC quiescence following exposure to ionizing radiation. foxo is required for entry in quiescence, while Tor is essential for cell cycle reentry. Importantly, we further show that the lack of regeneration in aging germ line stem cells after IR can be rescued by loss of foxo.

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