Number Of Intestinal Stem Cells Research Articles

Wuliangye strong aroma baijiu promotes intestinal homeostasis by improving gut microbiota and regulating intestinal stem cell proliferation and differentiation.

Wuliangye strong aroma baijiu (hereafter, Wuliangye baijiu) is a traditional Chinese grain liquor containing short-chain fatty acids, ethyl caproate, ethyl lactate, other trace components, and a large proportion of ethanol. The effects of Wuliangye baijiu on intestinal stem cells and intestinal epithelial development have not been elucidated. Here, the role of Wuliangye baijiu in intestinal epithelial regeneration and gut microbiota modulation was investigated by administering a Lieber-DeCarli chronic ethanol liquid diet in a mouse model to mimic long-term (8 weeks') light/moderate alcohol consumption (1.6 g kg-1 day-1) in healthy human adults. Wuliangye baijiu promoted colonic crypt proliferation in mice. According to immunofluorescence and reverse transcription-quantitative polymerase chain reaction analyses, compared with the ethanol-only treatment, Wuliangye baijiu increased the number of intestinal stem cells and goblet cells and the expression of enteroendocrine cell differentiation markers in the mouse colon. Furthermore, gut microbiota analysis showed an increase in the relative abundance of microbiota related to intestinal homeostasis following Wuliangye baijiu administration. Notably, increased abundance of Bacteroidota, Faecalibaculum, Lachnospiraceae, and Blautia may play an essential role in promoting stem-cell-mediated intestinal epithelial development and maintaining intestinal homeostasis. In summary, these findings suggest that Wuliangye baijiu can be used to regulate intestinal stem cell proliferation and differentiation in mice and to alter gut microbiota distributions, thereby promoting intestinal homeostasis. This research elucidates the mechanism by which Wuliangye baijiu promotes intestinal health. © 2024 Society of Chemical Industry.

DHX9 maintains epithelial homeostasis by restraining R-loop-mediated genomic instability in intestinal stem cells

Epithelial barrier dysfunction and crypt destruction are hallmarks of inflammatory bowel disease (IBD). Intestinal stem cells (ISCs) residing in the crypts play a crucial role in the continuous self-renewal and rapid recovery of intestinal epithelial cells (IECs). However, how ISCs are dysregulated in IBD remains poorly understood. Here, we observe reduced DHX9 protein levels in IBD patients, and mice with conditional DHX9 depletion in the intestinal epithelium (Dhx9ΔIEC) exhibit an increased susceptibility to experimental colitis. Notably, Dhx9ΔIEC mice display a significant reduction in the numbers of ISCs and Paneth cells. Further investigation using ISC-specific or Paneth cell-specific Dhx9-deficient mice demonstrates the involvement of ISC-expressed DHX9 in maintaining epithelial homeostasis. Mechanistically, DHX9 deficiency leads to abnormal R-loop accumulation, resulting in genomic instability and the cGAS-STING-mediated inflammatory response, which together impair ISC function and contribute to the pathogenesis of IBD. Collectively, our findings highlight R-loop-mediated genomic instability in ISCs as a risk factor in IBD.

Abstract SY07-02: Cell competition vs cell cooperation: Unlocking oncogenic potential of mutant clones in the gut

Abstract The epithelial monolayer of the intestine is one of the fastest renewing tissues of the human body. Every week, the entire epithelium is replaced by intestinal stem cells (ISCs) that reside in crypt-like structures within the monolayer. Within these crypts, a small number of ISCs continuously divide to provide all specialized cell types necessary to maintain normal intestinal homeostasis, while simultaneously competing with each other for a position within the crypt bottom. This competition, characterized by neutral loss-and-replacement events, can be drastically disturbed when one of the ISCs acquires a mutation that confers a competitive advantage. As a result, mutant ISCs will replace all normal ISCs, colonize the intestinal crypt, and facilitate the initiation of colorectal cancer (CRC). CRC development is a stepwise process characterized by the accumulation of mutations in cancer driver genes over time. Almost 80% of all CRCs present with early mutations in the tumor suppressor gene APC, leading to constitutive activation of the Wnt pathway and the formation of pre-malignant adenomas. We have previously demonstrated that, within individual crypts, Apc-mutant ISCs have a competitive advantage over normal ISCs, although the mechanism underlying this unequal competition remained unclear. Recently, we revealed that Apc-mutant ISCs in fact act as "supercompetitors" by secreting Wnt antagonists such as NOTUM, WIF1 and DKK2, that actively disadvantage neighboring wild type ISCs by forcing them do differentiate. We demonstrated that boosting wild type fitness by downstream pharmacological activation of the Wnt signaling pathway by GSK3β-inhibitor lithium rendered wild type ISCs resistant to the Wnt antagonists, reduced the number of Apc-mutant crypts, and prevented adenoma formation. Together, this work highlights that modulating cell competition between normal and mutant cells within the crypt can provide a novel approach to prevent CRC development. Our findings are particularly relevant for familial adenomatous polyposis (FAP) patients, that carry germline mutations in APC and are predisposed to the development of CRC. Based on our fundamental discoveries, we are currently performing a clinical trial to assess the chemopreventive effect of lithium on the expansion of APC-mutant clones in FAP patients [ClinicalTrials.gov: NCT05402891]. Importantly, it is becoming increasingly evident that intestinal adenomas do not form from a single Apc-mutant crypt, but require fields of multiple mutant crypts to facilitate tumor initiation. Although field cancerization by enhanced crypt fission has been observed for several oncogenic mutants in the gut (e.g. KrasG12D, PIK3CAH1047R), Apc-mutant cells are often contained within individual crypts. This indicates that multiple independent mutant clones may act cooperatively to facilitate tumorigenesis, thereby pointing towards a polyclonal origin of intestinal adenomas. Our current work investigates how Apc-mutant clones interact by modifying their local environment through reciprocal signalling with stromal and epithelial cells to escape the anatomical constraints of the crypt, thereby unlocking their oncogenic potential in the gut. Ultimately, unravelling the molecular mechanisms underlying clonal cooperation will inform potential chemoprevention strategies for CRC. Citation Format: Sanne van Neerven. Cell competition vs cell cooperation: Unlocking oncogenic potential of mutant clones in the gut [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr SY07-02.

The loss of antioxidant activities impairs intestinal epithelium homeostasis by altering lipid metabolism

Reactive oxygens species (ROS) are common byproducts of metabolic reactions and could be at the origin of many diseases of the elderly. Here we investigated the role of ROS in the renewal of the intestinal epithelium in mice lacking catalase (CAT) and/or nicotinamide nucleotide transhydrogenase (NNT) activities. Cat−/− mice have delayed intestinal epithelium renewal and were prone to develop necrotizing enterocolitis upon starvation. Interestingly, crypts lacking CAT showed fewer intestinal stem cells (ISC) and lower stem cell activity than wild-type. In contrast, crypts lacking NNT showed a similar number of ISCs as wild-type but increased stem cell activity, which was also impaired by the loss of CAT. No alteration in the number of Paneth cells (PCs) was observed in crypts of either Cat−/− or Nnt−/− mice, but they showed an evident decline in the amount of lysozyme. Cat deficiency caused fat accumulation in crypts, and a fall in the remarkable high amount of adipose triglyceride lipase (ATGL) in PCs. Notably, the low levels of ATGL in the intestine of Cat −/− mice increased after a treatment with the antioxidant N-acetyl-L-cysteine. Supporting a role of ATGL in the regulation of ISC activity, its inhibition halt intestinal organoid development. These data suggest that the reduction in the renewal capacity of intestine originates from fatty acid metabolic alterations caused by peroxisomal ROS.

Astragalus polysaccharide promotes the regeneration of intestinal stem cells through HIF-1 signalling pathway.

Ionizing radiation (IR)-induced intestinal injury is usually accompanied by high lethality. Intestinal stem cells (ISCs) are critical and responsible for the regeneration of the damaged intestine. Astragalus polysaccharide (APS), one of the main active ingredients of Astragalus membranaceus (AM), has a variety of biological functions. This study was aimed to investigate the potential effects of APS on IR-induced intestine injury via promoting the regeneration of ISCs. We have established models of IR-induced intestinal injury and our results showed that APS played great radioprotective effects on the intestine. APS improved the survival rate of irradiated mice, reversed the radiation damage of intestinal tissue, increased the survival rate of intestinal crypts, the number of ISCs and the expression of intestinal tight junction-related proteins after IR. Moreover, APS promoted the cell viability while inhibited the apoptosis of MODE-K. Through organoid experiments, we found that APS promoted the regeneration of ISCs. Remarkably, the results of network pharmacology, RNA sequencing and RT-PCR assays showed that APS significantly upregulated the HIF-1 signalling pathway, and HIF-1 inhibitor destroyed the radioprotection of APS. Our findings suggested that APS promotes the regeneration of ISCs through HIF-1 signalling pathway, and it may be an effective radioprotective agent for IR-induced intestinal injury.

Control of TGFβ signalling by ubiquitination independent function of E3 ubiquitin ligase TRIP12

Transforming growth factor β (TGFβ) pathway is a master regulator of cell proliferation, differentiation, and death. Deregulation of TGFβ signalling is well established in several human diseases including autoimmune disorders and cancer. Thus, understanding molecular pathways governing TGFβ signalling may help better understand the underlying causes of some of those conditions. Here, we show that a HECT domain E3 ubiquitin ligase TRIP12 controls TGFβ signalling in multiple models. Interestingly, TRIP12 control of TGFβ signalling is completely independent of its E3 ubiquitin ligase activity. Instead, TRIP12 recruits SMURF2 to SMAD4, which is most likely responsible for inhibitory monoubiquitination of SMAD4, since SMAD4 monoubiquitination and its interaction with SMURF2 were dramatically downregulated in TRIP12-/- cells. Additionally, genetic inhibition of TRIP12 in human and murine cells leads to robust activation of TGFβ signalling which was rescued by re-introducing wildtype TRIP12 or a catalytically inactive C1959A mutant. Importantly, TRIP12 control of TGFβ signalling is evolutionary conserved. Indeed, genetic inhibition of Drosophila TRIP12 orthologue, ctrip, in gut leads to a reduced number of intestinal stem cells which was compensated by the increase in differentiated enteroendocrine cells. These effects were completely normalised in Drosophila strain where ctrip was co-inhibited together with Drosophila SMAD4 orthologue, Medea. Similarly, in murine 3D intestinal organoids, CRISPR/Cas9 mediated genetic targeting of Trip12 enhances TGFβ mediated proliferation arrest and cell death. Finally, CRISPR/Cas9 mediated genetic targeting of TRIP12 in MDA-MB-231 breast cancer cells enhances the TGFβ induced migratory capacity of these cells which was rescued to the wildtype level by re-introducing wildtype TRIP12. Our work establishes TRIP12 as an evolutionary conserved modulator of TGFβ signalling in health and disease.

Protective effect of total flavonoids of Engelhardia roxburghiana Wall. leaves against radiation-induced intestinal injury in mice and its mechanism.

Irradiation-induced intestinal injury (RIII) often occurs during radiotherapy in patients, which would result in abdominal pain, diarrhea, nausea, vomiting, and even death. Engelhardia roxburghiana Wall. leaves, a traditional Chinese herb, has unique anti-inflammatory, anti-tumor, antioxidant, and analgesic effects, is used to treat damp-heat diarrhea, hernia, and abdominal pain, and has the potential to protect against RIII. To explore the protective effects of the total flavonoids of Engelhardia roxburghiana Wall. leaves (TFERL) on RIII and provide some reference for the application of Engelhardia roxburghiana Wall. leaves in the field of radiation protection. The effect of TFERL on the survival rate of mice was observed after a lethal radiation dose (7.2 Gy) by ionizing radiation (IR). To better observe the protective effects of the TFERL on RIII, a mice model of RIII induced by IR (13 Gy) was established. Small intestinal crypts, villi, intestinal stem cells (ISC) and the proliferation of ISC were observed by haematoxylin and eosin (H&E) and immunohistochemistry (IHC). Quantitative real-time PCR (qRT-PCR) was used to detect the expression of genes related to intestinal integrity. Superoxide dismutase (SOD), reduced glutathione (GSH), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the serum of mice were assessed. In vitro, cell models of RIII induced by IR (2, 4, 6, 8 Gy) were established. Normal human intestinal epithelial cells HIEC-6 cells were treated with TFERL/Vehicle, and the radiation protective effect of TFERL on HIEC-6 cells was detected by clone formation assay. DNA damage was detected by comet assay and immunofluorescence assay. Reactive oxygen species (ROS), cell cycle and apoptosis rate were detected by flow cytometry. Oxidative stress, apoptosis and ferroptosis-related proteins were detected by western blot. Finally, the colony formation assay was used to detect the effect of TFERL on the radiosensitivity of colorectal cancer cells. TFERL treatment can increase the survival rate and time of the mice after a lethal radiation dose. In the mice model of RIII induced by IR, TFERL alleviated RIII by reducing intestinal crypt/villi structural damage, increasing the number and proliferation of ISC, and maintaining the integrity of the intestinal epithelium after total abdominal irradiation. Moreover, TFERL promoted the proliferation of irradiated HIEC-6 cells, and reduced radiation-induced apoptosis and DNA damage. Mechanism studies have found that TFERL promotes the expression of NRF2 and its downstream antioxidant proteins, and silencing NRF2 resulted in the loss of radioprotection by TFERL, suggesting that TFERL exerts radiation protection by activating the NRF2 pathway. Surprisingly, TFERL reduced the number of clones of colon cancer cells after irradiation, suggesting that TFERL can increase the radiosensitivity of colon cancer cells. Our data showed that TFERL inhibited oxidative stress, reduced DNA damage, reduced apoptosis and ferroptosis, and improved IR-induced RIII. This study may offer a fresh approach to using Chinese herbs for radioprotection.

Restoration of the gut barrier integrity and restructuring of the gut microbiome in aging by angiotensin-(1-7).

Compromised barrier function of colon epithelium with aging is largely due to gut microbial dysbiosis. Recent studies implicate an important role for angiotensin converting enzymes, ACE and ACE2, angiotensins, and the receptors, AT1 receptor (AT1R) and Mas receptor (MasR), in the regulation of colon functions. The present study tested the hypothesis that leaky gut in aging is associated with an imbalance in ACE2/ACE and that the treatment with angiotenisn-(1-7) (Ang-(1-7)) will restore gut barrier integrity and microbiome. Studies were carried out in Young (3-4 months) and old (20-24 months) male mice. Ang-(1-7) was administered by using osmotic pumps. Outcome measures included expressions of ACE, ACE2, AT1R, and MasR, intestinal permeability by using FITC-dextran, and immunohistochemistry of claudin 1 and occludin, and intestinal stem cells (ISCs). ACE2 protein and activity were decreased in Old group while that of ACE were unchanged. Increased intestinal permeability and plasma levels of zonulin-1 in the Old group were normalized by Ang-(1-7). Epithelial disintegrity, reduced number of goblet cells and ISCs in the old group were restored by Ang-(1-7). Expression of claudin 1 and occludin in the aging colon was increased by Ang-(1-7). Infiltration of CD11b+ or F4/80+ inflammatory cells in the old colons were decreased by Ang-(1-7). Gut microbial dysbiosis in aging was evident by decreased richness and altered beta diversity that were reversed by Ang-(1-7) with increased abundance of Lactobacillus or Lachnospiraceae. The present study shows that Ang-(1-7) restores gut barrier integrity and reduces inflammation in the aging colon by restoring the layer of ISCs and by restructuring the gut microbiome.

Regulation of intestinal stem cell activity by a mitotic cell cycle regulator Polo in Drosophila.

Maintaining a definite and stable pool of dividing stem cells plays an important role in organ development. This process requires an appropriate progression of mitosis for proper spindle orientation and polarity to ensure the ability of stem cells to proliferate and differentiate correctly. Polo-like kinases (Plks)/Polo are the highly conserved serine/threonine kinases involved in the initiation of mitosis as well as in the progression of the cell cycle. Although numerous studies have investigated the mitotic defects upon loss of Plks/Polo in cells, little is known about the in vivo consequences of stem cells with abnormal Polo activity in the context of tissue and organism development. The current study aimed to investigate this question using the Drosophila intestine, an organ dynamically maintained by the intestinal stem cells (ISCs). The results indicated that the polo depletion caused a reduction in the gut size due to a gradual decrease in the number of functional ISCs. Interestingly, the polo-deficient ISCs showed an extended G2/M phase and aneuploidy and were subsequently eliminated by premature differentiation into enterocytes (ECs). In contrast, the constitutively active Polo (poloT182D) suppressed ISC proliferation, induced abnormal accumulation of β-tubulin in cells, and drove ISC loss via apoptosis. Therefore, Polo activity should be properly maintained for optimal stem cell function. Further analysis suggested that polo was a direct target gene of Sox21a, a Sox transcription factor that critically regulates stem cell activity. Together, this study provided a novel perspective on the correlation between the progression of mitosis and the ISC function in Drosophila.

Antibiotic-induced microbial depletion enhances murine small intestinal epithelial growth in a serotonin-dependent manner.

Regulation of small intestinal epithelial growth by endogenous and environmental factors is critical for intestinal homeostasis and recovery from insults. Depletion of the intestinal microbiome increases epithelial proliferation in small intestinal crypts, similar to the effects observed in animal models of serotonin potentiation. Based on prior evidence that the microbiome modulates serotonin activity, we hypothesized that microbial depletion-induced epithelial proliferation is dependent on host serotonin activity. A mouse model of antibiotic-induced microbial depletion (AIMD) was employed. Serotonin potentiation was achieved through either genetic knockout of the serotonin transporter (SERT) or pharmacologic SERT inhibition, and inhibition of serotonin synthesis was achieved with para-chlorophenylalanine. AIMD and serotonin potentiation increased intestinal villus height and crypt proliferation in an additive manner, but the epithelial proliferation observed after AIMD was blocked in the absence of endogenous serotonin. Using Lgr5-EGFP-reporter mice, we evaluated intestinal stem cell (ISC) quantity and proliferation. AIMD increased the number of ISCs per crypt and ISC proliferation compared to controls, and changes in ISC number and proliferation were dependent on the presence of host serotonin. Furthermore, western blotting demonstrated that AIMD reduced epithelial SERT protein expression compared to controls. In conclusion, host serotonin activity is necessary for microbial depletion-associated changes in villus height and ISC proliferation in crypts, and microbial depletion produces a functional serotonin-potentiated state through reduced SERT protein expression. These findings provide an understanding of how changes to the microbiome contribute to intestinal pathology and can be applied therapeutically.

Establishment and Evaluation of Intestinal Injury Model of Mouse Acute Graft Versus Host Disease Based on An Organoid Technology

To establish an intestinal organoid model that mimic acute graft versus host disease (aGVHD) caused intestinal injuries by using aGVHD murine model serum and organoid culture system, and explore the changes of aGVHD intestine in vitro by advantage of organoid technology. 20-22 g female C57BL/6 mice and 20-22 g female BALB/c mice were used as donors and recipients for bone marrow transplantation, respectively. Within 4-6 h after receiving a lethal dose (8.0 Gy) of γ ray total body irradiation, a total of 0.25 ml of murine derived bone marrow cells (1×107/mice, n=20) and spleen nucleated cells (5×106/mice, n=20) was infused to establish a mouse model of aGVHD (n=20). The aGVHD mice were anesthetized at the 7th day after transplantation, and the veinal blood was harvested by removing the eyeballs, and the serum was collected by centrifugation. The small intestinal crypts of healthy C57BL/6 mice were harvested and cultivated in 3D culture system that maintaining the growth and proliferation of intestinal stem cells in vitro. In our experiment, 5%, 10%, 20% proportions of aGVHD serum were respectively added into the organoid culture system for 3 days. The formation of small intestinal organoids were observed under an inverted microscope and the morphological characteristics of intestinal organoids in each groups were analyzed. For further evaluation, the aGVHD intestinal organoids were harvested and their pathological changes were observed. Combined with HE staining, intestinal organ morphology evaluation was performed. Combined with Alcian Blue staining, the secretion function of aGVHD intestinal organoids was observed. The distribution and changes of Lgr5+ and Clu+ intestinal stem cells in intestinal organoids were analyzed under the conditions of 5%, 10% and 20% serum concentrations by immunohistochemical stainings. The results of HE staining showed that the integrity of intestinal organoids in the 5% concentration serum group was better than that in the 10% and 20% groups. The 5% concentration serum group showed the highest number of organoids, the highest germination rate and the lowest pathological score among experimental groups, while the 20% group exhibited severe morphological destruction and almost no germination was observed, and the pathological score was the highest among all groups(t=3.668, 4.334,5.309,P<0.05). The results of Alican blue staining showed that the secretion function of intestinal organoids in serum culture of aGVHD in the 20% group was weaker than that of the 5% group and 10% of the organoids, and there was almost no goblet cells, and mucus was stainned in the 20% aGVHD serum group. The immunohistochemical results showed that the number of Lgr5+ cells of intestinal organoids in the 5% group was more than that of the intestinal organoids in the 10% aGVHD serum group and 20% aGVHD serum group. Almost no Clu+ cells were observed in the 5% group. The Lgr5+ cells in the 20% group were seriously injuried and can not be observed. The proportion of Clu+ cells in the 20% group significantly increased. The concentration of aGVHD serum in the culture system can affect the number and secretion function of intestinal organoids as well as the number of intestinal stem cells in organoids. The higher the serum concentration, the greater the risk of organoid injury, which reveal the characteristics of the formation and functional change of aGVHD intestinal organoids, and provide a novel tool for the study of intestinal injury in aGVHD.

Apical-basal polarity precisely determines intestinal stem cell number by regulating Prospero threshold.

Apical-basal polarity and cell-fate determinants are crucial for the cell fate and control of stem cell numbers. However, their interplay leading to a precise stem cell number remains unclear. Drosophila pupal intestinal stem cells (pISCs) asymmetrically divide, generating one apical ISC progenitor and one basal Prospero (Pros)+ enteroendocrine mother cell (EMC), followed by symmetric divisions of each daughter before adulthood, providing an ideal system to investigate the outcomes of polarity loss. Using lineage tracing and exvivo live imaging, we identify an interlocked polarity regulation network precisely determining ISC number: Bazooka inhibits Pros accumulation by activating Notch signaling to maintain stem cell fate in pISC apical daughters. A threshold of Pros promotes differentiation to EMCs and avoids ISC-like cell fate, and over-threshold of Pros inhibits miranda expression to ensure symmetric divisions in pISC basal daughters. Our work suggests that a polarity-dependent threshold of a differentiation factor precisely controls stem cell number.

Piwi maintains homeostasis in the Drosophila adult intestine.

PIWI genes are well known for their germline but not somatic functions. Here, we report the function of the Drosophila piwi gene in the adult gut, where intestinal stem cells (ISCs) produce enteroendocrine cells and enteroblasts that generate enterocytes. We show that piwi is expressed in ISCs and enteroblasts. Piwi deficiency reduced ISC number, compromised enteroblasts maintenance, and induced apoptosis in enterocytes, but did not affect ISC proliferation and its differentiation to enteroendocrine cells. In addition, deficiency of zygotic but not maternal piwi mildly de-silenced several retrotransposons in the adult gut. Importantly, either piwi mutations or piwi knockdown specifically in ISCs and enteroblasts shortened the Drosophila lifespan, indicating that intestinal piwi contributes to longevity. Finally, our mRNA sequencing data implied that Piwi may achieve its intestinal function by regulating diverse molecular processes involved in metabolism and oxidation-reduction reaction.

Heat stress disrupts intestinal stem cell migration and differentiation along the crypt–villus axis through FAK signaling

During heat stress (HS), the intestinal epithelium suffers damage due to imbalance of tissue homeostasis. However, the specific mechanism by which intestinal stem cells (ISCs) migrate and differentiate along the crypt–villus axis to heal lesions upon insult is unclear. In our study, C57BL/6 mice and IPEC-J2 cells were subjected to normal ambient conditions (25 °C for 7 days in vivo and 37 °C for 18 h in vitro) or 41 °C. The results showed that HS impaired intestinal morphology and barrier function. The numbers of ISCs (SOX9+ cells), mitotic cells (PCNA+ cells), and differentiated cells (Paneth cells marked by lysozyme, absorptive cells marked by Villin, goblet cells marked by Mucin2, enteroendocrine cells marked by Chromogranin A, and tuft cells marked by DCAMKL1) were reduced under high temperature. Importantly, BrdU incorporation confirmed the decreased migration ability of jejunal epithelial cells exposed to 41 °C. Furthermore, intestinal organoids (IOs) expanded from jejunal crypt cells in the HS group exhibited greater growth disadvantages. Mechanistically, the occurrence of these phenotypes was accompanied by FAK/paxillin/F-actin signaling disruption in the jejunum. The fact that the FAK agonist ZINC40099027 reversed the HS-triggered inhibition of IPEC-J2 cell differentiation and migration further confirmed the dominant role of FAK in response to high-temperature conditions. Overall, the present investigation is the first to reveal a major role of FAK/paxillin/F-actin signaling in HS-induced ISC migration and differentiation along the crypt–villus axis, which indicates a new therapeutic target for intestinal epithelial regeneration after heat injuries.

NMN ameliorated radiation induced damage in NRF2-deficient cell and mice via regulating SIRT6 and SIRT7.

Risk of cancer often increases with aging, and radiotherapy is an essential component of treatment. As for abdominal and pelvic cancer, radiotherapy always inevitably causes injury to intestines through direct DNA damage or overload of reactive oxygen species (ROS). Nuclear factor erythroid 2-related factor 2 (NRF2) has been identified as a key protective factor against ionizing-radiation induced damage through promoting DNA damage repair and antioxidant modulation. However, the level of NRF2 always decreases with aging. Here, we demonstrated that NRF2 deficiency aggravated cellular DNA damage and the intestinal pathological lesion. Overexpression of SIRT6 or SIRT7 could improve cell proliferation and protect against radiation injury in NRF2 knock-out (KO) cells by modulating oxidative-stress and DNA damage repair. Consistently, supplement of nicotinamide mononucleotide (NMN), the agonist of sirtuins, increased the level of SIRT6 and SIRT7 in NRF2 KO cells, concomitant with reduced cellular ROS level and ameliorated DNA damage. In vivo, long-term oral administration of NMN attenuated the radiation-induced injury of jejunum, increased the number of intestinal stem cells, and promoted the ability of intestinal proliferation in NRF2-/- mice. Together, our results indicated that SIRT6 and SIRT7 had involved in scavenging ROS and repairing DNA damage, and NMN could be a promising candidate for preventing radiation damage when NRF2 is lacking.

Zymosan-A promotes the regeneration of intestinal stem cells by upregulating ASCL2

Intestinal stem cells (ISCs) are responsible for intestinal tissue homeostasis and are important for the regeneration of the damaged intestinal epithelia. Through the establishment of ionizing radiation (IR) induced intestinal injury model, we found that a TLR2 agonist, Zymosan-A, promoted the regeneration of ISCs in vivo and in vitro. Zymosan-A improved the survival of abdominal irradiated mice (81.82% of mice in the treated group vs. 30% of mice in the PBS group), inhibited the radiation damage of intestinal tissue, increased the survival rate of intestinal crypts and the number of ISCs after lethal IR in vivo. Through organoid experiments, we found that Zymosan-A promoted the proliferation and differentiation of ISCs after IR. Remarkably, the results of RNA sequencing and Western Blot (WB) showed that Zymosan-A reduced IR-induced intestinal injury via TLR2 signaling pathway and Wnt signaling pathway and Zymosan-A had no radioprotection on TLR2 KO mice, suggesting that Zymosan-A may play a radioprotective role by targeting TLR2. Moreover, our results revealed that Zymosan-A increased ASCL2, a transcription factor of ISCs, playing a core role in the process of Zymosan-A against IR-induced intestinal injury and likely contributing to the survival of intestinal organoids post-radiation. In conclusion, we demonstrated that Zymosan-A promotes the regeneration of ISCs by upregulating ASCL2.

Back Cover: Modification of Diet to Reduce the Stemness and Tumorigenicity of Murine and Human Intestinal Cells

Mol. Nutr. Food Res. 2022, 66, 202200234 DOI: 10.1002/mnfr.202200234 Black raspberries (BRBs) play a role in colorectal cancer chemoprevention. The number of intestinal stem cells, the cells of origin of colorectal cancer, are reduced following a BRB diet. This decreases the chance of a mutation occurring within these stem cells and subsequently reduces the risk of developing colorectal cancer. This is reported by Stephanie May, Lee Parry, and co-workers in article number 2200234.

Lymphatics and fibroblasts support intestinal stem cells in homeostasis and injury.

Lgr5+ intestinal stem cells (ISCs) depend on niche factors for their proper function. However, the source of these ISC niche factors and how they support ISCs invivo remain controversial. Here, we report that ISCs depend on lymphatic endothelial cells (LECs) and RSPO3+GREM1+ fibroblasts (RGFs). In the intestine and colon, LECs are surrounded by RGFs and are located near ISCs at the crypt base. Both LECs and RGFs provide the critical ISC niche factor RSPO3 to support ISCs, where RSPO3 loss in both cell types drastically compromises ISC numbers, villi length, and repair after injury. In response to injury, LEC and RGF numbers expand and produce greater amounts of RSPO3 and other growth/angiocrine factors to foster intestinal repair. We propose that LECs represent a novel niche component for ISCs, which together with RGFs serve as the major invivo RSPO3 source for ISCs in homeostasis and injury-mediated regeneration.

Metformin Protects the Intestinal Barrier by Activating Goblet Cell Maturation and Epithelial Proliferation in Radiation-Induced Enteropathy.

Radiotherapy or accidental exposure to high-dose radiation can cause severe damage to healthy organs. The gastrointestinal (GI) tract is a radiation-sensitive organ of the body. The intestinal barrier is the first line of defense in the GI tract, and consists of mucus secreted by goblet cells and a monolayer of epithelium. Intestinal stem cells (ISCs) help in barrier maintenance and intestinal function after injury by regulating efficient regeneration of the epithelium. The Wnt/β-catenin pathway plays a critical role in maintaining the intestinal epithelium and regulates ISC self-renewal. Metformin is the most widely used antidiabetic drug in clinical practice, and its anti-inflammatory, antioxidative, and antiapoptotic effects have also been widely studied. In this study, we investigated whether metformin alleviated radiation-induced enteropathy by focusing on its role in protecting the epithelial barrier. We found that metformin alleviated radiation-induced enteropathy, with increased villi length and crypt numbers, and restored the intestinal barrier function in the irradiated intestine. In a radiation-induced enteropathy mouse model, metformin treatment increased tight-junction expression in the epithelium and inhibited bacterial translocation to mesenteric lymph nodes. Metformin increased the number of ISCs from radiation toxicity and enhanced epithelial repair by activating Wnt/β-catenin signaling. These data suggested that metformin may be a potential therapeutic agent for radiation-induced enteropathy.

A non-dividing cell population with high pyruvate dehydrogenase kinase activity regulates metabolic heterogeneity and tumorigenesis in the intestine

Although reprogramming of cellular metabolism is a hallmark of cancer, little is known about how metabolic reprogramming contributes to early stages of transformation. Here, we show that the histone deacetylase SIRT6 regulates tumor initiation during intestinal cancer by controlling glucose metabolism. Loss of SIRT6 results in an increase in the number of intestinal stem cells (ISCs), which translates into enhanced tumor initiating potential in APCmin mice. By tracking down the connection between glucose metabolism and tumor initiation, we find a metabolic compartmentalization within the intestinal epithelium and adenomas, where a rare population of cells exhibit features of Warburg-like metabolism characterized by high pyruvate dehydrogenase kinase (PDK) activity. Our results show that these cells are quiescent cells expressing +4 ISCs and enteroendocrine markers. Active glycolysis in these cells suppresses ROS accumulation and enhances their stem cell and tumorigenic potential. Our studies reveal that aerobic glycolysis represents a heterogeneous feature of cancer, and indicate that this metabolic adaptation can occur in non-dividing cells, suggesting a role for the Warburg effect beyond biomass production in tumors.