Ionizing radiation (IR) induced damages are common complications of radiotherapy for tumors, severely limiting the intensity and therapeutic efficacy of the radiotherapy program. Emerging data indicated that the cGAS-STING pathway has paradoxical effects on IR-induced damage. SR-717, as a non-nucleotide, small-molecule stimulator of interferon genes (STING) agonist, has been proven that it could activate the STING signaling pathway. In this work, we try to explore the radioprotection of the STING signaling pathway and figure out whether SR-717 could be a potential intestinal radioprotective agent. C57BL/6 mice were intraperitoneally treated with SR-717 or normal saline (NS). By analyzing the survival rate, body weight, and the number of peripheral blood cells after IR exposure, we found that SR-717 improved the survival rate and body weight of mice, protected the intestine from IR-induced damage as well as hematopoietic damage, and promoted the regeneration of intestinal stem cells (ISCs). Cell viability and apoptosis after irradiation were detected after stimulation of MODE-K cells with SR-717 or PBS. We found that SR-717 increased cell viability and inhibited apoptosis in vitro. The mechanism of SR-717 in intestinal radiation protection was investigated by RNA-seq. The results of RNA-seq and qRT-PCR suggested that SR-717 significantly activated the immune system via the STING-IL-6 signaling pathway. In addition, we discussed the role of TLR2 in SR-717-mediated anti-radiation activity, and TLR2 deletion significantly reversed the radioprotective effects of SR717. In conclusion, we proved STING signaling activation displayed anti-radiation activity and found SR-717 displayed anti-radiation activity via the STING-IL-6 signaling pathway, suggesting SR-717 could be a potential intestinal radioprotective agent.

The gastrointestinal (GI) tract constitutes a sophisticated system integral to digestion, nutrient absorption, and overall health, with its functionality predominantly hinging on the distinctive properties of diverse stem cell types. This review systematically investigates the pivotal roles of stem cells across the esophagus, stomach, small intestine, and colon, emphasizing their crucial contributions to tissue homeostasis, repair mechanisms, and regeneration. Each segment of the GI tract is characterized by specialized stem cell populations that exhibit distinct functional attributes, highlighting the necessity for tailored therapeutic approaches in the management of gastrointestinal disorders.Emerging research has shed light on the functional heterogeneity of GI stem cells, with ISCs in the small intestine displaying remarkable turnover rates and regenerative potential, whereas colonic stem cells (CSCs) are essential for the preservation of the colonic epithelial barrier. The intricate interplay between stem cells and their microenvironment-or niche-is fundamentally important for their functionality, with critical signaling pathways such as Wnt and Notch exerting substantial influence over stem cell behavior. The advent of organoid models derived from GI stem cells offers promising avenues for elucidating disease mechanisms and for the preclinical testing of novel therapeutic interventions.Despite notable advancements in foundational research on GI stem cells, the translation of these scientific discoveries into clinical practice remains limited. As of 2025, Japan's clinical GI disease guidelines do not endorse any stem cell-based therapies, underscoring the existing disconnect between research findings and clinical application. This scenario accentuates the urgent need for sustained efforts to bridge this divide and to cultivate innovative strategies that synergize stem cell technology with conventional treatment modalities.Future investigations should be directed toward unraveling the mechanisms that underpin stem cell dysfunction in various gastrointestinal pathologies, as well as exploring combination therapies that harness the regenerative capacities of stem cells in conjunction with immunomodulatory treatments. By fostering collaborative endeavors between basic researchers and clinical practitioners, we can deepen our understanding of GI stem cells and facilitate the translation of this knowledge into effective therapeutic interventions, ultimately enhancing patient outcomes in gastrointestinal diseases.

Acute radiation gastrointestinal syndrome (GI-ARS) develops when the intestine is rapidly exposed to large doses of ionizing radiation. In humans, GI-ARS occurs at radiation doses of 6 Gy, with doses of ≥10 Gy typically resulting in death within 10 days. This condition can be caused by various factors, including war, terrorism, nuclear power plant accidents, and cancer therapy-associated adverse events. Developing effective approaches for treating GI-ARS requires a comprehensive understanding of the syndrome. This review summarizes the current body of literature that defines GI-ARS as a consequence of intestinal irradiation. It highlights the paradigm shift in understanding which intestinal stem cells contribute to homeostasis, the critical role of vascular injury in the development of GI-ARS, and recent advances in research on crypt-villus regeneration following radiation injury.

Impaired colonic mucosal repair is a critical issue in inflammatory bowel diseases (IBD). SATB2 is essential for maintaining colonic epithelial homeostasis, but its role in mucosal repair is unclear. In this study, flow cytometry was used to assess SATB2's role in colonic epithelial repair in a radiation injury model. SATB2 knockout mice exhibited defective epithelial repair, with a marked reduction in goblet and enteroendocrine cells. Mechanistically, SATB2 directly regulated PPAR-γ transcription, and PYY was observed to translocate into the nucleus and promote the transcription of PPAR-γ target genes. In organoids derived from patients with Crohn's disease, PYY supplementation significantly improved epithelial regeneration, outperforming the PPAR-γ agonist rosiglitazone. In conclusion, SATB2 deficiency impairs colonic epithelial repair, which can be rescued by PYY through activation of PPAR-γ-dependent transcription. These findings suggest that PYY may serve as a promising therapeutic molecule to promote epithelial repair in IBD.

Intestinal stem cell (ISC) signaling maintains the balance of self-renewal and differentiation. The role of phosphatidylinositol 3-kinase (PI3K) signaling in ISC responses to radiation was interrogated using Villin-Cre pik3r1lox/lox (p85ΔIEC) mice and p85α-deficient human enteroids (shp85α). Lethal whole-body irradiation in mice was performed to monitor PI3K-mediated survival responses. Rectal biopsies from patients with radiation proctitis were examined by immunohistochemistry for the PI3K/Akt- and Wnt-target survivin. The intestinal epithelial cells (IECs) from p85ΔIEC mice showed increased protein levels of phosphorylated phosphatase and tensin homolog, phosphorylated AktSer473, survivin, cyclin D1, and ρ-β-cateninSer552, as well as increased mRNA for ISC/progenitor cell. In situ hybridization showed that enhanced PI3K signaling reduced Lgr5+ cells but expansion of Axin2+ cells. The shp85α enteroids showed increased mRNA expression of Wnt targets and transcription factor ASCL2, needed for dedifferentiation-mediated restoration of ablated ISCs. The p85α-deficient enteroids showed reduced HES1 mRNA and increases in secretory (ATOH1/MATH1) signaling determinants GFI1 and SPDEF, indicative of reduced NOTCH signaling. Seahorse analyses and phosphorylated p38 staining in IECΔp85 mice indicated that enhanced PI3K signaling led to increased IEC mitochondrial respiration and reactive oxygen species generation. Expression of survivin correlated with the radiation injury in patients. The current data indicate that PI3K signaling increases mitochondrial reactive oxygen species generation and ISC activation that improves IEC recovery from radiation-induced injury. The results suggest that increasing PI3K signaling and induced mitochondrial respiration may improve mucosal healing from radiation injury in patients.

Radiation-induced Intestinal Injury (RIII) affects quality of life in radiotherapy patients; Liangxue Guyuan Yishen Decoction (LGYD) offers protection but requires further study on its mechanism.

The aim of this study was to investigate the heterogeneity of cellular responses in the intestine at a single-cell level following radiation and LGYD treatment.

This study's design includes in vivo and in vitro assessments to evaluate LGYD's effects on intestinal cells post-radiation, targeting survival, recovery, and molecular pathways.

Mice were categorized into four groups: LGYD group, NC group, IR group, and Am group. Each group received daily drug administrations. All groups, except for the NC group, were subjected to a single whole-body irradiation at a dose rate of 70 R/min with a source-to-skin distance of 250 cm. Subsequent experiments were conducted following the irradiation, which led to severe survival impairments in the mice.

Our findings demonstrate that LGYD intervention substantially improves survival rates following lethal doses (8.5 Gy, 70R/min) of whole-body irradiation. Moreover, LGYD expedites the recovery period for intestinal injury on the fifth day after radiation by promoting repair mechanisms within intestinal tissue, with particular focus on mitigating intestinal stem cells (ISCs) damage and immune disorders. Through both in vivo and in vitro experiments, we have discovered that LGYD effectively treats RIII by activating Hes1 transcription factor activity through its key active ingredients in drug-containing serum. This activation further upregulates the downstream Stat3 and Akt gene, thereby facilitating repair processes within intestinal stem cells.

In this study, we discovered that LGYD can enhance the downstream expression and phosphorylation pathways of Stat3 and Akt by upregulating the expression of Hes1 gene following high-dose radiation exposure.

Collagenase and dispase treatment of intestinal tissue from adult mice generates cells growing in matrigel as stably replatable cystic spheroids, in addition to differentiated organoids. Contrary to classical EDTA-derived organoids, these spheroids display poor intestinal differentiation and grow independently of Rspondin, noggin and EGF. Their transcriptome strikingly resembles that of fetal intestinal spheroids, with downregulation of crypt base columnar cell (CBC) markers (Lgr5, Ascl2, Smoc2 and Olfm4). In addition, they display upregulation of inflammatory and mesenchymal genetic programs, together with robust expression of YAP target genes. Lineage tracing, cell-sorting and single cell RNA sequencing experiments demonstrate that adult spheroid-generating cells belong to a hitherto undescribed developmental lineage, independent of Lgr5-positive CBCs, and are involved in regeneration of the epithelium following CBC ablation.

This study aimed to establish an organoid culture model using small intestine tissues from male and female C57BL/6 mice and to compare it with rat organoid cultures derived from frozen tissues. Crypts were isolated from the small intestines of eight-week-old male and female mice and cultured in 3D extracellular matrix with Wnt, R-spondin, and Noggin. In addition, small intestine tissues from sixteen-week-old F344 rats were preserved in a storage solution immediately post-sacrifice and stored at -80°C before being transferred to a nitrogen tank. Upon thawing, crypts from frozen rat tissues failed to develop into organoids due to structural damage, suggesting the need for fresh tissues or optimized preservation methods. In contrast, mouse-derived organoids showed viability for 7 days, with distinct morphological changes and clear differentiation by Day 7. Quantitative real-time PCR analysis revealed that Lgr5, a stem cell marker, showed significantly higher expression in organoids than in tissues, confirming the successful establishment of the organoid culture. Among epithelial markers, the antimicrobial enzyme Lyz1 was more highly expressed in organoids, while Muc2, a key goblet cell marker, was more highly expressed in male tissues. The enterocyte marker Alp exhibited higher expression in male organoids compared to females, with no sex differences in tissues. These findings highlight sex-specific differences in gene expression related to small intestine differentiation and demonstrate the challenges in organoid culture from frozen rat tissues. The results suggest the importance of immediate tissue processing or improved preservation methods for successful organoid cultures.

Multiple next-generation molecules targeting estrogen receptor α (ERα) are being investigated in breast cancer clinical trials, encompassing thousands of women globally. Development of these molecules was partly motivated by the discovery of resistance-associated mutations in ESR1 (encodes ERα). Here, we studied the impact of ERα antagonist/degraders against Esr1 mutations expressed in mouse mammary glands. Inhibition of mutant ERα induced mixed-lineage cells, characterized by aberrant co-engagement of normally disparate master transcription factors. Lineage infidelity was also observed in Esr1-wild-type mice upon long-term estrogen deprivation. In ER+ breast cancer biopsy specimens, heavily pretreated tumors with no ESR1 mutation detected (NMD) frequently exhibited mixed-lineage features. ESR1-mutant tumors generally retained luminal features and higher ERα activity and exhibited an anti-proliferative response to the ERα antagonist giredestrant. ESR1-mutant tumors acquired mixed-lineage features following treatment. Lineage heterogeneity in advanced ER+ breast cancer may underpin the differential benefit of investigational ERα therapeutics observed in ESR1-mutant versus NMD contexts.

Plasticity is needed during development and homeostasis to generate diverse cell types from stem and progenitor cells. Following differentiation, plasticity must be restricted in specialized cells to maintain tissue integrity and function. For this reason, specialized cell identity is stable under homeostatic conditions; however, cells in some tissues regain plasticity during injury-induced regeneration. While precise gene expression controls these processes, the regulatory mechanisms that restrict or promote cell plasticity are poorly understood. Here we use the mouse small intestine as a model system to study cell plasticity. We find that H3K36 methylation reinforces expression of cell-type-associated genes to maintain specialized cell identity in intestinal epithelial cells. Depleting H3K36 methylation disrupts lineage commitment and activates regenerative gene expression. Correspondingly, we observe rapid and reversible remodelling of H3K36 methylation following injury-induced regeneration. These data suggest a fundamental role for H3K36 methylation in reinforcing specialized lineages and regulating cell plasticity and regeneration.

Leucine-rich repeat-containing G-protein-coupled receptors regulate stem cell activity and tissue homeostasis within female reproductive organs, primarily through their interaction with the Wnt/β-catenin signaling pathway. LGR4-6 are increasingly recognized for their roles in organ development, regeneration, and cancer. This review aims to provide a comprehensive overview of the roles of LGR4-6 in female reproductive organs, highlighting their significance in normal physiology and disease states, specifically in the context of ovarian cancer. LGR4 is essential for the proper development of the female reproductive system; its deficiency leads to significant reproductive abnormalities, including delayed menarche and follicle development issues. LGR5 is a well-established marker of stem cells in the ovary and fallopian tubes. It has been implicated in the pathogenesis of high-grade serous ovarian cancer. LGR6, while less studied, shares functional similarities with LGR5 and can maintain stemness. It contributes to chemoresistance in ovarian cancer. LGR6 is a marker for fallopian tube stem cells and is involved in stem cell maintenance and differentiation. LGR4-6 regulate the pathophysiology of female reproductive tissues. LGR4-6 are promising therapeutic targets for treating reproductive cancers and other related disorders. Molecular mechanisms underlying the functions of LGR4-6 should be studied.

Tissue damage and repair are hallmarks of inflammation. Despite a wealth of information on the mechanisms that govern tissue damage, mechanistic insight into how inflammation affects repair is lacking. Here, we investigated how interferons influence tissue repair after damage to the intestinal mucosa. We found that type III, not type I or type II, interferons delay epithelial cell regeneration by inducing the upregulation of Z-DNA-binding protein 1 (ZBP1). Z-nucleic acids formed following intestinal damage are sensed by ZBP1, leading to caspase-8 activation and the cleavage of gasdermin C (GSDMC). Cleaved GSDMC drives epithelial cell death by pyroptosis and delays repair of the large or small intestine after colitis or irradiation, respectively. The type III interferon/ZBP1/caspase-8/GSDMC axis is also active in patients with inflammatory bowel disease (IBD). Our findings highlight the capacity of type III interferons to delay gut repair, which has implications for IBD patients or individuals exposed to radiation therapies.

The increasing use of radiation presents a risk of radiation exposure, making the development of radioprotectors necessary. In the previous study, it is investigated that Deinococcus radiodurans (R1-EVs) exert the antioxidative properties. However, the radioprotective activity of R1-EVs remains unclear. In the present study, the protective effects of R1-EVs against total body irradiation (TBI)-induced acute radiation syndrome (ARS) are investigated. To assess R1-EVs' radioprotective efficacy, ARS is induced in mice with 8 Gy of TBI, and protection against hematopoietic (H)- and gastrointestinal (GI)-ARS is evaluated. The survival rate of irradiated mice group decreases substantially after irradiation. In contrast, pretreatment with R1-EVs increases the survival rates of the mice. The administration of R1-EVs provides effective protection against radiation-induced death of bone marrow cells and splenocytes by scavenging reactive oxygen species (ROS). Additionally, R1-EVs protect both intestinal stem and epithelial cells from radiation-induced apoptosis. R1-EVs stimulate the production of short-chain fatty acids in the gastrointestinal tract, suppress proinflammatory cytokines, and increase regulatory T cells in pretreated mice versus the irradiation-only group. Proteomic analysis shows that the R1-EV proteome is significantly enriched with proteins involved in oxidative stress response. These findings highlight R1-EVs as potent radioprotectors with applications against radiation damage and ROS-mediated diseases.

Mucosal healing is associated with better clinical outcomes in patients with inflammatory bowel disease. But the epithelial-specific contribution to mucosal healing in vivo is poorly understood. We evaluated mucosal healing in an acute dextran sulfate sodium mouse model that shows an alleviated colitis response after epithelial-specific loss of Smad4. We find that enhanced epithelial wound healing alleviates the fibrotic response. Dextran sulfate sodium caused increased mesenchymal collagen deposition-indicative of fibrosis-within a week in the WT but not in the Smad4 KO colon. The fibrotic response correlated with decreased epithelial proliferation in the WT, whereas uninterrupted proliferation and an expanded zone of proliferation were observed in the Smad4 KO colon epithelium. Furthermore, the Smad4 KO colon showed epithelial extracellular matrix alterations that promote epithelial regeneration. Our data suggest that epithelium is a key determinant of the mucosal healing response in vivo, implicating mucosal healing as a strategy against fibrosis in inflammatory bowel disease patients.

Progenitors and mature cells can maintain the intestinal epithelium by dedifferentiation and facultative intestinal stem cell (fISC) function when active ISCs (aISCs) are lost to damage. Here, we modeled fISC activation in mouse intestinal organoids with doxorubicin (DXR) treatment, a chemotherapeutic known to ablate Lgr5+ aISCs in vivo. Similar fISC gene activation was observed between organoids treated with low versus high DXR, despite significantly decreased survival at the higher dose. aISCs exhibited dose-dependent loss after DXR treatment but survived at doses compatible with organoid survival. We ablated residual aISCs after DXR treatment using a Lgr52A-DTR allele and observed that aISC survival of the initial genotoxic insult is required for organoid survival following DXR treatment. These results suggest that although typical fISC genes are activated by DXR-induced injury in organoids, functional stemness remains dependent on the aISC pool. Finally, we show that human intestinal organoids require higher doses of DXR to induce loss of survival and downregulation of LGR5. Our data establish a reproducible model of DXR-induced injury in intestinal organoids and reveal differences in in vitro responses to an established in vivo damage modality.

Lgr5+ intestinal stem cells (ISCs) are crucial for the intestinal epithelium renewal and regeneration after injury. However, the mechanism underlying the interplay between Wnt and BMP signaling in this process is not fully understood. Here we report that Bcl11b, which is downregulated by BMP signaling, enhances Wnt signaling to maintain Lgr5+ ISCs and thus promotes the regeneration of the intestinal epithelium upon injury. Loss of Bcl11b function leads to a significant decrease of Lgr5+ ISCs in both intestinal crypts and cultured organoids. Mechanistically, BMP suppresses the expression of Bcl11b, which can positively regulate Wnt target genes by inhibiting the function of the Nucleosome Remodeling and Deacetylase (NuRD) complex and facilitating the β-catenin-TCF4 interaction. Bcl11b can also promote intestinal epithelium repair after injuries elicited by both irradiation and DSS-induced inflammation. Furthermore, Bcl11b deletion prevents proliferation and tumorigenesis of colorectal cancer cells. Together, our findings suggest that BMP suppresses Wnt signaling via Bcl11b regulation, thus balancing homeostasis and regeneration in the intestinal epithelium.

Radiation accidents and misuse of nuclear weapons elevate the risk of development of acute life-threatening injuries as well as their late effects are noted in survivors. Currently, no countermeasure agents are available for the management of radiation-induced GI injury (RIGI) in humans. In the present study, the radiomitigative potential of Trichostatin A (TSA) was evaluated against acute and late RIGI.

15 Gy gamma radiation was delivered to the whole abdomen of C57BL/6 mice, followed by intravenous TSA (150 ng/kg) administration after 1 h and 24 h. Acute changes were checked 24 h and 3.5 days post irradiation. Mice were monitored for development of fibrosis, survival for 1 year and alteration in different signaling pathways.

15 Gy abdominal irradiation activated the DNA damage marker (γ-H2AX) by nearly 3.2 ± 0.29 fold and regulated the repair proteins, XRCC1 and PARP1 in the intestine, which was differentially regulated by TSA. The Wnt signaling pathway and stem cell proliferation in the intestine were also positively regulated by TSA. The TSA administered mice demonstrated improved intestinal morphology. 12.5% of TSA administered mice survived upto 1 year whereas 100% of 15 Gy exposed mice died by 6 months. The surviving mice that had received TSA showed reduced intestinal fibrosis than 15 Gy group, possibly via downregulation of TGFβ/Smad signaling.

The findings suggest that TSA have the potential to mitigate both acute and late effects of radiation in the intestine and can be explored as promising agent in the management of RIGI.

The study of intestinal stem cells is a prerequisite for the development of therapies aimed at regenerating the gut. To enable investigation of adult slow-cycling H2B-GFP-retaining putative small intestinal (SI) stem cells in vitro, we have developed a three-dimensional (3D) SI organoid culture model based on the Tet-Op histone 2 B (H2B)-green fluorescent protein (GFP) fusion protein (Tet-Op-H2B-GFP) transgenic mouse. SI crypts were isolated from 6- to 12-week-old Tet-Op-H2B-GFP transgenic mice and cultured with appropriate growth factors and an animal-derived matrix (Matrigel). For in vitro transgene expression, doxycycline was added to the culture medium for 24 h. By pulse-chase experiments, H2B-GFP expression and retention were assessed through direct GFP fluorescence observations, both by confocal and fluorescence microscopy and by immunohistochemistry. The percentages of H2B-GFP-retaining putative SI stem cells and H2B-GFP-retaining Paneth cells persisting in organoids were determined by scoring relevant GFP-positive cells. Our results indicate that 24 h exposure to doxycycline (pulse) induced ubiquitous expression of H2B-GFP in the SI organoids. During subsequent culture, in the absence of doxycycline (chase), there was a gradual loss (due to cell division) of H2B-GFP. At 6-day chase, slow-cycling H2B-GFP-retaining putative SI stem cells and H2B-GFP-retaining Paneth cells were detected in the SI organoids. The developed culture model allows detection of slow-cycling H2B-GFP-retaining putative SI stem cells and will enable the study of self-renewal and regeneration for further characterization of these cells.

Radiation injury to the intestine is one of the most common complications in patients undergoing abdominal or pelvic cavity radiotherapy, limiting the clinical application of this treatment. Evidence shows the potential benefits of dietary restriction in improving metabolic profiles and age-related diseases. The present study investigated the effects and mechanisms of dietary restriction in radiation-induced intestinal injury. The mice were randomly divided into the control group, 10 Gy total abdominal irradiation (TAI) group, and groups pretreated with 30% caloric restriction (CR) for 7 days or 24 h fasting before TAI. After radiation, the mice were returned to ad libitum. The mice were sacrificed 3.5 days after radiation, and tissue samples were collected. CR and fasting reduced radiation-induced intestinal damage and promoted intestinal recovery by restoring the shortened colon length, improving the impaired intestinal structure and permeability, and remodeling gut microbial structure. CR and fasting also significantly reduced mitochondrial damage and DNA damage, which in turn reduced activation of the cyclic GMP-AMP synthase/stimulator of interferon gene (cGAS/STING) pathway and the production of type I interferon and other chemokines in the jejunum. Since the cGAS/STING pathway is linked with innate immunity, we further showed that CR and fasting induced polarization to immunosuppressive M2 macrophage, decreased CD8+ cytotoxic T lymphocytes, and downregulated proinflammatory factors in the jejunum. Our findings indicated that CR and fasting alleviate radiation-induced intestinal damage by reducing cGAS/STING-mediated harmful immune responses.

Metabolism serves not only as the organism's energy source but also yields metabolites crucial for maintaining tissue homeostasis and overall health. Intestinal stem cells (ISCs) maintain intestinal homeostasis through continuous self-renewal and differentiation divisions. The intricate relationship between metabolic pathways and intestinal homeostasis underscores their crucial interplay. Metabolic pathways have been shown to directly regulate ISC self-renewal and influence ISC fate decisions under homeostatic conditions, but the cellular and molecular mechanisms remain incompletely understood. Understanding the intricate involvement of various pathways in maintaining intestinal homeostasis holds promise for devising innovative strategies to address intestinal diseases. Here, we provide a comprehensive review of recent advances in the regulation of intestinal homeostasis. We describe the regulation of intestinal homeostasis from multiple perspectives, including the regulation of intestinal epithelial cells, the regulation of the tissue microenvironment, and the key role of nutrient metabolism. We highlight the regulation of intestinal homeostasis and ISC by nutrient metabolism. This review provides a multifaceted perspective on how intestinal homeostasis is regulated and provides ideas for intestinal diseases and repair of intestinal damage.

Cancer Stem Cells (CSCs) are highly tumorigenic, chemoresistant, and immune evasive. They emerge as a central driver that gives rise to the bulk of tumoral mass, modifies the tumor microenvironment (TME), and exploits it, leading to poor clinical outcomes for patients with cancer. The existence of CSCs thus accounts for the failure of conventional therapies and immune surveillance. Identifying CSCs in solid tumors remains a significant challenge in modern oncology, with the use of cell surface markers being the primary strategy for studying, isolating, and enriching these cells. In this review, we explore CSC markers, focusing on the underlying signaling pathways that drive CSC self-renewal, which simultaneously makes them intrinsically chemoresistant and immune system evaders. We comprehensively discuss the autonomous and non-autonomous functions of CSCs, with particular emphasis on their interactions with the tumor microenvironment, especially immune cells. This reciprocal network enhances CSCs malignancy while compromising the surrounding niche, ultimately defining therapeutic vulnerabilities associated with each CSC marker. The most common CSCs surface markers addressed in this review-CD44, CD133, ICAM1/CD54, and LGR5-provide insights into the interplay between chemoresistance and immune evasion, two critically important phenomena in disease eradication. This new perspective on the state-of-the-art of CSCs will undoubtedly open new avenues for therapy.

Radiotherapy is a pivotal intervention for cancer patients, significantly impacting their treatment outcomes and survival prospects. Nevertheless, in the course of treating those with abdominal, pelvic, or retroperitoneal malignant tumors, the procedure inadvertently exposes adjacent intestinal tissues to radiation, posing risks of radiation-induced enteropathy upon reaching threshold doses. Stem cells within the intestinal crypts, through their controlled proliferation and differentiation, support the critical functions of the intestinal epithelium, ensuring efficient nutrient absorption while upholding its protective barrier properties. Intestinal stem cells (ISCs) regulation is intricately orchestrated by diverse signaling pathways, among which are the WNT, BMP, NOTCH, EGF, Hippo, Hedgehog and NF-κB, each contributing to the complex control of these cells' behavior. Complementing these pathways are additional regulators such as nutrient metabolic states, and the intestinal microbiota, all of which contribute to the fine-tuning of ISCs behavior in the intestinal crypts. It is the harmonious interplay among these signaling cascades and modulating elements that preserves the homeostasis of intestinal epithelial cells (IECs), thereby ensuring the gut's overall health and function. This review delves into the molecular underpinnings of how stem cells respond in the context of radiation enteropathy, aiming to illuminate potential biological targets for therapeutic intervention. Furthermore, we have compiled a summary of several current treatment methodologies. By unraveling these mechanisms and treatment methods, we aspire to furnish a roadmap for the development of novel therapeutics, advancing our capabilities in mitigating radiation-induced intestinal damage.

Vasoactive intestinal peptide (VIP) is a neuronal peptide with prominent distribution along the enteric nervous system. While effects of VIP on intestinal motility, mucosal vasodilation, secretion, and mucosal immune cell function are well-studied, the direct impact of VIP on intestinal epithelial cell turnover and differentiation remains less understood. Intestinal stem and progenitor cells are essential for the maintenance of intestinal homeostasis and regeneration, and their functions can be modulated by factors of the stem cell niche, including neuronal mediators. Here, we investigated the role of VIP in regulating intestinal epithelial homeostasis and regeneration following irradiation-induced injury.

Jejunal organoids were derived from male and female C57Bl6/J, Lgr5-EGFP-IRES-CreERT2 or Lgr5-EGFP-IRES-CreERT2/R26R-LSL-TdTomato mice and treated with VIP prior to analysis. Injury conditions were induced by exposing organoids to 6 Gy of irradiation (IR). To investigate protective effects of VIP in vivo, mice received 12 Gy of abdominal IR followed by intraperitoneal injections of VIP.

We observed that VIP promotes epithelial differentiation towards a secretory phenotype predominantly via the p38 MAPK pathway. Moreover, VIP prominently modulated epithelial proliferation as well as the number and proliferative activity of Lgr5-EGFP+ progenitor cells under homeostatic conditions. In the context of acute irradiation injury in vitro, we observed that IR injury renders Lgr5-EGFP+ progenitor cells more susceptible to VIP-induced modulations, which coincided with the strong promotion of epithelial regeneration by VIP. Finally, the observed effects translate into an in vivo model of abdominal irradiation, where VIP showed to prominently mitigate radiation-induced injury.

VIP prominently governs intestinal homeostasis by regulating epithelial progenitor cell proliferation and differentiation and promotes intestinal regeneration following acute irradiation injury.

The continuous expansion of intestinal stem cells (ISCs) is crucial for maintaining the renewal of the intestinal epithelium, particularly in inflammatory conditions. It remains largely unknown how the internal microbiota repair damage to the internal mucosal barrier. Hence, investigating potential anti-inflammatory probiotics from the intestinal symbolic microbes of broilers and analyzing their mechanism of action to support the intestinal mucosal barrier function can offer novel regulatory tools to alleviate broiler enteritis. In this research, we utilized in vivo broilers plus ex vivo organoids model to thoroughly examine the effectiveness of Lactobacillus reuteri (LR) in protecting the integrity of the intestinal mucosa during lipopolysaccharide-induced (LPS-induced) enteritis in broilers. The findings indicated that LR feeding maintained intestinal morphological and structural integrity, enhanced proliferation of intestinal epithelial cells, and inhibited cell apoptosis and inflammatory response against the deleterious effects triggered by LPS. Simultaneously, LR enhanced ISCs activity and stimulated intestinal epithelial regeneration to protect the intestinal barrier during LPS-induced injury conditions. The coculture system of LR and ileum organoids revealed that LR increased the growth of organoids and attenuated LPS-stimulated damage to organoids. Furthermore, the LPS-induced decrease in ISC activity was rescued by reactivation of Wnt/β-catenin signaling by LR ex vivo and in vivo. This research revealed that LR promoted the expansion of ISCs and intestinal epithelial cell renewal by regulating the Wnt/β-catenin signaling pathway, thereby maintaining the integrity of the intestinal mucosal barrier. This finding provided theoretical support for lactobacillus as a probiotic additive in livestock feed to improve intestinal inflammation and treat intestinal diseases.

The β-catenin-dependent canonical Wnt signaling is pivotal in organ development, tissue homeostasis, and cancer. Here, we identified an upstream enhancer of Ctnnb1 - the coding gene for β-catenin, named ieCtnnb1 (intestinal enhancer of Ctnnb1), which is crucial for intestinal homeostasis. ieCtnnb1 is predominantly active in the base of small intestinal crypts and throughout the epithelia of large intestine. Knockout of ieCtnnb1 led to a reduction in Ctnnb1 transcription, compromising the canonical Wnt signaling in intestinal crypts. Single-cell sequencing revealed that ieCtnnb1 knockout altered epithelial compositions and potentially compromised functions of small intestinal crypts. While deletion of ieCtnnb1 hampered epithelial turnovers in physiologic conditions, it prevented occurrence and progression of Wnt/β-catenin-driven colorectal cancers. Human ieCTNNB1 drove reporter gene expression in a pattern highly similar to mouse ieCtnnb1. ieCTNNB1 contains a single-nucleotide polymorphism associated with CTNNB1 expression levels in human gastrointestinal epithelia. The enhancer activity of ieCTNNB1 in colorectal cancer tissues was stronger than that in adjacent normal tissues. HNF4α and phosphorylated CREB1 were identified as key trans-factors binding to ieCTNNB1 and regulating CTNNB1 transcription. Together, these findings unveil an enhancer-dependent mechanism controlling the dosage of Wnt signaling and homeostasis in intestinal epithelia.

Recent advances in human genetics have shed light on the genetic factors contributing to inflammatory diseases, particularly Crohn's disease (CD), a prominent form of inflammatory bowel disease. Certain risk genes associated with CD directly influence cytokine biology and cell-specific communication networks. Current CD therapies primarily rely on anti-inflammatory drugs, which are inconsistently effective and lack strategies for promoting epithelial restoration and mucosal balance. To understand CD's underlying mechanisms, we investigated the link between CD and the FGFR1OP gene, which encodes a centrosome protein. FGFR1OP deletion in mouse intestinal epithelial cells disrupted crypt architecture, resulting in crypt loss, inflammation, and fatality. FGFR1OP insufficiency hindered epithelial resilience during colitis. FGFR1OP was crucial for preserving non-muscle myosin II activity, ensuring the integrity of the actomyosin cytoskeleton and crypt cell adhesion. This role of FGFR1OP suggests that its deficiency in genetically predisposed individuals may reduce epithelial renewal capacity, heightening susceptibility to inflammation and disease.

Age at exposure is a critical modifier of the risk of radiation-induced cancer. However, the effects of age on radiation-induced carcinogenesis remain poorly understood. In this study, we focused on tissue stem cells using Lgr5-eGFP-ires-CreERT2 mice to compare radiation-induced DNA damage responses between Lgr5+ and Lgr5- intestinal stem cells. Three-dimensional immunostaining analyses demonstrated that radiation induced apoptosis and the mitotic index more efficiently in adult Lgr5- stem cells than in adult Lgr5+ stem cells but not in infants, regardless of Lgr5 expression. Supporting this evidence, rapid and transient p53 activation occurred after irradiation in adult intestinal crypts but not in infants. RNA sequencing revealed greater variability in gene expression in adult Lgr5+ stem cells than in infant Lgr5+ stem cells after irradiation. Notably, the cell cycle and DNA repair pathways were more enriched in adult stem cells than in infant stem cells after irradiation. Our findings suggest that radiation-induced DNA damage responses in mouse intestinal crypts differ between infants and adults, potentially contributing to the age-dependent susceptibility to radiation carcinogenesis.

Stem cells are critical for the development and homeostasis of the gastrointestinal (GI) tract. Inflammatory molecules are known to regulate the activity of stem cells. A comprehensive review specifically describing the role of inflammatory molecules in the regulation of stem cells within the GI tract and in GI cancers (GICs) is not available. This review focuses on understanding the role of inflammatory molecules and stem cells in maintaining homeostasis of the GI tract. We further discuss how inflammatory conditions contribute to the transformation of stem cells into tumor-initiating cells. We also describe the molecular mechanisms of inflammation and stem cell-driven progression and metastasis of GICs. Furthermore, we report on studies describing the prognostic value of cancer stem cells and the clinical trials evaluating their therapeutic utility. This review provides a detailed overview on the role of inflammatory molecules and stem cells in maintaining GI tract homeostasis and their implications for GI-related malignancies.

Anti-Cluster of differentiation (CD)-40-induced colitis, driven by innate inflammatory responses in the intestine, is a potent animal model exhibiting IBD pathophysiology including diarrhea. However, the ion transport basis of diarrhea and some key mucosal pathways (Paneth cells, stem cell niche, and mechanosensory) in this model have not been investigated.

Mucosal scrapings and intestinal tissue from control and CD40 antibody (150 µg) treated Rag2-/- mice were examined for gut inflammation, Paneth cell numbers, expression of key transporters, tight/adherens junction proteins, stem cell niche, and mechanosensory pathway via hematoxylin and eosin staining, quantitative polymerase chain reaction, and western blotting.

Compared with control, anti-CD40 antibody treatment resulted in a significant loss of body weight (P < .05) and diarrhea at day 3 postinjection. Distal colonic tissues of anti-CD40 mice exhibited increased inflammatory infiltrates, higher claudin-2 expression, and appearance of Paneth cell-like structures indicative of Paneth cell metaplasia. Significantly reduced expression (P < .005) of downregulated in adenoma (key Cl- transporter), P-glycoprotein/multidrug resistantance-1 (MDR1, xenobiotic transporter), and adherens junction protein E-cadherin (~2-fold P < .05) was also observed in the colon of anti-CD40 colitis mice. Interestingly, there were also marked alterations in the stem cell markers and upregulation of the mechanosensory YAP-TAZ pathway, suggesting the activation of alternate regeneration pathway post-tissue injury in this model.

Our data demonstrate that the anti-CD40 colitis model shows key features of IBD observed in the human disease, hence making it a suitable model to investigate the pathophysiology of ulcerative colitis (UC).

Radiation-induced intestinal injury (RIII) commonly occur during abdominal-pelvic cancer radiation therapy; however, no effective prophylactic or therapeutic agents are available to manage RIII currently. This study aimed to clarify the potential of probiotic consortium supplementation in alleviating RIII.

Male C57BL/6J mice were orally administered a probiotic mixture comprising Bifidobacterium longum BL21, Lactobacillus paracasei LC86, and Lactobacillus plantarum Lp90 for 30 days before exposure to 13 Gy of whole abdominal irradiation. The survival rates, clinical scores, and histologic changes in the intestines of mice were assessed. The impacts of probiotic consortium treatment on intestinal stem cell proliferation, differentiation, and epithelial barrier function; oxidative stress; and inflammatory cytokines were evaluated. A comprehensive examination of the gut microbiota composition was conducted through 16S rRNA sequencing, while changes in metabolites were identified using liquid chromatography-mass spectrometry.

The probiotic consortium alleviated RIII, as reflected by increased survival rates, improved clinical scores, and mitigated mucosal injury. The probiotic consortium treatment exhibited enhanced therapeutic effects at the histologic level compared with individual probiotic strains, although there was no corresponding improvement in survival rates and colon length. Moreover, the probiotic consortium stimulated intestinal stem cell proliferation and differentiation, enhanced the integrity of the intestinal epithelial barrier, and regulated redox imbalance and inflammatory responses in irradiated mice. Notably, the treatment induced a restructuring of the gut microbiota composition, particularly enriching short-chain fatty acid-producing bacteria. Metabolomic analysis revealed distinctive metabolic changes associated with the probiotic consortium, including elevated levels of anti-inflammatory and antiradiation metabolites.

The probiotic consortium attenuated RIII by modulating the gut microbiota and metabolites, improving inflammatory symptoms, and regulating oxidative stress. These findings provide new insights into the maintenance of intestinal health with probiotic consortium supplementation and will facilitate the development of probiotic-based therapeutic strategies for RIII in clinical practice.

Interferon-gamma (IFNγ) is traditionally recognized for its pro-inflammatory role during intestinal inflammation. Here, we demonstrate that IFNγ also functions as a pro-repair molecule by increasing TNFα receptor 2 (TNFR2 protein/TNFRSF1B gene) expression on intestinal epithelial cells (IEC) following injury in vitro and in vivo. In silico analyses identified binding sites for the IFNγ signaling transcription factor STAT1 in the promoter region of TNFRSF1B. Scratch-wounded IEC exposed to IFNγ exhibited a STAT1-dependent increase in TNFR2 expression. In situ hybridization revealed elevated Tnfrsf1b mRNA levels in biopsy-induced colonic mucosal wounds, while intraperitoneal administration of IFNγ neutralizing antibodies following mucosal injury resulted in impaired IEC Tnfrsf1b mRNA and inhibited colonic mucosal repair. These findings challenge conventional notions that "pro-inflammatory" mediators solely exacerbate damage by highlighting latent pro-repair functions. Moreover, these results emphasize the critical importance of timing and amount in the synthesis and release of IFNγ and TNFα during the inflammatory process, as they are pivotal in restoring tissue homeostasis.

Graft-versus-host disease (GVHD) is a major life-threatening complication that occurs after allogeneic hematopoietic cell transplantation (HCT). Although adult tissue stem cells have been identified as targets of GVHD in the skin and gut, their role in hepatic GVHD is yet to be clarified. In the current study, we explored the fate of bile duct stem cells (BDSCs), capable of generating liver organoids in vitro, during hepatic GVHD after allogeneic HCT. We observed a significant expansion of biliary epithelial cells (BECs) on injury early after allogeneic HCT. Organoid-forming efficiency from the bile duct was also significantly increased early after allogeneic HCT. Subsequently, the organoid-forming efficiency from bile ducts was markedly decreased in association with the reduction of BECs and the elevation of plasma concentrations of bilirubin, suggesting that GVHD targets BDSCs and impairs the resilience of BECs. The growth of liver organoids in the presence of liver-infiltrating mononuclear cells from allogeneic recipients, but not from syngeneic recipients, was significantly reduced in a transforming growth factor-β (TGF-β)-dependent manner. Administration of SB-431542, an inhibitor of TGF-β signaling, from day 14 to day 28, protected organoid-forming BDSCs against GVHD and mitigated biliary dysfunction after allogeneic HCT, suggesting that BDSCs are a promising therapeutic target for hepatic GVHD.