Intestinal Cell Differentiation Research Articles

Identification of feature genes in intestinal epithelial cell types.

The intestine, is responsible for food digestion, nutrient absorption, endocrine secretion, food residue excretion, and immune defense. These function performances are based on the intricate composition of intestinal epithelial cells, encompassing differentiated mature cells, rapidly proliferative cells, and intestinal stem cells. Although the characteristics of these cell types are well-documented, in-depth exploration of their representative markers and transcription factors is critical for comprehensive cell fate trajectory analysis. Here, we unveiled the feature genes in different cell types of the human and mouse gut through single-cell RNA sequencing analysis. Further, the locations of some specific transcription factors and membrane proteins were determined by immunofluorescence staining, and their role in regulating the proliferation and differentiation of intestinal epithelial cells were explored by CRISPR/Cas9 knockout. Therefore, this study not only reports new markers for various intestinal epithelial cell types but also elucidates the involvement of relevant genes in the determination of epithelial cell fate and maintenance of stem cell homeostasis, which facilitates the tracing and functional elucidation of intestinal epithelial cells.

Regulation of the Intestinal Stem Cell Pool and Proliferation in Drosophila

Understanding the regulation of somatic stem cells, both during homeostasis and in response to environmental challenges like injury, infection, chemical exposure, and nutritional changes, is critical because their dysregulation can result in tissue degeneration or tumorigenesis. The use of models such as the Drosophila and mammalian adult intestines offers valuable insights into tissue homeostasis and regeneration, advancing our knowledge of stem cell biology and cancer development. This review highlights significant findings from recent studies, unveiling the molecular mechanisms that govern self-renewal, proliferation, differentiation, and regeneration of intestinal stem cells (ISCs). These insights not only enhance our understanding of normal tissue maintenance but also provide critical perspectives on how ISC dysfunction can lead to pathological conditions such as colorectal cancer (CRC).

Targeting Gut microbiota as a therapeutic target in T2DM: a review of Multi-target interactions of probiotics, prebiotics, postbiotics, and synbiotics with the Intestinal Barrier

The global epidemic of type 2 diabetes mellitus (T2DM) imposes a substantial burden on public health and healthcare expenditures, thereby driving the pursuit of cost-effective preventive and therapeutic strategies. Emerging evidence suggests a potential association between dysbiosis of gut microbiota and its metabolites with T2DM, indicating that targeted interventions aimed at modulating gut microbiota may represent a promising therapeutic approach for the management of T2DM. In this review, we concentrated on the multifaceted interactions between the gut microbiota and the intestinal barrier in the context of T2DM. We systematically summarized that the imbalance of beneficial gut microbiota and its metabolites may constitute a viable therapeutic approach for the management of T2DM. Meanwhile, the mechanisms by which gut microbiota interventions, such as probiotics, prebiotics, postbiotics, and synbiotics, synergistically improve insulin resistance in T2DM are summarized. These mechanisms include the restoration of gut microbiota structure, upregulation of intestinal epithelial cell proliferation and differentiation, enhancement of tight junction protein expression, promotion of mucin secretion by goblet cells, and the immunosuppressive functions of regulatory T cells (Treg) and M2 macrophages. Collectively, these actions contribute to the amelioration of the body's metabolic inflammatory status. Our objective is to furnish evidence that supports the clinical application of probiotics, prebiotics, and postbiotics in the management of T2DM.

Assessment of equine intestinal epithelial junctional complexes and barrier permeability using a monolayer culture system.

Gastrointestinal disease is a leading cause of death in mature horses. A lack of in vitro modeling has impeded the development of novel therapeutics. The objectives of this study were to develop and further characterize a small intestinal monolayer cell culture derived from equine jejunum including establishing normal measurements of intestinal permeability and restitution. Three-dimensional enteroids, derived from postmortem sampling of equine jejunum, were utilized to develop confluent epithelial monolayers. The presence of differentiated intestinal epithelial cell types and tight junctions were confirmed using histology, reverse transcription PCR (RT-PCR), RNAscope, protein immunofluorescence and transmission electron microscopy. Transepithelial resistance (TER) and macromolecule flux were assessed as measurements of paracellular and transcellular permeability. Scratch assays were utilized to model and assess intestinal restitution. Monolayer cell cultures reached 100% confluency by ~5-7 days. Equine jejunum monolayers were confirmed as epithelial in origin, with identification of differentiated intestinal epithelial cell types and evidence of tight junction proteins. Function of the intestinal barrier was supported by acquisition of physiologically normal TER values (179.9 ± 33.7 ohms*cm2) and limited macromolecule flux (22 ± 8.8% at 60 min). Additionally, following a scratch wound, epithelial cell monolayers migrated to close gap defects within 24 h. In conclusion, this study describes the development of a novel intestinal epithelial monolayer cell culture for equine jejunum, and provides evidence of intestinal epithelial cell differentiation, formation of physiologically relevant barrier function and use as a model of intestinal restitution to test potential therapeutics for equine colic.

Exploring the impact of sEH inhibition on intestinal cell differentiation and Colon Cancer: Insights from TPPU treatment

Inhibition of soluble epoxide hydrolase (sEH) appears to be promising for the treatment of many diseases. Studies have focused on the beneficial effects of epoxyeicosatrienoic acids (EETs), which are sEH substrates. However, our recent studies have shown that the sEH activity is crucial for the proper intestinal cell differentiation. In this recent study, we investigated the impact of TPPU, an inhibitor of sEH, on the colon cancer cell lines Caco2 and HT-29. We analysed the changes in the expression of the cytoskeletal protein ezrin and the phosphorylated protein kinase p38 (p-p38). Our results showed a decrease in ezrin expression in differentiated cells and an increase in p-p38 expression after TPPU treatment. Immunocytochemical staining revealed a higher staining intensity of p-p38 in the nuclei of HT-29 cells following TPPU treatment. Immunohistochemical staining was performed on human samples of normal colon tissue, grade 2 tumours, and embryonal/foetal tissues. The staining intensity of ezrin in tumours was reduced in the surface area compared to the crypts. Additionally, we observed the translocation of p-p38 expression from the cytoplasm to the nucleus during differentiation. The tumour samples exhibited higher levels of p-p38 in the cytoplasm, similar to normal undifferentiated tissue. To observe the disruption of the cytoskeleton after TPPU treatment, confocal microscopy was used. It was found that β-actin associated with ezrin forms clusters under the plasma membranes. All of these results are significant because sEH inhibitors are being tested in clinical trials, but they could cause an unexpected adverse effects.

Sight of Aged Microplastics Adsorbing Heavy Metal Exacerbated Intestinal Injury: A Mechanistic Study of Autophagy-Mediated Toxicity Response.

Contaminant-bearing polystyrene microplastics (PSMPs) may exert significantly different toxicity profiles from their contaminant-free counterparts, with the role of PSMPs in promoting contaminant uptake being recognized. However, studies investigating the environmentally relevant exposure and toxic mechanisms of aged PSMPs binding to Cr are limited. Here, we show that loading of chromium (Cr) markedly alters the physicochemical properties and toxicological profiles of aged PSMPs. Specifically, Cr-bearing aged PSMPs induced severe body weight loss, oxidative stress (OS), autophagy, intestinal barrier injury, inflammation-pyroptosis response, and enteropathogen invasion in mice. Mechanistic investigations revealed that PSMPs@Cr exacerbated the OS, resulting in intestinal barrier damage and inflammation-pyroptosis response via overactivated Notch signaling and autophagy/cathepsin B/IL-1β pathway, respectively, which ultimately elevated mortality related to bacterial pathogen infection. In vitro experiments confirmed that autophagy-mediated reactive oxygen species (ROS) overproduction resulted in severe pyroptosis and impaired intestinal stem cells differentiation alongside the overactivation of Notch signaling in PSMPs@Cr-exposed organoids. Overall, our findings provide an insight into autophagy-modulated ROS overproduction within the acidic environment of autophagosomes, accelerating the release of free Cr from PSMPs@Cr and inducing secondary OS, revealing that PSMPs@Cr is a stable hazard material that induces intestinal injury. These findings provided a potential therapeutic target for environmental MPs pollution caused intestinal disease in patients.

Vasoactive intestinal peptide promotes secretory differentiation and mitigates radiation-induced intestinal injury

BackgroundVasoactive 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.MethodsJejunal 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.ResultsWe 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.ConclusionsVIP prominently governs intestinal homeostasis by regulating epithelial progenitor cell proliferation and differentiation and promotes intestinal regeneration following acute irradiation injury.

Loss of ARID3A perturbs intestinal epithelial proliferation-differentiation ratio and regeneration.

Intestinal stem cells at the crypt divide and give rise to progenitor cells that proliferate and differentiate into various mature cell types in the transit-amplifying (TA) zone. Here, we showed that the transcription factor ARID3A regulates intestinal epithelial cell proliferation and differentiation at the TA progenitors. ARID3A forms an expression gradient from the villus tip to the upper crypt mediated by TGF-β and WNT. Intestinal-specific deletion of Arid3a reduces crypt proliferation, predominantly in TA cells. Bulk and single-cell transcriptomic analysis shows increased enterocyte and reduced secretory differentiation in the Arid3a cKO intestine, accompanied by enriched upper-villus gene signatures of both cell lineages. We find that the enhanced epithelial differentiation in the Arid3a-deficient intestine is caused by increased binding and transcription of HNF1 and HNF4. Finally, we show that loss of Arid3a impairs irradiation-induced regeneration with sustained cell death and reprogramming. Our findings imply that Arid3a functions to fine-tune the proliferation-differentiation dynamics at the TA progenitors, which are essential for injury-induced regeneration.

GZMA suppressed GPX4-mediated ferroptosis to improve intestinal mucosal barrier function in inflammatory bowel disease

BackgroundOur previous study has demonstrated a decreased colonic CD8+CD39+ T cells, enrichment of granzyme A (GZMA), was found in pediatric-onset colitis and inflammatory bowel disease (IBD) characterized by impaired intestinal barrier function. However, the influence of GZMA on intestinal barrier function remains unknown.MethodsWestern blotting(WB), real-time PCR (qPCR), immunofluorescence (IF) and in vitro permeability assay combined with intestinal organoid culture were used to detect the effect of GZMA on intestinal epithelial barrier function in vivo and in vitro. Luciferase, immunoprecipitation (IP) and subcellular fractionation isolation were performed to identify the mechanism through which GZMA modulated intestinal epithelial barrier function.ResultsHerein, we, for the first time, demonstrated that CD8+CD39+ T cells promoted intestinal epithelial barrier function through GZMA, leading to induce Occludin(OCLN) and Zonula Occludens-1(ZO-1) expression, which was attributed to enhanced CDX2-mediated cell differentiation caused by increased glutathione peroxidase 4(GPX4)-induced ferroptosis inhibition in vivo and in vitro. Mechanically, GZMA inhibited intestinal epithelial cellular PDE4B activation to trigger cAMP/PKA/CREB cascade signaling to increase CREB nuclear translocation, initiating GPX4 transactivity. In addition, endogenous PKA interacted with CREB, and this interaction was enhanced in response to GZMA. Most importantly, administration of GZMA could alleviate DSS-induced colitis in vivo.ConclusionThese findings extended the novel insight of GZMA contributed to intestinal epithelial cell differentiation to improve barrier function, and enhacement of GZMA could be a promising strategy to patients with IBD.

MiR-27a-3p regulates intestinal cell proliferation and differentiation through Wnt/β-catenin signalling.

Intestinal stem cells differentiate into absorptive enterocytes, characterised by increased brush border enzymes such as intestinal alkaline phosphatase (IAP), making up the majority (95%) of the terminally differentiated cells in the villus. Loss of integrity of the intestinal epithelium plays a key role in inflammatory diseases and gastrointestinal infection. Here, we show that the intestinal microRNA (miR)-27a-3p is an important regulator of intestinal epithelial cell proliferation and enterocyte differentiation. Repression of endogenous miR-27a-3p leads to increased enterocyte differentiation and decreased intestinal epithelial cell proliferation in mouse and human small intestinal organoids. Mechanistically, miR-27a-3p regulates intestinal cell differentiation and proliferation at least in part through the regulation of retinoic acid receptor α (RXRα), a modulator of Wnt/β-catenin signalling. Repression of miR-27a-3p increases the expression of RXRα and concomitantly, decreases the expression of active β-catenin and cyclin D1. In contrast, overexpression of miR-27a-3p mimic decreases the expression of RXRα and increases the expression of active β-catenin and cyclin D1. Moreover, overexpression of the miR-27a-3p mimic results in impaired enterocyte differentiation and increases intestinal epithelial cell proliferation. These alterations were attenuated or blocked by Wnt inhibition. Our study demonstrates an miR-27a-3p/RXRα/Wnt/β-catenin pathway that is important for the maintenance of enterocyte homeostasis in the small intestine.

Selenium nanoparticles affect chicken offspring's intestinal health better than other selenium sources

This study aimed to assess the effects of maternal diets containing various selenium (Se) sources on the intestinal mucosal function in the jejunum of chicken offspring. A total of 630, 18-wk-old Hy-Line Grey hens and 70 18-wk-old Hy-Line Grey breeders were randomly allocated into 7 groups, with 5 replicates in each group (18 hens and tow roosters). After 4 wk of Se depletion, the birds were fed either a nonsupplemented basal diet (control) or the same basal diet supplemented with 0.15 mg/kg selenium nanoparticles (Nano-Se), 0.30 mg/kg Nano-Se, 0.30 mg/kg selenocysteine (Sec), 0.30 mg/kg sodium selenite (SS), 0.30 mg/kg selenomethionine (SeMet), or 0.15 mg/kg Nano-Se + 0.15 mg/kg Sec, for 8 wk. Frtilized eggs were collected and incubated during the final week of the experiment. Jejunal tissues from embryonic d 18 and the hatch day were collected for analysis, and the 7-d survival rate of the offspring was recorded. Compared to the control, the maternal diet of 0.30 mg/kg Nano-Se, 0.30 mg/kg Sec, and 0.30 mg/kg SeMet significantly increased the survival of 7-day-old offspring (P < 0.05). The maternal diet supplemented with 0.30 mg/kg Nano-Se significantly increased intestinal villus height and the villus height/crypt depth ratio in chicks at embryonic d 18 and in 1-day-old (P < 0.05). The maternal diet containing 0.30 mg/kg Nano-Se and Sec increased the mRNA expression levels of tight junction proteins in 1-day-old offspring (P < 0.05). Supplemental 0.30 mg/kg Nano-Se significantly increased the mRNA expression levels of marker genes in intestinal enteroendocrine, stem, and Paneth cells (P < 0.05). In 1-day-old chicks, the number of intestinal goblet cells, as well as the mRNA expression levels of intestinal mucin2 (Muc2) and goblet cell differentiation factors (Spdef and C-myc), were the highest in diets supplemented with 0.30 mg/kg Nano-Se. Moreover, the expression levels of intestinal Muc2 and Spdef in chicks at embryonic d 18 was the highest with 0.30 mg/kg Nano-Se supplementation (P < 0.05). Supplementing with 0.30 mg/kg Nano-Se significantly reduced reactive oxygen species levels and decreased the mRNA expression levels of apoptosis-related genes in 1-day-old chicks (P < 0.05). Additionally, 0.30 mg/kg Nano-Se supplementation significantly down-regulated NLRP3 pathway gene expression in 1-day-old chicks (P < 0.05). In conclusion, maternal dietary supplementation with Nano-Se improved jejunal microarchitecture, antioxidant levels, and the expression of tight-junction protein in chicken offspring along with supporting goblet cell development by inhibiting the NLRP3 signaling pathway.

Electroacupuncture Promotes the Generation of Intestinal Treg Cells After Ischemic Stroke by Foxp3 Acetylation Regulation.

Electroacupuncture (EA) has been shown to ameliorate brain injury and protect against intestinal injury after ischemic stroke. These protective effects are closely associated with the enhancement of regulatory T (Treg) cell numbers and function in the intestine, as well as the inhibition of intestinal γδ T cell production and their migration to the brain. This study aimed to elucidate the potential mechanism by which EA regulates intestinal Treg cell differentiation after stroke. Sprague-Dawley rats were divided into three groups: the sham group, the middle cerebral artery occlusion (MCAO) group, and the MCAO plus EA (MEA) group. The MCAO model was generated by occluding the middle cerebral artery. EA was applied to Baihui (GV20) acupoint once daily. Samples were collected 3days after reperfusion. Our results showed that EA reduced the inflammatory response in the brain and intestine after ischemic stroke. EA treatment increased the percentage of Treg cells in the small intestine of rats. EA increased the levels of SCFAs, while also inhibiting histone deacetylase activity (HDAC). Additionally, acetylated Foxp3 protein in the small intestine was increased after EA treatment. These results suggest that EA at GV20 alleviates brain and intestinal inflammatory injury in stroke rats, potentially through the enhancement of SCFA-mediated Foxp3 acetylation in Treg cells.

Effect of Butyrate on Food-Grade Titanium Dioxide Toxicity in Different Intestinal In Vitro Models.

Short-chain fatty acids (SCFA) are an important energy source for colonocytes and crucial messenger molecules both locally in the intestine and systemically. Butyrate, one of the most prominent and best-studied SCFA, was demonstrated to exert anti-inflammatory effects, improve barrier integrity, enhance mucus synthesis in the intestine, and promote cell differentiation of intestinal epithelial cells in vitro. While the physiological relevance is undisputed, it remains unclear if and to what extent butyrate can influence the effects of xenobiotics, such as food-grade titanium dioxide (E171, fgTiO2), in the intestine. TiO2 has been controversially discussed for its DNA-damaging potential and banned as a food additive within the European Union (EU) since 2022. First, we used enterocyte Caco-2 monocultures to test if butyrate affects the cytotoxicity and inflammatory potential of fgTiO2 in a pristine state or following pretreatment under simulated gastric and intestinal pH conditions. We then investigated pretreated fgTiO2 in intestinal triple cultures of Caco-2, HT29-MTX-E12, and THP-1 cells in homeostatic and inflamed-like state for cytotoxicity, barrier integrity, cytokine release as well as gene expression of mucins, oxidative stress markers, and DNA repair. In Caco-2 monocultures, butyrate had an ambivalent role: pretreated but not pristine fgTiO2 induced cytotoxicity in Caco-2 cells, which was not observed in the presence of butyrate. Conversely, fgTiO2 induced the release of interleukin 8 in the presence but not in the absence of butyrate. In the advanced in vitro models, butyrate did not affect the characteristics of the healthy or inflamed states and caused negligible effects in the investigated end points following fgTiO2 exposure. Taken together, the effects of fgTiO2 strongly depend on the applied testing approach. Our findings underline the importance of the experimental setup, including the choice of in vitro model and the physiological relevance of the exposure scenario, for the hazard testing of food-grade pigments like TiO2.

Toxicity Evaluation of Potassium Sorbate In Vivo with Drosophila Melanogaster.

Potassium sorbate (PS) is a preservative widely used in the food, pharmaceutical, and cosmetics industries. Improper and careless use of PS can lead to various health issues and potential environmental problems. Drosophila is capable of making rapid and sensitive responses to stress or other stimuli. Here we utilized Drosophila as a model organism to evaluate the potential toxicity of PS. Our study revealed that PS ingestion reduced the lifespan and fecundity of Drosophila. In addition, excessive PS ingestion led to cell apoptosis and ROS accumulation in the midgut. Furthermore, PS intake also enhanced the mitophagy of midgut cells. Strikingly, PS affected the cell differentiation progression as well, leading to the production of more enteroendocrine (EE) cells. We further demonstrated that the expression of notch (N), a vital player in intestinal stem cell (ISC) differentiation, was down-regulated in the midgut. This indicates that the differentiation progression was affected potentially by repressing the N expression.

Establishment of an in-vitro inflammatory bowel disease model using immunological differentiation of Caco-2 cells

Studies on intestinal cell differentiation, particularly in dextran sodium sulfate (DSS)-induced inflammatory bowel disease (IBD), have predominantly focused on the disruption of intestinal crypts and suppressive effects on the intestinal microbiota; however, repeated administration of DSS is required to induce inflammatory pathology, and there is a lack of observation of early responses and consideration of differentiation stages. Although colonic adenocarcinoma (Caco-2) cells can be used as intestinal cell models, research on these cells in an immature state is limited. We, therefore, investigated the relationship between Caco-2 cell culture duration and immunological differentiation using α-defensin5 (DEFA5) as an indicator of intestinal immunity and differentiation. Changes in protein and gene expression levels in response to DSS were examined at each differentiation stage. Expression of immune- and differentiation-related proteins, including DEFA5 and lysozyme, was evident from Day 8 of culture. Immune responses to DSS varied with the differentiation stage, affecting cell viability and cytokine expression.•Caco-2 cell culture duration correlates with the differentiation stage of Paneth cells.•DSS exposure elicits different effects depending on the differentiation stage.•Our in-vitro model of IBD facilitates the characterization of the cell differentiation process and provides a methodology to help elucidate the causal mechanisms of IBD.

Characterizing the effects of triclosan and triclocarban on the intestinal epithelial homeostasis using small intestinal organoids

Intestinal epithelium has the largest surface of human body, contributes dramatically to defense of toxicant-associated intestinal injury. Triclosan (TCS) and triclocarban (TCC), extensively employed as antibacterial agents in personal care products (PCPs) and healthcare facilities, caused serious damage to human intestine. However, the role of the intestinal epithelium in TCS/TCC-induced intestinal toxicity and its underlying toxic mechanisms remain incompletely understood. In this study, a novel 3D intestinal organoid model was utilized to investigate that exposure to TCS/TCC led to a compromised self-renewal and differentiation of intestinal stem cells (ISCs). Consequently, this disrupted intestinal epithelial homeostasis ultimately caused a reduction in nutrient absorption and deficient of epithelial defense to exogenous and endogenous pathogens stimulation. The inhibition of the Wnt signaling pathway in intestinal stem cell was contributed to the intestinal toxicity of TCS/TCC. These results were further confirmed in vivo with mice exposed to TCS/TCC. The findings of this study provide evidence that TCS/TCC possess the capacity to disturb the homeostasis of the intestinal epithelium, and emphasize the credibility of organoids as a valuable model for toxicological studies, as they could faithfully recapitulate in vivo phenomena.

Transcriptomic Landscape of Colorectal Mucinous Adenocarcinoma has Similarity with Intestinal Goblet Cell Differentiation

Introduction: Colorectal mucinous adenocarcinoma (MC) differs from adenocarcinoma (AD) in clinical features and molecular characteristics. The current treatment of colorectal MC is not precise enough, and the molecular characteristics remain unclear. The study aims to explore the difference between colorectal MC and AD on the transcriptome level for the possibility of treating colorectal MC precisely. Methods: The data of colorectal cancer (CRC) patients from The Cancer Genome Atlas (TCGA) database was assessed, and then differential analysis and weighted gene co-expression network analysis (WGCNA) were performed to identify the differential hub RNAs between colorectal MC and AD. Differential hub lncRNAs and hub RNA of significant modules were validated by quantitative real-time PCR (qRT-PCR) among different colon cancer cell lines. Results: In total, 1680 differential expressed RNAs (DERs) were found by comparing colorectal MC (52, 13.3%) with AD (340, 86.7%). Through the WGCNA, a mucin-associated RNA module was identified, while some others might be associated with unique immune progress. Finally, 6 differential hub RNAs in the mucin-associated RNA module (CTD-2589M5.4, RP11-234B24.2, RP11-25K19.1 and COLCA1) were validated by qRT-PCR and showed higher expression levels in mucin-producing colorectal cell lines (Ls174T and HT-29). Conclusion: This study suggests that clinical treatments for colorectal MC should be differentiated from AD. Further exploration of enterocyte (goblet cell) differentiation with tumor genesis and the distinct immune progression of MC may help to identify key therapeutic targets for colorectal MC. Further research on the application of immunotherapy to colorectal MC is needed.

Sirtuin 6 inhibits group 3 innate lymphoid cell function and gut immunity by suppressing IL-22 production.

Group 3 innate lymphoid cells (ILC3s) are enriched in the intestinal mucosa and play important roles in host defense against infection and inflammatory diseases. Sirtuin 6 (SIRT6) is a nicotinamide adenine dinucleotide (NAD+)- dependent deacetylase and has been shown to control intestinal epithelial cell differentiation and survival. However, the role of SIRT6 in ILC3s remains unknown. To investigate the role of SIRT6 in gut ILC3s, we generated SIRT6 conditional knockout mice by crossing Rorccre and Sirt6flox/flox mice. Cell number and cytokine production was examined using flow cytometry. Citrobacter rodentium infection and dextran sodium sulfate-induced colitis models were used to determine the role of SIRT6 in gut defense. RT-qPCR, flow cytometry and immunohistochemistry were used to assess the intestinal inflammatory responses. Here we show that SIRT6 inhibits IL-22 expression in intestinal ILC3s in a cell-intrinsic manner. Deletion of SIRT6 in ILC3s does not affect the cell numbers of total ILC3s and subsets, but results in increased IL-22 production. Furthermore, ablation of SIRT6 in ILC3s protects mice against Citrobacter rodentium infection and dextran sodium sulfate-induced colitis. Our results suggest that SIRT6 may play a role in ILC3 function by regulating gut immune responses against bacterial infection and inflammation. Our finding provided insight into the relation of epigenetic regulators with IL-22 production and supplied a new perspective for a potential strategy against inflammatory bowel disease.

WTAP promotes the progression of ulcerative colitis by silencing the expression of CES2 through m6A modification

ObjectiveThis study will explore the function of WTAP, the critical segment of m6A methyltransferase complex, in UC and its regulation on immune response. MethodsThe expression levels of key proteins were detected in colon tissues which were derived from UC patients and mice. Macrophage polarization and CD4+ T cell infiltration were detected by flow cytometry and IF staining. ELISA assay was utilized to analyze the level of the inflammatory cytokines. m6A-RIP-PCR, actinomycin D test, and RIP assays were utilized to detect the m6A level, stability, and bound proteins of CES2 mRNA. A dual luciferase reporter assay was conducted to confirm the transcriptional interactions between genes. A co-culture system of intestinal epithelium-like organs was constructed to detect the primary mouse intestinal epithelial cells (PMIEC) differentiation. The interaction between proteins was detected via Co-IP assay. ResultsThe expression of WTAP and CES2 in UC tissues was increased and decreased, respectively. Knockdown of WTAP inhibited the progression of UC in mice by inhibiting M1 macrophage polarization and CD4+ T cell infiltration. WTAP combined YTHDF2 to promote the m6A modification of CES2 mRNA and inhibited its expression. CES2 co-expressed with EPHX2 and overexpression of CES2 promoted the differentiation of PMIEC. The inhibitory effect of WTAP knockdown on the progress of UC was partially abrogated by CES2 knockdown. ConclusionWTAP/YTHDF2 silences CES2 by promoting its m6A modification and then promotes the progression of UC. WTAP could be a promoting therapy target of UC.

Heterogeneity effects of bisphenol A and its substitute, fluorene-9-bisphenol, on intestinal homeostasis

Bisphenol A (BPA) and its substitute fluorene-9-bisphenol (BHPF) are used in consumer products; however, their toxic effects on intestinal epithelium remain largely unknown. In this study, we combined intestinal organoids and single-cell RNA sequencing to investigate the impact of BPA and BHPF exposure on intestinal cell composition, differentiation, and function. Both compounds inhibited the growth of small intestinal organoids, with BHPF exhibiting a more potent inhibitory effect. BPA and BHPF did not significantly alter the overall cell type composition; however, they led to different alterations in cell–cell communications. Gene Ontology enrichment analysis showed that BPA and BHPF exposures affected various biological processes, such as glutathione transferase activity, antioxidant activity, and lipid metabolism, in cell type-specific and compound-dependent manners. Trajectory analysis demonstrated that BPA and BHPF altered the differentiation trajectory of the intestinal cells. To further connect the cellular mechanism to the phenotypic impact in vivo, we constructed a mouse model exposed to BPA or BHPF and observed significant alterations in intestinal morphology, including reduced crypt depth and villus length and impaired stem cell proliferation and self-renewal. These results provide novel insights into the cell type-specific effects of BPA and BHPF on the intestinal epithelium and highlight the potential risks of exposure to these compounds. Our findings underscore the importance of evaluating the safety of BPA substitutes and contribute to a better understanding of the effects of environmental chemicals on gut health.