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OBJECTIVE: Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD) with a rising global incidence and significant socioeconomic impact. While macrophages are key drivers of intestinal inflammation, their heterogeneity and molecular characteristics in UC remain poorly defined. This study aimed to dissect macrophage subpopulations and validate potential therapeutic targets through integrated single-cell and experimental analyses. METHODS: We analyzed publicly available single-cell RNA sequencing (scRNA-seq) datasets from colonic tissues of healthy controls (HC), non-inflamed UC, and inflamed UC patients. Data preprocessing, clustering, and cell-type annotation were performed using Seurat. Macrophage subsets were identified and characterized through pseudotime trajectory analysis (Monocle3) and intercellular communication profiling (CellChat). High-dimensional weighted gene co-expression network analysis (hdWGCNA) was employed to uncover macrophage-associated gene modules. Candidate genes were evaluated for diagnostic potential in an independent bulk transcriptomic UC cohort using nine machine learning algorithms. Functional validation of the key gene was conducted in vivo using a dextran sulfate sodium (DSS)-induced colitis mouse model and in vitro with LPS-stimulated RAW264.7 macrophages. RESULTS: scRNA-seq analysis revealed 18 major cell types across all samples, with macrophages notably enriched in inflamed tissues. Five distinct macrophage subsets were identified: APOE+, S100A8+, LSP1+, IGHM+, and IL1B+ macrophages. Among these, S100A8+ and IL1B+ macrophages were significantly expanded in inflamed UC and displayed non-classical M1/M2 polarization patterns. CellChat analysis demonstrated that these subsets were predominant contributors to pro-inflammatory signaling and mucosal injury. hdWGCNA uncovered five macrophage-specific co-expression modules, with the macrophage-M2 module-active in S100A8+ macrophages-enriched in S100A4, S100A6, and VCAN. Among nine machine learning models, the Random Forest algorithm achieved the highest diagnostic accuracy (AUC = 0.89) in predicting UC based on these gene signatures. Functionally, both pharmacological inhibition (niclosamide) and lentiviral shRNA-mediated knock-down of S100A4 in RAW264.7 macrophages significantly attenuated DSS-induced colitis in vivo and suppressed LPS-driven M1 polarization while restoring M2 markers in vitro, validating S100A4 as a critical therapeutic node. CONCLUSION: This study provides a comprehensive single-cell landscape of macrophage heterogeneity in UC, identifying S100A8+ and IL1B+ macrophages as key mediators of mucosal inflammation. Integrative analysis pinpointed S100A4 as a robust diagnostic and therapeutic candidate, with experimental validation underscoring its translational potential. These findings offer new insights into macrophage-driven pathogenesis and open avenues for targeted UC therapies.

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Ulcerative colitis (UC), a chronic inflammatory bowel disease, significantly impairs patients' quality of life. Although conventional therapies remain widely utilized, their effectiveness is often constrained by undesirable side effects and frequent relapses. Therefore, new treatment strategies are urgently needed to minimize toxicity and enhance sustainability. In this study, we investigated the protective effects and underlying mechanisms of vanillic acid (VA) against UC using both cells and mouse models. In vitro experiments demonstrated that VA pretreatment effectively attenuated the lipopolysaccharide (LPS) induced accumulation of reactive oxygen species and superoxide anions in RAW 264.7 cells. Furthermore, VA decreased the expression of NFκB, IL-6, and IL-1ß while increasing the expression of Claudin-1, Occluding, and ZO-1 in LPS stimulated NCM460 cells. In vivo, UC was induced by dextran sulfate sodium (DSS) and VA was administered orally. The results indicated that, treatment with VA could significantly alleviate weight loss, shorten colon length, decrease DAI score, and alleviate intestinal damage caused by DSS. Mechanistically, VA ameliorated UC by reducing oxidative stress, suppressing inflammation, regulating macrophage polarization, and enhancing intestinal barrier integrity. Moreover, transcriptome analysis combined with KEGG pathway enrichment revealed that VA modulated key pathways in colon tissue, including the AMPK signaling pathways, PPAR signaling pathways, neuroactive ligand-receptor interaction, and linoleic acid metabolism. The results indicate the therapeutic potential of VA in the treatment of UC, providing a new insight into its mechanisms of action and establishing a foundation for future clinical applications.

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BACKGROUND & AIMS: Cytotoxic T cells have been postulated to facilitate the destruction of intestinal epithelium in inflammatory bowel diseases (IBDs). CADM1, which encodes a membrane adhesion protein that can bind the T cell receptor CRTAM, was markedly up regulated in colons of IBD patients compared with non-IBD (NIBD) patients. METHODS: We performed comprehensive small RNA and RNA profiling on colon tissue from IBD and NIBD control patients in addition to characterizing the serum cleaved ectodomain of CADM1 (sCADM1) function in lamina propria mononuclear cells isolated from these patients. Last, a conditional loss-of-function mouse was developed to assess Cadm1 function in the myeloid compartment during chemical-induced colitis. RESULTS: We identified CADM1 enrichment in multiple immune cell clusters including macrophages and dendritic cells in the colons of IBD patients. Increased numbers of CADM1+ myeloid cells were measured adjacent to CD8+ T cells within colons of ulcerative colitis patients compared with NIBD patients. Conditional deletion of Cadm1 in myeloid cells resulted in reduced numbers of activated T cell populations and protected mice from chemical-induced colitis. Similarly, administration of a Cadm1 "neutralizing" antibody, which binds its extracellular domain reduced tissue inflammation and breakdown of the intestinal epithelium and crypts after induction of colitis in mice. Last, serum levels of sCADM1 were elevated in IBD patients compared with NIBD control subjects and treatment of lamina propria mononuclear cells with recombinant sCADM1 enhanced inflammatory STAT3 phosphorylation. CONCLUSIONS: CADM1 is a mediator of proinflammatory signaling cascades in the colon and a potential therapeutic target for the IBDs.

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BACKGROUND: Microscopic colitis (MC), comprising lymphocytic colitis (LC) and collagenous colitis (CC), is an inflammatory bowel disease with increasing incidence. MC etiopathogenesis remains unknown; however, altered colonic epithelial integrity may underlie uncontrolled luminal antigen passage, triggering immuno-inflammatory responses. OBJECTIVE: The aim of this study was to further define the involvement of the colonic epithelium in MC. METHODS: A paired transcriptomic and proteomic analysis followed by epithelial ultrastructural examination was performed on colonic biopsies from LC and CC patients, and from irritable bowel syndrome with a predominance of diarrhoea (IBS-D) and healthy subjects (H) as control groups. The impact of budesonide therapy on the epithelial structure was also evaluated in CC. RESULTS: MC patients exhibited decreased expression of inter-microvilli adhesion and actin-bundling proteins, accompanied by increased expression of actin-membrane connection proteins compared to both control groups. Distinct molecular differentiated CC and LC, which translated into differential ultrastructure abnormalities. The colonic microvilli in CC patients were shorter in length and fewer in number, with partial restoration following budesonide treatment, whereas LC showed a reduction solely in microvilli number. A negative correlation was found between daily stool frequency and SPATN1 and ATP8B1 protein levels in CC patients. CONCLUSIONS: Molecular dysregulation and aberrant ultrastructure of the colonic brush border feature the colonic epithelium in LC and CC. These previously undescribed findings provide new perspectives for further defining MC pathogenesis and identifying biomarkers for diagnosis, prognosis and treatment of this debilitating and prevalent disease.

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Ulcerative colitis (UC) is a chronic inflammatory disease of the gastrointestinal tract, characterized by ongoing intestinal inflammation, epithelial damage, and mucosal injury. Despite the identification of C-C motif chemokine ligand 5 (CCL5) as a key mediator in UC, the precise mechanisms underlying its role in immune activation and inflammation remain unclear. This study aimed to investigate CCL5 as a critical immune modulator in UC, focusing on its effects on immune cell activation, particularly regulatory T cell (Treg) formation, and the molecular pathways involved in these processes. Using the dextran sulfate sodium salt (DSS)-induced UC model and CCL5 knockout (Ccl5-KO) mice, we demonstrated that CCL5 deficiency exacerbates intestinal inflammation during the acute phase of colitis, partly due to impaired interleukin-33 (IL-33)-induced Treg formation. In addition, we observed a positive correlation between CCL5 expression and forkhead box protein 3 (FOXP3) levels in inflamed colon tissues of UC patients, suggesting a role for CCL5 in Treg regulation. Mechanistically, CCL5 deficiency disrupted the PI3K/Akt/NF-κB signaling pathway, resulting in reduced IL-33 expression, which in turn impaired CD4+ T cell activation and FOXP3+ Treg formation via the JAK1/STAT5 pathway. In vivo rescue experiments confirmed that restoring IL-33 signaling could alleviate inflammation and partially recover Treg function. Collectively, these findings highlight CCL5 as a novel immune modulator of Treg formation and immune responses in UC and suggest that targeting CCL5 may offer a promising therapeutic strategy for managing UC and related inflammatory diseases.

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The colon immune activity of high branched rhamnogalacturonan-I type pectin from Lycium barbarum fruit (LBP-P4) was unclear. This study comparatively investigated the colon immunity activity of LBP-P4 and inulin and explored its mechanism via simulated colon biological and mechanical barriers models. LBP-P4 could be slowly degraded and utilized by intestinal microorganisms with degradation rate of 88.88 %. Meanwhile, LBP-P4 increased the relative abundance of Bifidobacterium, especially Bifidobacterium longum subsp. longum, which was confirmed by single Bifidobacterium fermentation. In the normal colon mechanical barrier coupled with macrophage model, the colonic metabolites of LBP-P4 enhanced macrophage viability and phagocytic ability. In the damaged colon mechanical barrier coupled with macrophage model, the colonic metabolites of LBP-P4 enhanced mechanical barrier integrity, reduced the production of NO and IL-6 and improved macrophage viability and phagocytic ability. The colon immunity activity of LBP-P4 was stronger than that of inulin, which was because the contents of immunity related metabolites i.e. short chain fatty acids and tryptophan metabolites, were significantly higher (about 1-3 fold) in LBP-P4 group than those in inulin group. Our findings provided new insights into the probiotic and intestinal immunity function of LBP-P4 and offered theoretical basis for the development of functional products of LBP-P4.

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Toxoplasma gondii (T. gondii) impairs gut barrier integrity in hosts, which helps the establishment of long-term infection and is closely associated with disease progression. However, the precise mechanisms remain largely unclear. This study investigated whether pyroptosis, a form of inflammatory programmed cell death, contributes to the impairment of colonic barrier integrity induced by T. gondii chronic infection in mice. We found that the infection resulted in pronounced pathological damage in the colon, characterised by reduced colon length, decreased expression of tight junction proteins, and elevated serum LPS levels. Interestingly, the infection activated the pyroptosis pathway, supported by the higher expression of NLRP3 (NOD-like receptor thermal protein domain associated protein 3), Caspase-1, gasdermin D (GSDMD), GSDMD-N, as well as IL-1ß and IL-18. In addition, dimethyl fumarate, an anti-pyroptogenic drug, significantly attenuated the infection-induced pyroptosis and colonic pathology. Notably, macrophages exhibited pyroptosis post T. gondii infection, which was attenuated by DMF treatment. Overall, our results uncover that pyroptosis is vital for the impairments of gut barrier integrity post T. gondii chronic infection.

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BACKGROUNDFecal microbiota transplantation (FMT) is the most effective therapy for recurrent Clostridioides difficile infection (rCDI), yet its mechanism of action remains poorly understood.METHODSWe report the results of a clinical trial of patients undergoing FMT therapy for rCDI (n = 16), which analyzed colon biopsies, plasma, PBMCs, and stool at the time of FMT and 2-month follow-up. Plasma and colon biopsy samples were also collected from healthy controls for comparison with patients with rCDI. Microbiome composition, colonic gene expression, and immune changes were evaluated through high-throughput sequencing and immunoprofiling via flow cytometry.RESULTSNo patients experienced recurrence at follow-up. FMT significantly altered the intestinal microbiome but had no significant impact on the systemic immune system. In contrast, FMT promoted broad changes in colonic transcriptional profiles compared with both pre-FMT and healthy control biopsies, inhibiting genes associated with proinflammatory signaling and upregulating type 2 immunity and proliferative pathways (Myc and mTORC1). FMT increased expression of IL-33 and the type 2 immune EGFR family ligand amphiregulin, potentially explaining upregulation of Myc and mTORC1 pathways. Spatial transcriptomics demonstrated that these changes were localized to the colonic epithelium. Comparison of transcriptional profiles with available single-cell gene sets determined that post-FMT biopsies were enriched in signatures associated with proliferative cell types while repressing signatures of differentiated colonocytes.CONCLUSIONWe conclude that FMT promotes proliferation of the colonic epithelium in patients with rCDI, which may drive regeneration and protect against subsequent CDI.TRIAL REGISTRATIONClinicaltrials.gov NCT02797288.FUNDINGThis work was funded by grants from the NIH.

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Single-cell studies have revealed that intestinal macrophages maintain gut homeostasis through the balanced actions of reactive (inflammatory) and tolerant (noninflammatory) subpopulations. How such balance is impaired in inflammatory bowel diseases (IBDs), including Crohn's disease (CD) and ulcerative colitis (UC), remains unresolved. Here, we define colon-specific macrophage states and reveal the critical role of noninflammatory colon-associated macrophages (niColAMs) in IBD recovery. Through trans-scale analyses-integrating computational transcriptomics, proteomics, and in vivo interventional studies-we identified GIV (CCDC88A) as a key regulator of niColAMs. GIV emerged as the top-ranked gene in niColAMs that physically and functionally interacts with NOD2, an innate immune sensor implicated in CD and UC. Myeloid-specific GIV depletion exacerbates infectious colitis, prolongs disease, and abolishes the protective effects of the NOD2 ligand muramyl dipeptide in colitis and sepsis models. Mechanistically, GIV's C-terminus binds the terminal leucine-rich repeat 10 (LRR 10) of NOD2 and is required for NOD2 to dampen inflammation and clear microbes. The CD-associated 1007fs NOD2 variant, which lacks LRR 10, cannot bind GIV, which provides critical insights into how this clinically relevant variant impairs microbial sensing and clearance. These findings illuminate a critical GIV•NOD2 axis essential for gut homeostasis and highlight its disruption as a driver of dysbiosis and inflammation in IBD.

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BACKGROUND: The mechanisms by which microbiota from disease-resistant populations or animals improve intestinal immune defense remain incompletely elucidated. Tibetan pig, a renowned disease-resistant breed, serve as a valuable research subject for the health of humans and economic animals. RESULTS: In this study, fecal microbiota transplantation from Tibetan piglets into mice conferred enhanced resistance to C. rodentium DBS100. Further microbiota profiling and metabolomics analysis showed this protection may be partly ascribed to C. butyricum SLZX19-05 in recipients' colon. Administration of C. butyricum SLZX19-05 to germ-free mice resulted in the significantly increased lysozyme expression within colonic macrophages, subsequently bolstering the resistance to C. rodentium infection. In mice and piglets, this C. butyricum similarly elevated the lysozyme level in colon and decreased diarrhea incidence. Conversely, lyz1-knockout heightened mice's susceptibility to C. rodentium, highlighting lysozyme's critical role in immune defense. Mechanistically, this study systematically revealed that C. butyricum enhanced lysozyme expression by inhibiting mTORC1-HDAC3/8 pathway, leading to the increased H4K31 Crotonylation (H4K31Cr) and openness of an upstream region of lyz1 promoter via butyrate in macrophages. Additionally, H4K31-mutant mice showed the leukopenia, further validating the significance of H4K31Cr in immune regulation. CONCLUSIONS: Collectively, mTORC1-HDAC3/8-H4K31Cr pathway is a key mechanism by which butyrate-producing commensal bacteria enhance immune defense in gut. This discovery provides a novel foundation for the screening and application of the next generation of butyrate-producing probiotics. Video Abstract.

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Background: Akkermansia muciniphila (A. muciniphila) is a mucin-degrading commensal bacterium with established roles in maintaining intestinal homeostasis and modulating host immune responses. However, its therapeutic potential and mechanisms in chronic spontaneous colitis remain incompletely defined. Interleukin-10 knockout (IL-10-/-) mice are a well-established model of spontaneous colitis due to impaired anti-inflammatory cytokine signaling and disrupted immune regulation. In this study, we aimed to determine whether A. muciniphila and its culture-derived supernatant could ameliorate intestinal inflammation in IL-10-/- mice. Methods: We administered A. muciniphila or its culture supernatant (A. muciniphila supernatant [A.m-SN]) to IL-10-/- mice and monitored disease progression over 30 days. Clinical assessments included body weight, disease activity index (DAI), colon length, histopathology, and endoscopy. Tight junction proteins (zonula occludens-1 [ZO-1] and Occludin) and colonic messenger ribonucleic acid levels of pro- and anti-inflammatory cytokines were evaluated. Gut microbiota composition was assessed using 16S ribosomal deoxyribonucleic acid sequencing, and untargeted metabolomic profiling was performed on the culture medium to identify potential anti-inflammatory components. Results: Mice treated with A. muciniphila or A.m-SN showed significantly reduced colitis severity, including reduced body weight loss, lower DAI, preserved colon length, and improved histological scores. Immunofluorescence revealed increased expression of ZO-1 and Occludin in colonic tissues. Pro-inflammatory cytokines (tumor necrosis factor-α, IL-6, IL-17, IL-1ß, and interferon-γ) were downregulated, while anti-inflammatory cytokines (IL-4, transforming growth factor-ß, and IL-22) were upregulated in treated groups. Gut microbiota analysis showed partial restoration of microbial diversity and structure. Metabolomic profiling identified distinct signatures in A.m-SN, with bioactive molecules such as adenosine and tryptophan derivatives implicated in the anti-inflammatory effects. Conclusion: The culture supernatant of A. muciniphila alleviated spontaneous colitis in IL-10-/- mice by suppressing pro-inflammatory cytokines, enhancing anti-inflammatory mediators, and strengthening epithelial barrier function. Metabolomic profiling identified candidate bioactive components, including adenosine and tryptophan derivatives, supporting its potential as a postbiotic intervention for inflammatory bowel disease.

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Ulcerative colitis (UC) is a severe inflammatory bowel disease affecting millions of people worldwide, but the factors driving the condition are poorly understood. In tissue samples from individuals with UC, we found that macrophages were depleted from areas of the colon that did not yet exhibit overt epithelial inflammation. We hypothesized that toxins produced by bacteria could impair macrophages and that this could promote wider inflammation. We isolated a variant of Aeromonas genus from stool samples from UC patients, which we termed macrophage-toxic bacteria (MTB), because aerolysin secreted by MTB caused macrophage death. MTB colonized mice under pathogenic conditions and triggered colitis. Antibodies against aerolysin alleviated colitis induced by Aeromonas in mice. In a cohort, UC patients more frequently tested positive for Aeromonas than healthy controls did.

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The intestine is a highly compartmentalized organ, with distinct segments exhibiting both varying susceptibilities and responses to enteric pathogens, although the cellular and molecular bases of these responses remain elusive. Here, we used Salmonella Typhimurium (S. Tm), a prominent enteric pathogen that causes human colitis, to establish a murine model of Salmonella enterocolitis. By integrating bulk RNA-seq, single-cell RNA-seq, and spatial RNA-seq data, we present a comprehensive spatiotemporal single-cell transcriptomic landscape of the colon over a week-long time course of infection. We identified the distal colon as the intestinal segment where most of the host responses were initiated, with distal colonocytes (DCCs) being the most responsive epithelial cells upon the onset of infection. Furthermore, by correlating our findings with human intestinal single-cell transcriptome data, we identified a human colonocyte population that shares many characteristics with murine DCCs. Our study advances the understanding of the cellular and molecular basis of compartmentalized intestinal responses to pathogenic insults and may pave the way for novel preventive and therapeutic strategies to mitigate intestinal damage and combat intestinal infections.

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To study the roles of bacterial secretome in immune cell polarization and gut mucosal homeostasis, we developed a 3-dimensional co-culture model using commercially available inserts in a back-to-back stacked format. We termed this system 3D Flipwells, and successfully co-cultured bacteria, colon epithelial, and immune cells in stratified layers to recapitulate the gut mucosal microenvironment. This co-culture system enables the seeding of colon epithelial cells on one side of the membrane and undifferentiated non-adherent monocytes on the other, allowing for the synchronous progression of colon epithelial polarization, mucus formation, and monocyte-macrophage differentiation and polarization within a single co-culture without the need for detaching and replating pre-differentiated macrophages. This system is easy to construct and only requires a few readily available materials and tools. We tested its utility by analyzing the responses of co-culture components to sepiapterin (SEP), the endogenous precursor of BH4 (tetrahydrobiopterin) -- a cofactor of nitric oxide synthase (NOS). We demonstrated that SEP treatment of 3D Flipwell induced biofilm formation by gut bacteria, mucus production by colon epithelium, and pro-immunogenic polarization of macrophages. This indicated that SEP triggered activation of a mucosal defense mechanism through crosstalk between different cellular components. Further experiments are needed to investigate the role of mucosal communications that lead to immune cell reprogramming and to evaluate the utility of this co-culture system to screen for novel immunomodulatory therapies.

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OBJECTIVE: This study aims to elucidate how butyrate, a short-chain fatty acid, regulates the Treg/Th17 balance in ulcerative colitis (UC) via the cAMP-PKA/mTOR signaling pathway, offering novel treatment strategies. METHODS: Dextran sulfate sodium (DSS) was used to induce ulcerative colitis in a mouse model. Various butyrate dosages were administered to the mice. The mice's body weight, colon length, spleen index (SI), and disease activity index (DAI) were all assessed.HE staining and Masson staining were used for histopathological evaluation. Immunohistochemistry and RT-qPCR were applied to detect fibrosis markers. Flow cytometry, RT-qPCR, and ELISA were employed to analyze immune cell subsets and cytokines. RT-qPCR and Western blotting were utilized to explore the cAMP-PKA/mTOR signaling pathway. Specific inhibitors were used to further confirm the mechanism of its action. RESULTS: Butyrate treatmentreduced DAI and SI, and reversed pathological changes (weight loss, colon shortening, splenomegaly) in DSS-induced UC model mice, with the high-dose group showing the best recovery. It inhibited colon fibrosis,and decreased fibrosis markers. By regulating the regulatory T cell (Treg)/T helper 17 cell (Th17) balance, butyrate restored immune homeostasis. Flow cytometry showed DSS-induced immune imbalance was reversed in a dose-dependent manner. Additionally, butyrate modulated the cAMP-PKA/mTOR signaling pathway, reversing DSS-induced gene and protein expression changes. Specific inhibitor experiments confirmed that butyrate exerted its therapeutic effects via this pathway. CONCLUSION: Butyrate can markedly alleviate acute UC intestinal inflammation and block chronic fibrosis progression. The bidirectional regulation of the cAMP-PKA/mTOR signaling pathway is the key mechanism for butyrate to restore immune homeostasis.

Butyrate dose-dependently improves the pathological state of UC mice, reducing DAI and SI and restoring colon length.Butyrate effectively alleviates acute-phase inflammation in UC, blocks chronic fibrosis, and inhibits collagen proliferation and the expression of fibrosis markers.Butyrate regulates theTreg/Th17 balance and reshapes immune homeostasis by bidirectionally regulatingthe cAMP-PKA/mTOR signaling pathway.

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The intestinal epithelium consists of a single layer of cells that includes enterocytes, enteroendocrine and goblet cells. This monolayer is in close contact with the underlying lamina propria and spatially segregates the gut microbiota and other environmental stimuli from the mucosal immune system. Under certain circumstances, however, gut bacterial components and/or metabolites can perturb tissue homeostasis resulting in abnormal immune responses and tissue damage. In this context, some bacterial toxins, besides stimulating an inflammatory response, may contribute to the activation of pathways related to carcinogenesis. Since immunosurveillance represents a critical barrier to an emerging tumor, characterizing tissue resident immune cells can prove a relevant tool for translational research. The protocol described herein describes the required steps for the isolation and flow cytometry phenotype analysis of tissue resident immune cells from mouse colon following repeated exposure to a pathogenic E. coli toxin as model stimulus.

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Beetroot has been traditionally used in folk medicine to treat various gastrointestinal disorders and is commonly consumed for its high antioxidant content. The primary objective of this study is to demonstrate, for the first time, the potential therapeutic effects of beetroot juice (BJ) in a rat model suffering from ulcerative colitis (UC) induced by dextran sulfate sodium (DSS). In addition, the study aims to identify the bioactive compounds in beetroot responsible for these effects. UC was induced in rats through the oral administration of DSS dissolved in drinking water (5%) during the final week of a 21-day treatment with BJ (5 and 10 mL/kg body weight) and compared to a reference drug, mesalazine. Phytochemical analysis revealed that BJ is a rich source of secondary metabolites such as polyphenols, flavonoids, soluble sugars, and betalains. The pretreatment with BJ significantly and dose-dependently reduced DSS-induced colonic mucosa lesions and associated histopathological alterations. The protective effects of BJ on the colonic mucosa were closely linked to its ability to mitigate oxidative damage, inflammatory injury, and biochemical imbalances. Specifically, BJ counteracted DSS-induced lipid peroxidation and restored levels of both enzymatic and nonenzymatic antioxidants. Moreover, BJ significantly decreased the levels of inflammatory markers and cytokines in plasma and colonic mucosa, while enhancing plasma scavenging activity and maintaining the homeostasis of intracellular mediators such as hydrogen peroxide, free iron, and ionizable calcium. These findings supported the use of BJ as a complementary approach for managing UC and underscore the need for further investigation into its potential clinical applications.

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Enteric neurons are essential regulators of intestinal physiology, yet their responses to varying microbial and immune environments along the intestinal tract and or during challenges remain poorly understood. In this study, we regionally profiled enteric neurons across gnotobiotic, allergic, and parasite-infected mice. Timing and complexity of microbial perturbations and type 2 inflammation result in motor neuron state shifts and alter multiple functionally distinct sensory neurons, including interleukin-13- and leukotriene-responsive Nmu-hi cells and Grp-hi neurons, which expand in germ-free colonic tissue and interact with Grpr+ interstitial cells of Cajal. Leveraging adeno-associated virus-based Perturb-seq, we identified Edf1 and Mitf as controllers of motor neuron state transition and gastrointestinal transit time, directly linking enteric neuron states to physiology.

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BACKGROUND AND AIMS: Colitis-associated cancer (CAC) is the most severe complication of inflammatory bowel disease (IBD). We hypothesized that chronic inflammation activates endogenous anti-inflammatory mechanisms that promote dysplasia by undermining immunosurveillance. Our aim was to determine chronic inflammation-induced immune cell reprogramming in IBD patients at risk for developing CAC. METHODS: This cohort study utilized GeoMx digital spatial profiling and imaging mass cytometry in 11 patients with either CAC or sporadic colorectal cancer (SCRC). Results from this discovery cohort were validated using immunohistochemistry/immunofluorescence in an independent cohort of CAC and SCRC patients (n = 10 and n = 14, respectively), as well as in an independent cohort of IBD patients with (n = 6) and without dysplasia (n = 18). RESULTS: Histologically uninflamed colon from patients who developed CAC displayed upregulated metabolism and stress response pathways as compared to SCRC patients, indicating ongoing epithelial stress-responses. Endogenous anti-inflammatory mechanisms included increased IL-10 expression by lamina propria IgA+ plasma cells and CD163+ macrophages. T cell recruitment and effector pathways were downregulated in CAC, which was associated with a decrease in CD8+ intraepithelial T cells (IELs) and reduced levels of granzyme B within CD8+ IELs. Decreased CD8+ IEL density was associated with CAC susceptibility, as IBD patients who developed dysplasia showed significantly lower levels of CD8+ IELs than IBD patients who never developed dysplasia. CONCLUSIONS: Chronic inflammation induces endogenous mechanisms to protect from inflammation-induced damage, including increased anti-inflammatory cytokine production and decreased levels of CD8+ IELs. While this may limit inflammation, these mechanisms may also reduce immunosurveillance, favoring the development of CAC.

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Introduction: Crohn's disease (CD) is a clinical subset of inflammatory bowel disease that is characterized by patchy transmural inflammation across the gastrointestinal tract. Although the exact etiology remains unknown, recent findings suggest that it is a complex multifactorial disease with contributions from the host genetics and environmental factors such as the microbiome. We have previously shown that the T cell repertoire of individuals with CD harbors a group of highly expanded T cells which hints toward an antigen-mediated pathology. Methods: We simultaneously profiled the αß and γδ T cell repertoire in addition to the B cell repertoire of both the blood and the colonic mucosa of 27 treatment-naïve individuals with CD and 27 age-matched symptomatic controls. Results: Regardless of disease status, we observed multiple physiological differences between the immune repertoire of blood and colonic mucosa. Additionally, by comparing the repertoire of individuals with CD relative to controls, we observed different alterations that were only detected in the blood or colonic mucosa. These include a depletion of mucosal-associated invariant T (MAIT) cells and an expansion of TRAV29/DV5-TRAJ5 + clonotypes in the blood repertoire of individuals with CD. Also, a significant depletion of multiple IGHV3-33-IGHJ4+ and IGHV3-33-IGHJ6+ clonotypes in the blood and gut IGH repertoire of individuals with CD. Discussion: Our findings highlight the importance of studying the immune repertoire in a tissue-specific manner and the need to profile the T and B cell immune repertoire of gut tissues as not all disease-induced alterations will be detected in the blood.