Inflammatory Bowel Diseases

IntroductionIntestinal barrier failure is a key characteristic of both kinds of inflammatory bowel disease (IBD), ulcerative colitis (UC) and Crohns disease (CD). The intestinal barrier, or gut mucosal barrier, is composed of mucus and a tightly interconnected single layer of intestinal epithelial cells (IEC) that line the gut. Together, these components work to contain the gut microbiota within the gut lumen and when they fail, microbes, microbial products, or microbial components cause damage, inflammation, and immune activation in host tissue. We previously reported that High mobility group box 1 (HMGB1) in colonic mucus aggregates bacteria, limits bacterial invasion through mucus, and prevents bacteria from adhering to host tissue. Epithelial surface-associated HMGB1 is decreased in active UC lesions and low levels of HMGB1 are associated with high levels of tissue-adherent bacteria expressing adhesins carrying the molecular target of HMGB1 (ToH1). The study reported here was designed to determine whether HMGB1 defense is also compromised in active lesions from CD patients. MethodsImmunofluorescence microscopy was used to visualize mucus and HMGB1 in tissue from colonic resections performed in CD and non-IBD control patients. ResultsActive CD lesions had areas where the IEC were absent or pulling away from underlying tissue along with areas of increased mucus thickness and goblet cells full of mucus highly positive for alpha-linked-fucose residues. The surface associated HMGB1 was also decreased in active CD lesions. ConclusionTissue from CD patients exhibited cellular and acellular intestinal barrier defects in comparison to control patients. We observed the previously reported loss of IEC barrier integrity and abnormalities in the amount and distribution of mucus in CD lesions. We also report for the first time that CD is associated with decreased HMGB1 defense at the colon surface.

Background and AimsStem cell-derived organoids are promising platforms for therapeutic screening in inflammatory bowel disease (IBD), but identifying functional organoid readouts with translational utility is challenging. Colon epithelial organoids from patients with ulcerative colitis (UC) overexpress chemokines CXCL1, CXCL11, CCL2, and CCL28, yet whether these inflammatory signatures correlate with disease activity and treatment response is unknown. This short report investigates whether organoid-retained chemokines correlate with disease activity and therapeutic outcomes. MethodsWe interrogated three bulk and two single-cell transcriptomic datasets from IBD clinical trials encompassing anti-TNF and anti-integrin therapies to determine whether epithelial chemokines retained in UC organoids track clinical response and distinguish treatment responders from non-responders to biologic therapy across multiple IBD patient cohorts. ResultsIn bulk transcriptomic data, CXCL1, CXCL11, and CCL2 were elevated in active UC and normalized only in patients achieving clinical remission, independent of therapy class, with persistent chemokine overexpression in non-responders. Single-cell analysis demonstrated widespread chemokine overexpression in UC epithelial clusters, with consistent normalization of CXCL1, CXCL11, and CCL28 in LGR5-positive stem compartment of patients who achieved clinical remission, but not in non-responders. In Crohns disease, the resolution of these epithelial chemokines was not associated with clinical response. ConclusionsEpithelial chemokines, particularly CXCL1, CXCL11, and CCL28, track clinical remission in UC and represent candidate biomarkers and functional endpoints for epithelial-directed therapeutic strategies using stem cell-derived UC organoid models.

BackgroundVedolizumab, a gut-selective anti-integrin therapy, is effective in IBD, but response rates remain variable. Conventional clinical and biochemical markers, including C-reactive protein and faecal calprotectin, have limited predictive value. Although recent transcriptomic studies have implicated T-cell-related signatures in predicting vedolizumab response, these findings lack validation across independent cohorts. MethodsWe analyzed pre-treatment transcriptomic profiles from whole blood and T-cell subsets across five independent cohorts comprising 100 patients with UC and CD. The primary outcome was clinical response. Secondary outcomes included clinical and biochemical remission. ResultsAmong the 100 patients, 61 were responders and 39 non-responders, with no significant baseline clinical differences. Gene set enrichment analyses revealed downregulation of interferon alpha and gamma signalling in responders baseline blood samples, a finding validated across independent cohorts. Downregulated interferon signalling at baseline was also observed in patients who achieved clinical and biochemical remission. To build a predictive model, an adaptive elastic net logistic regression model was applied to baseline whole-blood RNA-sequencing data. The classifier achieved an AUC of 1.0 in training, 0.71-0.83 in UC validation cohorts, and 0.64-1.0 in CD cohorts. Reduced interferon signalling was observed across CD4{square} and CD8{square} T-cell subsets, including regulatory T cells, suggesting a broad immune signature rather than cell-type specificity. ConclusionsDownregulated interferon signalling in peripheral blood prior to treatment is a reproducible molecular signature predictive of vedolizumab response and biochemical remission. Whole-blood transcriptomics revealed a robust interferon-axis signal that predicted vedolizumab response across independent cohorts, with stronger performance in UC than CD. Given heterogeneous clinical endpoints and assessment windows, these data provide proof-of-concept that warrants validation with standardised, endoscopy-based outcomes.

BackgroundTryptophan (Trp) metabolism is a central immunometabolic axis in inflammatory bowel disease (IBD) and has been linked to inflammatory activity and immune regulation. While individual Trp metabolites have been associated with disease severity and treatment response, systems-level frameworks to define metabolic subtypes in IBD are lacking. ObjectiveTo identify reproducible Trp-related metabolic subtypes ("metabotypes") in IBD and assess their association with disease activity, clinical outcomes, and early disease development. DesignWe applied unsupervised clustering to serum concentrations of 16 Trp-related metabolites in a discovery cohort of patients with IBD undergoing biologic induction therapy (n=134). Metabotypes were validated in three independent IBD cohorts (total n>2,800), a healthy reference population, and a prospective cohort of first-degree relatives at risk for Crohns disease. Associations with disease activity, longitudinal outcomes, and metabolic pathways were assessed using multivariable regression and survival analysis. ResultsFour reproducible metabotypes with distinct metabolite profiles were identified across cohorts: Low Kyna, High Kyna, High Quin, and Balanced. Low Kyna and High Quin metabotypes were consistently associated with increased inflammatory activity and adverse clinical outcomes, including increased risk of treatment escalation and disease progression. Pathway-level analyses revealed alterations in NAD-related, lipid, and amino acid pathways between inflammatory metabotypes. A metabotype resembling inflammatory disease states was enriched in individuals who later developed Crohns disease in a prospective pre-disease cohort. ConclusionTrp-linked metabotypes define reproducible immunometabolic states in IBD that associate with disease activity and clinical outcomes and may precede disease onset. These findings provide a framework for metabolic stratification and biomarker-guided clinical trials targeting immunometabolic pathways. What is already known on this topicTryptophan metabolism through the kynurenine pathway is a central immunometabolic axis in inflammatory bowel disease (IBD) and has been linked to inflammatory activity and immune regulation. Individual tryptophan metabolites have been associated with disease severity and treatment response, but their clinical utility for patient stratification remains limited. Systems-level approaches to define clinically meaningful metabolic subtypes in IBD are lacking. What this study addsWe identify four reproducible tryptophan-related metabolic subtypes ("metabotypes") that are consistently associated with disease activity across multiple independent IBD cohorts. Inflammation-associated metabotypes show distinct pathway-level alterations, including differences in NAD-related metabolism and broader metabolic programs. A metabotype resembling inflammatory disease states is detectable before clinical diagnosis in individuals who later develop Crohns disease. How this study might affect research, practice or policyMetabotype-based classification provides a framework for molecular stratification of patients in mechanistic studies and clinical trials targeting immunometabolic pathways. This approach may support biomarker-guided monitoring of disease activity and disease progression in IBD. Identification of preclinical metabolic states highlights the potential of metabolomics for early disease detection and prevention-oriented research strategies.

Background: Histological examination of mucosal tissue in inflammatory bowel diseases (IBD) is a sensitive tool to measure disease activity, and histological remission is emerging as a potentially important treatment target. There are several existing histopathological indices, but they often encompass caveats such as not primarily having been designed to measure the degree of inflammation, encompassing subjective components with poor intra- and interindividual reproducibility, and requiring expert pathologists who are scarce, thus resulting in extended response times. Aim: To construct a new computerized, automated index to objectively measure histological disease activity in the ileal and colonic mucosa, applicable to both Crohn's disease (CD) and ulcerative colitis (UC). Materials and methods: Ileocolonic biopsies were collected from control subjects and patients with CD or UC. A group of CD patients was sampled before and after 12 weeks of anti-TNF therapy. Another group of CD and UC patients functioned as a small validation cohort. Epithelial cells, neutrophils, macrophages, and T cells were immunohistochemically stained, followed by digitalization of the color signal and computerized delineation of the epithelial and lamina propria compartments. The various immune cell types within the epithelium and the lamina propria, respectively, were enumerated, and the numbers were compared between control subjects and patients with CD or UC. Results: The numbers of neutrophils and macrophages in the epithelium, and neutrophils in the lamina propria, showed the highest sensitivity and specificity for distinguishing control-subject tissues from CD and UC tissues. These three parameters were thus chosen to construct a new index, named QiC3 1.0, that could separate tissues from control subjects and patients with CD or UC with high precision. It performed equally well in a small validation cohort of patients. The QiC3 index correlated well with previously described histopathological indices, fecal calprotectin, and endoscopic scores in UC, but showed worse correlation with endoscopic scores in CD and symptomatic scores. When applying the new index to tissues from CD patients before and after therapy, it showed good responsiveness, demonstrating a distinct amelioration in the microscopic inflammatory status that corresponded well to improvements in histopathological scores. Conclusion: We describe a new quantitative, computerized, automated, non-subjective, and response-sensitive immunohistological index (QiC3) for measuring disease activity in ileal and colonic mucosal biopsies, suitable for both CD and UC.

Background and Aims: Fatigue is a prevalent and disabling symptom in inflammatory bowel disease (IBD), yet its underlying biological mechanisms remain poorly understood. We aimed to characterize fatigue-associated molecular signatures in IBD patients by integrating DNA methylation and mRNA expression analyses. Methods: Peripheral blood was collected from 40 patients with Crohn's disease (CD), 29 with ulcerative colitis (UC), and 10 healthy controls. Fatigue severity was assessed continuously using the Multidimensional Fatigue Inventory (MFI). Epigenome-wide DNA methylation profiling and mRNA sequencing were performed, identifying differentially methylated regions (DMRs) and differentially expressed genes (DEGs) for active and quiescent CD and UC, adjusting for age, sex, and smoking status. Pathway enrichment analysis was performed on genes with differential methylation and expression. Results: In active CD, more severe fatigue was associated with transcriptional suppression of immune and metabolic pathways (246 DMRs; 1,090 DEGs), versus upregulation of mitochondrial and metabolic processes in quiescent CD (200 DMRs; 1,619 DEGs). In active UC, fatigue was associated with anabolic pathway upregulation and epigenetic silencing of neuroactive pathways (6,927 DMRs; 343 DEGs; 56 concordant genes). Quiescent UC showed transcriptional changes without significant epigenetic pathway enrichment (1,710 DMRs; 3,224 DEGs). Healthy controls exhibited a distinct profile spanning metabolic, immune, and neuronal pathways (8,621 DMRs; 395 DEGs). Fatigue-associated signatures were largely non-overlapping across all five groups. Conclusions: Fatigue-associated molecular profiles differed substantially by disease subtype and activity state, highlighting the biological heterogeneity of IBD-related fatigue and laying the foundation for multi-omics approaches to identify biomarkers and potential therapeutic targets.

Background and Aims: Diet plays a critical role in managing Crohns disease (CD) inflammation. We assessed whether dietary replacement of animal protein (AnimalP) by soy-pea protein (SoyP) decreases the pro-inflammatory potential of gut microbiota and intestinal inflammation in CD patients. Design: In an open-label, randomized controlled feeding trial at University Hospitals Cleveland Medical Center, CD participants and healthy controls were randomized (1:1) to a soy-pea or animal protein diet for 7-days. Primary outcomes were the absolute difference (d7-d0) in; Crohns Disease Activity Index (CDAI) score and fecal myeloperoxidase (MPO). Secondary outcomes included fecal calprotectin (FC) and high-sensitivity C-reactive protein (hsCRP). Murine fecal transplantation experiments were performed to determine the inflammatory potential of diet-altered gut microbiota. Results: The study randomized 66 participants and 60 were included in the final analysis (n=31 CD, n=29 HC). After 7 days, CD-SoyP participants were more likely than CD-AnimalP to show reductions in HBI (RR=4.68, 95% CI: 1.22-17.98, P=0.009) and fecal MPO (RR=2.30, 95% CI: 1.04-4.85, P=0.032), with a similar directional trend for CDAI (RR=1.52, 95% CI: 0.89-2.58, P=0.135). No participants experienced worsening of CDAI. The rank-based composite CDAI-MPO score was lower in the CD-SoyP vs CD-AnimalP group (median [IQR]: 5 [4-6] vs 8 [7-9]; P=0.012). Stratified analyses showed significant reductions in fecal MPO among CD participants with lower baseline disease activity (CDAI <150; P<0.0001), but not in those with higher activity (P=0.799) Conclusion: Short-term addition of plant-based soy-pea protein within a controlled diet exerted a beneficial, anti-inflammatory effect in CD, with evidence of greater effects among participants with lower baseline disease activity. ClinicalTrials.gov, Number NCT04065048.

Malnutrition is common in Crohns disease (CD), and its assessment requires multiple tools. Comprehensive evaluation of nutritional status in a population with CD, predominantly characterized by metabolic phenotype, was inadequately reported. This study evaluated the nutritional status of CD patients using anthropometric, clinical, and biochemical measures and compared patients with active disease with those in remission. This cross-sectional study included 127 adults with CD: 63 with active disease and 64 in remission. Disease activity was classified using the Crohns Disease Activity Index, the Simple Endoscopic Score for Crohns Disease, and magnetic resonance enterography. Nutritional assessment included body mass index (BMI), mid-upper arm circumference, calf circumference, triceps skinfold thickness, mid-arm muscle circumference, Mini Nutritional Assessment-Short Form (MNA-SF), and biochemical markers including hemoglobin, serum iron, folate, vitamin B12, albumin, and zinc. Malnutrition was defined using the Global Leadership Initiative on Malnutrition criteria. Overall, 47.2% of participants were malnourished. Malnutrition was significantly more frequent in active disease than in remission (81.0% vs. 14.1%, P<0.001). Patients with active CD had lower anthropometric indices, MNA-SF score, hemoglobin, serum iron, albumin, and zinc (all P<0.001), whereas folate and vitamin B12 did not differ significantly. BMI showed positive correlations with other anthropometric measures and MNA-SF score (r=0.854-0.914, all P<0.001), whereas correlations with biochemical parameters were weaker and disappeared after subgroup stratification. Overall, the findings indicate that malnutrition is highly prevalent in CD, particularly during active disease. Anthropometric measures and MNA-SF were strongly concordant, whereas biochemical markers were less consistent, supporting a multidimensional nutritional assessment approach in CD.

Background and AimsPerianal fistulizing Crohns disease (PFCD) is a challenging complication with poorly understood pathogenesis and limited treatment options, largely due to the lack of clinically relevant animal models. Interferon-gamma (IFN-{gamma}) signaling is hyperactivated in human PFCD. We aimed to establish mouse models recapitulating human PFCD and to evaluate IFN-{gamma} as a new therapeutic target. MethodsPerianal fistulas were established in three mouse models with concurrent Crohns disease-like intestinal inflammation: wild-type (WT) mice with 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced proctocolitis, Il10-/-mice, and TNF{Delta}69AU/+ mice. A modified MAGNIFI-CD index was developed for longitudinal fistula assessment in mice. Transcriptomic analysis and flow cytometry were conducted on mouse fistula tissue. Re-analysis of single-cell and spatial transcriptomics of human PFCD tissues was performed. Therapeutic benefits of anti-TNF-, upadacitinib, and IFN-{gamma} pathway antagonists were evaluated in the PFCD models. ResultsAll three PFCD models sustained chronic perianal fistula tracts for at least 5 weeks after wire removal. All three models closely recapitulate the pathological and molecular features of PFCD in patients, as confirmed by clinical examination, MRI, histopathology, immunostaining, flow cytometry, and transcriptomics. IFN-{gamma} signaling emerged as a central and conserved pathway across all three mouse models and human PFCD. Targeting of the IFN-{gamma} pathway promptly improved fistula healing with mitigation of IFN-{gamma} signaling, inflammation, and epithelial-to-mesenchymal transition (EMT). Moreover, combining IFN-{gamma} and TNF- blockade demonstrated augmented therapeutic efficacy compared to anti-TNF- monotherapy. ConclusionsThese PFCD mouse models and imaging tools provide first reliable and clinically relevant platforms for mechanistic studies and therapeutic evaluation. IFN-{gamma} signaling represents a potential therapeutic target warranting clinical investigation.

The intestinal epithelium relies on rapid repair to maintain homeostasis after injury, and dysregulation of this process contributes to the pathogenesis of inflammatory bowel disease and colorectal cancer. Interleukin-11 (IL-11), a fibroblast-derived cytokine elevated in these diseases, has well-documented effects on stromal cells, but its direct action on intestinal epithelial cells remains poorly characterized. Here, we used mouse colon organoids as an isolated epithelial system to directly examine the effects of IL-11 on epithelial cells. IL-11 stimulation activated the canonical JAK/STAT3 pathway, as evidenced by increased STAT3 phosphorylation and Socs3 induction in a concentration-dependent manner. In a pipetting-based mechanical disruption model, IL-11 significantly increased the number of organoids recovered. Although mechanical disruption dominated the overall transcriptional landscape, RNA-seq analysis identified coordinated upregulation of STAT3 target genes and proliferation-related pathways specifically in response to IL-11. Pharmacological inhibition of STAT3 attenuated the IL-11-induced promotion of organoid recovery, indicating that STAT3 signaling mediates the epithelial response to IL-11 and maintains organoid size under basal conditions. Together, these findings demonstrate that IL-11 directly promotes intestinal epithelial repair after mechanical disruption through STAT3-dependent signaling, providing a mechanistic basis for its protective role in acute colonic injury.

Background Recent mouse model data demonstrate that chronic colonization with toxigenic Clostridioides difficile promotes colonic tumorigenesis via intraluminal toxin B (TcdB), its main virulence factor. In a prior multisite hospital cohort, we found that history of positive tcdB stool testing was associated with increased CRC risk in a dose-dependent manner, though limited by small sample size. We aimed to validate this association in a larger cohort with extended follow-up and greater geographic distribution using the Veterans Health Administration (VHA) Corporate Data Warehouse (CDW). Methods We conducted a retrospective cohort study among adults receiving care through the VA from 2000-2025 who underwent C. difficile testing. Data collected from the VHA CDW and National Death Index (NDI) included demographics, comorbidities, medications, CRC risk factors, and cancer incidence and death. The first C. difficile test date defined cohort entry; individuals with prior CRC were excluded. Ever C. difficile positivity was defined by a positive PCR or EIA results. The number of positive tests (episodes) was also determined to define recurrent positivity. Follow-up time ended at the first occurrence of CRC incidence or mortality, death from other causes, or censor date. Follow-up time was split for individuals who converted from negative to positive, with follow-up time updated accordingly. Multivariable Cox proportional hazards models were used to estimate hazard ratios (HRs) for C. difficile exposure and CRC incidence and mortality after adjustment for confounders. Tests for linear trend and tests for interaction were conducted to assess effect modification by sex and IBD status, while time-lag intervals were evaluated for 1, 3, 5, and 10 years before the outcome. Results Among 806,844 veterans with C. difficile testing, those with positive tests were more likely to be older, male, to have diabetes, to use aspirin, and to have a lower BMI than those with negative tests. Race and IBD prevalence were similar between the groups. There was no overall association between ever C. difficile positivity and CRC incidence (HR = 0.99, 95% CI 0.93-1.05). However, recurrent C. difficile positivity was associated with increased risk in a dose-response manner [2-3 episodes HR = 1.30 (95% CI 1.16-1.47), and >3 episodes HR = 1.58 (95% CI 1.17-2.14) compared to negative tests; ptrend< 0.001]. Further, ever C. difficile positivity was associated with increased CRC mortality risk (HR = 1.21, 95% CI 1.13-1.30; p < 0.001). Recurrent C. difficile positivity was associated with increased mortality risk but was particularly strong for those with >3 episodes among individuals with IBD (HR=3.84, 95% CI 1.98-7.45). In sensitivity analyses, the increased risk of CRC incidence and mortality attenuated beyond 10 years. Conclusion Prior positive C. difficile testing was associated with increased CRC incidence and mortality in a dose-dependent manner, particularly among patients with IBD. These findings extend animal model evidence, epidemiologically establishing C. difficile presence as an independent risk factor for subsequent colorectal tumorigenesis and supporting investigation into recurrent CDI, especially among patients with IBD, as a potential modifiable CRC risk factor.

Coeliac Disease (CeD) is a chronic gastrointestinal inflammatory disease initiated by dietary gluten in genetically predisposed individuals. While the inflammatory processes which drive tissue destruction in the coeliac duodenum have been extensively characterised, an increased oxidative stress (OS) response has also been suggested to contribute to CeD pathogenesis. However, the precise mechanisms which regulate OS in the coeliac mucosa and whether they impact inflammation remain ill defined. The master anti-oxidant transcriptional regulator Nuclear factor erythroid 2-related factor 2 (Nrf2), and its inhibitor, Kelch like ECH-associated protein 1 (Keap1) have been implicated in chronic gastrointestinal inflammatory diseases, such as ulcerative colitis but have been largely unexplored in the context of CeD. To investigate redox balance in the CeD duodenum, we utilised single cell transcriptomics to assess overall OS and cytoprotective Nrf2 activation across cell subsets in duodenal biopsies from CeD patients. OS induced gene expression was broadly increased across multiple cell subsets in the CeD mucosa. Simultaneously, specific markers of Nrf2 activation were decreased in cell subtypes central to pathogenesis of CeD, including activated CD4+ T cells and intraepithelial T lymphocytes, indicating a distinct redox imbalance in these cells. Furthermore, pharmacological activation of Nrf2 significantly decreased gliadin induced IFNG expression in CeD duodenal biopsies. Taken together, our findings demonstrate that redox imbalance represents a therapeutic opportunity for the modulation of proinflammatory responses that drive the pathogenesis of CeD.

Cell division cycle 42 (Cdc42) is a member of the Rho family of small GTPases, which plays crucial roles in regulating cytoskeletal remodeling, and membrane trafficking. While previous studies implicated Cdc42 in controlling intestinal epithelial homeostasis, the involvement of this small GTPase in the process of intestinal fibrogenesis remains unexplored. Our study was designed to determine whether Cdc42 regulates the fibrogenic activation of intestinal myofibroblasts in vitro. The study was conducted using a CCD-18Co normal human colonic fibroblast cell line, and primary human intestinal myofibroblasts (HIMF) isolated from Crohns disease (CD) patients. CCD-18Co and HIMF cells were stimulated by transforming growth factor-{beta}1 (TGF-{beta}1). Cdc42 was inhibited either genetically, using siRNA-mediated knockdown, or pharmacologically using specific Cdc42 inhibitors, ML141 and CASIN. Genetic and pharmacologic inhibition of Cdc42 markedly reduced TGF-{beta}1 induced expression of the major contractile cytoskeletal proteins, -smooth muscle actin, calponin 1 and L-caldesmon. Furthermore, Cdc42 inhibition significantly attenuated expression of key extracellular matrix (ECM) proteins, fibronectin and collagen I, in activated CCD-18Co cells and HIMF. Interestingly, decreased expression of contractile and ECM proteins in Cdc42-depleted myofibroblasts was not due to downregulation of the TGF-{beta}1 signaling, decreased mRNA transcription or increased lysosomal or proteasomal degradation of these proteins. Such suppressed pro-fibrotic activation of Cdc42-deficient CCD-18Co cells and HIMF involved a selective inhibition of protein translation due to inactivation of the AKT-mammalian target of rapamycin (mTOR) signaling module. These findings highlight Cdc42 as a key regulator of intestinal fibrosis that controls mTOR activation to enhance ECM production and contractile actomyosin cytoskeleton in intestinal myofibroblasts.

Background Unplanned hospital admissions in Inflammatory Bowel Diseases (IBD) account for nearly three-quarters of IBD inpatient stays in the United Kingdom. Although costly to services and distressing for patients, research exploring experiences and potential drivers of admissions is limited. We undertook a qualitative study to explore the healthcare experiences and access needs of people with IBD who had unplanned admissions, along with their caregivers and clinicians. Methods Semi-structured interviews with 25 participants from a single tertiary IBD service in England (17 people with IBD, 3 informal caregivers, 5 clinicians) were conducted. We applied thematic framework analysis, guided by the Candidacy Framework, and worked with 2 patient and public contributors to generate final themes. Results We identified four themes: 1) Difficulties in Identifying flares and asserting severity before admission, summarised the prevailing uncertainty in identifying a flare and access to timely IBD care. 2) Navigating a disjointed healthcare system, highlighted how lack of care plans and systemic barriers can delay access. 2) Emergency care access challenges highlighted the gaps in emergency and inpatient care during flares. Whilst 4) fighting for care and individual advocacy needs, described the persistent assertion for care that may disproportionally impact access to vulnerable groups, also highlighting the importance of positive interpersonal relationships. Conclusions Individual, interpersonal and healthcare factors across the patient pathway were perceived to shape access to care in unplanned IBD admissions. Potentially reducing admissions requires proactive strategies, including the integration of patient education, monitoring tools, establishment of specialist rapid-access pathways, and formal psychological support to address barriers to access.

Clostridioides difficile infection (CDI) susceptibility and severity are strongly associated with preexisting colonic inflammation. However, chronic inflammatory conditions such as cystic fibrosis rarely progress to symptomatic CDI despite high rates of C. difficile colonization, suggesting that inflammation alone is insufficient to explain disease vulnerability. Notably, populations relatively protected from symptomatic CDI exhibit impaired regenerative capacity within the colon epithelium. Here, we used single cell RNA sequencing of human colonoid monolayers to map markers of CDI susceptibility and severity to cell populations associated with inflammation and epithelial repair. We identified an inducible microfold-like (M-like) population that is largely absent from the healthy colon but emerges during inflammation and regeneration. These cells were enriched for markers of severe CDI, C. difficile toxin interaction genes, and elevated CCL20 and CFTR expression. Spatial imaging localized CCL20-producing cells to wound-like gaps in mock and CDI-treated colonoids, identifying a repair-associated niche active independent of infection. Following exposure to C. difficile, wound-healing transcription within the M-like lineage declined while tuft-like populations expanded and upregulated genes associated with immune cell recruitment. These findings demonstrate that epithelial regeneration shapes host CDI vulnerability. IMPORTANCEClostridioides difficile infection can lead to severe illness and death in vulnerable populations despite available treatments. Clinical signs of inflammation during active Clostridioides difficile infection are strongly associated with disease outcome, yet these responses primarily reflect tissue damage already underway, limiting opportunities to prevent progression. In contrast, conditions linked to severe disease, including inflammatory bowel disease and antibiotic exposure, are associated with colonic inflammation before infection or at the time of diagnosis, highlighting an opportunity for earlier identification of high-risk individuals. Using human colonoid single cell transcriptomics and spatial imaging, we identified a microfold-like cell population enriched for inflammatory mediators and Clostridioides difficile toxin interaction genes linked to severe disease. This population was active even in the absence of infection, suggesting that repair-associated populations within the inflamed colon may help identify susceptibility to severe CDI before clinical progression occurs.

Background: Heterogeneity in symptom presentation and treatment response in irritable bowel syndrome (IBS) remains poorly understood. The gut microbiota may contribute to this variability, but its role in shaping symptom trajectories and responses to self-management interventions is unclear. Objective: To identify symptom trajectory phenotypes and determine whether gut microbiota composition and function distinguish these phenotypes and predict multidimensional responses to pain self-management interventions in young adults with IBS. Design: Ancillary data analysis from a randomized control trial (NCT03332537). Methods: Participants with longitudinal data (n = 62) were analyzed using longitudinal k-means clustering (KML) based on trajectories of measures in IBS quality of life (QOL), Brief Pain Inventory (BPI), and psychoneurological outcomes (anxiety, applied cognition, depression, fatigue, global health, positive affect, and sleep disturbance) over 12 weeks. Baseline differences between clusters were assessed with Wilcoxon rank-sum tests, and longitudinal changes were evaluated with linear mixed models. Gut microbiota composition and predicted functional pathways were compared between phenotypes. Bayesian Additive Regression Trees (BART) models were used to identify baseline microbial taxa and pathways predictive of longitudinal changes in QOL, BPI pain interference, and severity. Results: Two distinct trajectory-defined response phenotypes were identified: a Constrained Response Phenotype (Phenotype A, n = 35) and an Adaptive Multidomain Response Phenotype (Phenotype B, n = 27). At baseline, Phenotype B showed lower pain severity and interference, but higher levels of anxiety, depression, and fatigue compared to Phenotype A. Over 12 weeks, both phenotypes showed improvements in pain outcomes (all p < 0.05), but only Phenotype B demonstrated broad improvements across psychoneurological domains and QOL (all p < 0.05). Phenotype A exhibited more limited improvements and worsening in several psychoneurological domains. Gut microbiota functional pathways differed between phenotypes, including pathways related to xenobiotic degradation, amino acid metabolism, bile secretion, and immune-related processes (all raw p < 0.05), although these did not remain significant after multiple testing correction. Machine learning models identified distinct, phenotype-specific microbial predictors of intervention response. In Phenotype A, genera such as Alistipes and Sutterella were consistently identified across models, whereas in Phenotype B, predictors included Phascolarctobacterium, Collinsella, and Parabacteroides. Functional pathways also differed between phenotypes, suggesting distinct microbiome-linked mechanisms underlying symptom trajectories and responses to pain interventions. Conclusions: Young adults with IBS exhibit distinct multidimensional response phenotypes that are associated with differential clinical and microbiome profiles. Baseline gut microbiota composition and functional capacity demonstrate phenotype-specific predictive signatures of treatment response, supporting a microbiome-informed framework for stratifying patients and advancing personalized self-management strategies in IBS.

Bidirectional communication between the nervous and immune systems has been demonstrated to limit or enhance immune cell function across organ systems and conditions. Although these neuroimmune circuits can become activated as an anti-inflammatory reflex, vagus nerve stimulation (VNS) reduces inflammation in models of endotoxemia, rheumatoid arthritis, and intestinal inflammation. In the spleen during endotoxemia, VNS activates a "cholinergic anti-inflammatory pathway" (CAIP), whereby choline acetyltransferase (ChAT)-expressing T cells release acetylcholine to reduce macrophage activation. VNS can also drive CAIP-independent pathways to reduce inflammation in the spleen and intestinal tract, although the circuitry modulating colonic inflammation remains underexplored. Here, we demonstrate that left cervical VNS reduces acute LPS-induced inflammation, evidenced by reduced Tnfa expression in colon and spleen and decreased circulating TNF. In the colon, these protective effects required efferent but not afferent VNS and were independent of ChAT+ T cells, IL10, {beta}-adrenergic signaling, and colonic sympathetic innervation. Critically, the ability of VNS to modulate colonic inflammation depended on prior tissue-specific inflammation. Mice recovering from DSS colitis, despite near-complete histological recovery, were refractory to the protective effects of VNS in the colon. This lack of efficacy in the colon was not reflected in measures of inflammation in the spleen or serum, highlighting the need for target-organ-specific monitoring. This loss of efficacy after colonic inflammation was transient, with restoration occurring upon complete recovery. These findings demonstrate that VNS efficacy in colonic inflammation depends on circuitry distinct from canonical systemic anti-inflammatory pathways, and that tissue responsiveness is shaped by anatomical site and inflammatory history. Key PointsO_LIElectrical stimulation of the left cervical vagus nerve reduces LPS-induced inflammation in the mouse colon. C_LIO_LIThis colonic anti-inflammatory effect requires vagal efferents but not afferent signaling. C_LIO_LIUnlike canonical splenic anti-inflammatory pathways, the colonic response does not require ChAT+ T cells, IL-10, {beta}-adrenergic signaling, or local sympathetic innervation. C_LIO_LIRecent DSS colitis abolishes colonic responsiveness to VNS despite preserved splenic and systemic anti-inflammatory effects. C_LIO_LIRecovery of VNS anti-inflammatory efficacy after colitis shows that neuroimmune responsiveness in the colon is dynamically shaped by inflammatory history. C_LI

BackgroundEmerging evidence suggests that the oral microbiome may contribute to aberrant gut immune responses in Inflammatory Bowel Disease (IBD). MethodsWe performed a comprehensive, harmonised analysis of aggregated oral microbiome 16S rRNA datasets across multiple cohorts. Data were processed using a unified bioinformatics pipeline including DADA2 for taxonomic assignment, PICRUSt2 for functional prediction, MaAsLin2 for multivariable modelling, and machine learning. ResultsAcross 25 studies (n = 1,136 IBD; n = 759 controls), meta-analysis showed significantly reduced oral microbial Shannon diversity in IBD (standardised mean difference -0.31, p = 0.007). Secondary bioinformatics analysis of six datasets plus in-house data confirmed this reduction (Shannon diversity; Hedges SMD = -0.372, p < 0.001), driven primarily by Crohns disease (CD). Beta diversity demonstrated global compositional shifts, with CD demonstrating greater divergence from controls than ulcerative colitis (UC). Multivariable modelling identified reproducible taxa enriched in IBD, including Corynebacterium, Serratia and Streptoccocus, while Porphyromonas and Ruminococcaceae.G1 were enriched in controls. Functional pathway prediction indicated reduced butyrate metabolism in IBD sub-types and increased aromatic amino acid and related metabolite degradation pathways. Machine learning classifiers achieved modest discrimination (mean AUC [~]0.67), supporting the potential of saliva-based microbiome profiling to study dysbiosis in IBD. ConclusionsThese findings demonstrate that the oral microbiome in IBD is characterised by reduced diversity and reproducible structural community reorganisation. Together, these data support a contributory role for the oral-gut axis in CD pathogenesis and provide a rationale for targeted mechanistic and longitudinal studies to define causal links between oral dysbiosis and intestinal inflammation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/26351936v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@57306corg.highwire.dtl.DTLVardef@2c0ef0org.highwire.dtl.DTLVardef@88b0b3org.highwire.dtl.DTLVardef@8ed62_HPS_FORMAT_FIGEXP M_FIG C_FIG

Antibiotic exposure is a significant risk factor for Crohns disease, yet the tissue-specific consequences of antibiotic-driven dysbiosis remain poorly defined. Adherent-invasive Escherichia coli (AIEC), a pathobiont enriched in Crohns disease, expands following antibiotic treatment, but whether discrete mucosal niches support this expansion is unknown. Peyers patches are specialized lymphoid structures that coordinate mucosal immunity and are frequently associated with early disease lesions, suggesting they may represent a vulnerable site for pathobiont colonization. Here, we show that vancomycin disrupts the Peyers patch-associated microbiome, creating a permissive niche that is selectively exploited by AIEC and associated with focal inflammation. Antibiotic treatment markedly increased AIEC burden within Peyers patches. AIEC localized within the lymphoid follicle was accompanied by focal tissue pathology and a distinct cytokine signature. In contrast, expansion of resident E. coli in the absence of AIEC did not elicit comparable inflammation, indicating that the pathogenic traits of AIEC are required to trigger disease-relevant responses in this niche. Supporting this, genetic disruption of flagellin, long polar fimbriae, or antimicrobial peptide resistance in AIEC attenuated Peyers patch colonization or inflammation, revealing separable mechanisms governing niche access and immunopathology. Together, these findings identify Peyers patches as a previously unrecognized reservoir for antibiotic-driven AIEC expansion and define a localized host-microbe interaction that links dysbiosis to focal intestinal inflammation. These results provide a mechanistic framework for understanding how antibiotic exposure may precipitate site-specific pathology in Crohns disease. Further, these findings highlight that mucosal lymphoid tissues should be considered when evaluating microbiome-targeted therapeutic interventions in Crohns disease.

The pathogenic bacterium Clostridioides difficile is a major cause of antibiotic-associated diarrheal disease. Treatment of the disease is challenging because antibiotics used for treatment may also perpetuate the conditions that contributed to initial susceptibility. Elucidating the mechanisms of C. difficile/intestinal epithelium interaction is needed to facilitate the development of new therapeutic options. The studies described in this communication demonstrate the development of a tissue culture system that supported the growth of C. difficile in co-culture with a model of the human intestinal epithelium produced from colonoids, organoids derived from human colonic biopsies. Epithelial cell responses to C. difficile included upregulation of CCL20, encoding a chemokine. Glucosylating toxin production by the bacteria was required for upregulation of CCL20. Additionally, bacteria associated with the monolayer in a non-toxin dependent manner. This system will support future investigation of epithelium/C. difficile interactions during CDI and identification of mechanisms that drive pathogenesis by C. difficile in the human intestine.