The naked mole-rat (NMR; Heterocephalus glaber) is a subterranean rodent native to the arid regions of the Horn of Africa. The NMR is the longest-lived rodent and is known for its distinctive physiological and social traits. This species has become a notable model organism for studying aging, cancer biology, behavioral ecology, and reproduction. Recently, NMRs have gained attention because their gastrointestinal tract features an exceptionally strong intestinal barrier, a large number of goblet cells, a thicker mucin layer, and reduced gut permeability. The NMR gut microbiome, similar to that observed in human centenarians, is highly diverse and characterized by a high microbial load. In fact, Hart et al (2026) demonstrated that spontaneous infection with Citrobacter braakii in the NMR causes clinical symptoms and histopathological changes that are very similar to those observed in human colitis. If left untreated, the disease can progress and become fatal. However, probiotic treatment can reverse the clinical and histopathological phenotypes. These findings indicate that, in addition to serving as a powerful model for aging, cancer, and reproduction, the NMR may also serve as a powerful tool for studying human diseases such as gut dysbiosis, gut barrier dysfunction, and colitis. © 2026 The Pathological Society of Great Britain and Ireland.

Inflammatory bowel disease (IBD), which includes Crohn's disease and ulcerative colitis (UC), comprises chronic relapsing inflammatory disorders of the gastrointestinal tract. Although the acute dextran sodium sulfate (DSS)-induced murine colitis model does not reproduce chronic human IBD, it reflects acute colonic epithelial injury and superficial mucosal inflammation, which are more characteristic of UC than of Crohn's disease. Therefore, the present study investigated whether the root extract of Acanthopanax senticosus Harms (ASH) exerts a prophylactic effect in an acute DSS-induced experimental colitis model in mice. Mice were fed a chow diet containing 5% ASH for 28 days, followed by water containing 3% DSS for 7 days to induce acute colitis. After DSS administration, mice fed the ASH diet exhibited significant inhibition of colon shortening and a reduction in clinical disease severity compared with the control group. Furthermore, the elevated levels of pro-inflammatory cytokines and oxidative stress markers in the serum were mitigated. Mechanistically, ASH pretreatment was associated with reduced NADPH oxidase 2 expression, relative restoration of nuclear factor erythroid 2-related factor 2 expression and increased Cu/Zn-superoxide dismutase expression in the colon. These findings suggested that ASH pretreatment attenuated acute DSS-induced experimental colitis by alleviating mucosal inflammation and restoring redox homeostasis.

Multiple members (DUSP1-29) of dual-specificity phosphatase (DSP) family are key regulators of mitogen-activated protein kinases (MAPKs), which regulate numerous physiological responses. Eight DUSPs are also named MAPK phosphatases (MKPs). DUSP dysregulation contributes to the pathogenesis of various human inflammatory and chronic diseases. Downregulation of DUSP1, DUSP3, DUSP11, and DUSP22, as well as upregulation of DUSP4, DUSP6, and DUSP23 are involved in human autoimmune diseases. Besides autoimmune diseases, reduction of DUSP1, DUSP2, and DUSP14, as well as induction of DUSP8 contribute to the pathogenesis of allergic diseases. Additionally, decreased levels of DUSP2, DUSP11, DUSP22, and DUSP28 are associated with human inflammatory bowel diseases. Moreover, deficiency of 10 DUSPs, as well as induction of DUSP4 are associated with metabolic diseases. Downregulation of 5 DUSPs are involved in cardiovascular disease pathogenesis; in contrast, upregulation of other 5 DUSPs are correlated with cardiovascular diseases. Collectively, dysregulated DUSPs could be diagnostic biomarkers and therapeutic targets for inflammatory diseases. Due to complex expression patterns of DUSPs, it is crucial to study the regulatory mechanisms of individual DUSPs in various inflammatory and chronic diseases. In this review, we summarize the roles and regulatory mechanisms of DUSPs in human inflammatory and chronic diseases. We also discuss the potential therapeutic applications of DUSP agonists/inhibitors in human inflammatory and chronic diseases.

Spatiotemporal analysis reveals distinct inflammatory programs underlying chronic colitis.

Shiga toxin-producing Escherichia coli (STEC) is a pathotype of E. coli associated with a wide variety of diarrhoea in neonatal calves, causing a global economic loss in the dairy industry with significant zoonotic risks via STEC and intimin-producing STEC, resulting in enteric and systemic illness, including diarrhoea, haemorrhagic colitis (HC), and haemolytic uremic syndrome (HUS) in humans. The presence of multidrug-resistant (MDR) STEC in neonatal diarrhoeic calves represents a significant public health concern and limits available therapeutic options. This study investigated the presence of extended-spectrum β-lactamase (ESBL) and carbapenemase-producing STEC isolates and their genotypic combinations of virulence and resistance genes in diarrhoeal calves. A total of 75 rectal swabs from diarrhoeic calves aged ≤ 12 weeks were sampled from three districts of Bangladesh, screened using selective culture and polymerase chain reaction (PCR), followed by phenotypic antimicrobial susceptibility profiling and phenotypic and genotypic screening for ESBL- and carbapenemase-production. The E. coli isolation rate among diarrhoeic calves was 80% (60/75; 95% CI: 69.17-88.35). PCR screening revealed the presence of stx1, stx2, and eae virulence genes, and the prevalence of E. coli isolates harbouring these virulence genes was 13.33% (10/75; 95% CI: 6.5-23.1). The ten virulent isolates included five stx-positive, four stx/eae-positive and one eae-positive E. coli. All the virulent isolates (100%) were resistant to ciprofloxacin and meropenem; however, eight (80%) of the tested isolates were susceptible to gentamycin, and 60% were susceptible to amoxicillin-clavulanate. All virulent E. coli isolates were MDR. We observed that 5 (50%) of the virulent E. coli isolates were ESBL producers, and 7 (70%) were carbapenemase producers. The results for phenotypes and genotypes of ESBL- and carbapenemase-producing strains were not concordant. At least one ESBL gene was present in nine of the ten virulent isolates examined, including four ESBL phenotype-negative isolates. Carbapenemase-producing genotypes were significantly more common in isolates with multiple virulence genes than in those with a single virulence gene (p = 0.001). To the best of our knowledge, this is the first report on ESBL- and carbapenemase-producing MDR STEC in diarrhoeic calves in Bangladesh.

Th17 cell function is highly context-dependent and can be categorized into pathogenic and nonpathogenic Th17 cell subsets. Understanding the molecule control of pathogenic Th17 (pTh17) cell immunity will benefit the treatment for related autoimmune diseases. Here, we revealed that cytochrome P450 1B1 (CYP1B1) is highly upregulated during mice and human colitis. CYP1B1 promoted both colon inflammatory diseases and colitis-associated colorectal cancer via pTh17-dependent but microbiota-independent manner. Notably, CYP1B1 specifically dictated the differentiation and pathogenicity of pTh17 cells, while having no effects on nonpathogenic Th17 cell generation. Mechanistically, CYP1B1 deficiency disrupted intracellular redox homeostasis via decreased glutathione synthetase, leading to increased ROS and mitochondrial dysfunction of pTh17 cells. ROS elimination by N-acetylcysteine or ectopic glutathione synthetase expression restored mitochondrial fitness and promoted pTh17 cell survival and generation. Taken together, our findings uncover a T cell intrinsic CYP1B1-ROS-mitochondrial axis in driving pTh17 cell generation, interfering with this hub may be beneficial for pTh17 cell-related immunopathology.

Dysregulation of the immune system, gut microbiota alteration, and epithelial dynamics in the colon contribute to the pathogenesis of inflammatory bowel disease (IBD). However, the role of epithelial dynamics, particularly epithelial regeneration, remains incompletely understood. CADM1 encodes an immunoglobulin-superfamily cell adhesion molecule involved in epithelial adhesion, immune cell interactions, and tumor suppression in colon and various cancers. Here, we investigated the role of CADM1 in IBD using a murine model of colitis induced by dextran sulfate sodium in both wild-type and conventional Cadm1-deficient (Cadm1−/−) mice. Cadm1−/− mice exhibited more severe colitis than wild-type mice with increased mortality (64% vs. 10%) and delayed recovery. Cadm1−/− mice showed reduced numbers of Ki-67-positive cells in colonic crypts and delayed epithelial regeneration, whereas no significant differences were observed in epithelial apoptosis, intestinal permeability, or immune responses. Immunohistochemistry revealed that CADM1 expression was restricted to regenerative crypt cells in wild-type mice with nuclear accumulation of β-catenin and phospho-Akt. Furthermore, CADM1 overexpression in colon epithelial cells enhanced Tcf-transcriptional activity in a β-catenin-dependent manner. Immunohistochemistry of human IBD materials revealed that CADM1 expression also correlated with nuclear β-catenin accumulation in crypt epithelial cells. Collectively, CADM1 appears to promote colonic epithelial regeneration through the PI3K/Akt/β-catenin axis to protect against severe epithelial injury in IBD.

Group 3 innate lymphoid cells (ILC3s) preserve intestinal barrier integrity by producing IL-22 and IL-17A, yet the molecular mechanisms that maintain these cytokines during inflammation are incompletely defined. Here, we identify DAB2IP as a cell-intrinsic regulator of ILC3 effector function. In human inflammatory bowel disease mucosa, DAB2IP expression is reduced and associated with transcriptional programs linked to impaired epithelial repair. In murine models, inflammatory cues dynamically modulate Dab2ip in ILC3s, and genetic loss of DAB2IP diminishes IL-22 and IL-17A, compromising host defense during Citrobacter rodentium infection, and exacerbates dextran sulfate sodium-induced colitis. Mechanistically, DAB2IP enables efficient NF-κB activation, promoting IκBα degradation, p65 nuclear accumulation, and thus transcription of Il22/Il17a and NF-κB targets. These results reveal a context-dependent role for DAB2IP as a positive regulator of NF-κB in ILC3s, highlighting its previously unknown function in mucosal immunity and epithelial repair, and suggesting that restoring DAB2IP signaling could enhance barrier protection during intestinal inflammation.

CD4⁺ helper T (TH) cells consist of multiple functional subsets defined by specific effector cytokines and transcription factors. Recently, single-cell transcriptomic analyses have revealed possible existence of additional populations. Here we identify a unique CD4⁺ T cell subset in mouse and human colitis characterized by high levels of granzyme K (Gzmk) expression, designated as THK cells. These cells exhibit unique transcriptional signatures, with minimal expression of classical TH-defining factors but rather prominent Eomesodermin (Eomes) expression. Notably, THK cell differentiation is independent of TH1, TH2 and TH17 lineages in colitis. EOMES is both necessary and sufficient for THK cell induction, by directly driving the expression of Gzmk and associated effector molecules. Genetic ablation of Eomes ameliorates intestinal immunopathology in a T cell-induced colitis model. The THK transcriptional program seems to be conserved across species and in diverse disease contexts. Our findings establish THK cells as a distinct TH cell subtype, and the EOMES-THK axis may serve as a potential therapeutic target in inflammatory diseases.

Ethyl acetate extract of Lactococcus lactis KR-050L suppresses IL-6/STAT3 signaling in Hep3B cells and alleviates DSS-induced colitis in mice.

Microbiome alterations and host-pathogen interactions in paratuberculosis: A one health perspective.

Chronic inflammation of the colon has been described as an inflammatory condition of the colon that is characterized by recurring flare-ups of the disease (ulcerative colitis). Ulcerative colitis is a complex inflammatory disorder of the colon that is determined by the combination of several mechanisms: abnormal bacterial flora, inappropriate immune response, and the breakdown of the protective lining of the colon. The last 10years have seen the emergence of plant derived polysaccharides as new biological active compounds with the potential to intervene at the different levels of the disease. Plant derived polysaccharides can be divided into two separate regulatory modes; the first mode of action occurs when the polysaccharides work in collaboration with the microbiota to modify the structure of the microbiota and create a positive environment for the growth of the beneficial bacteria. The second mode of action occurs without the use of the microbiota and works by creating anti-inflammatory signals to regulate the body's immune system and protect the integrity of the intestinal lining. The authors of this paper used a systematic method to identify 103 papers published on the subject of plant derived polysaccharides and their effects on the disease mechanism of ulcerative colitis. The results of the analysis of these papers indicated that the polysaccharides act to reduce the inflammatory process through the downregulation of the three major inflammatory signaling pathways of NF-κB, PI3K/AKT, and JAK/STAT. In addition, the polysaccharides were shown to restore the balance of macrophages and T-cells and increase the production of mucus producing cells and tighten the tight junctions between the intestinal epithelial cells. In recent studies it was found that the metabolic products of the microbes and the intracellular signaling processes are part of a coordinated regulatory network. Therefore, based on the results of the study, plant derived polysaccharides appear to be multi-target modulators of the disease process rather than just acting on one pathway of the disease process. Although the study indicates that there has been significant progress in understanding how the plant derived polysaccharides interact with the disease process, there are still many barriers to overcome before the plant derived polysaccharides can be used as a treatment for ulcerative colitis in humans. These barriers include; the structural heterogeneity of the polysaccharides, the lack of clinical trials evaluating the safety and efficacy of the polysaccharides as a treatment for ulcerative colitis, and the large variation in how individuals respond to changes in the composition of the gastrointestinal tract microbiota. Therefore, this study represents a comprehensive review of the current state of knowledge on the interaction of plant derived polysaccharides with the pathophysiology of ulcerative colitis, and highlights areas of future research necessary to develop polysaccharide based treatments for ulcerative colitis that are safe, effective, and can be translated into the clinic.

ABSTRACT Adipose tissue dysfunction is integral to the pathophysiology of ulcerative colitis (UC), yet the conservation of adipose immunometabolic responses across species remains unclear. Here, we employed a comparative transcriptomic approach to analyze adipose remodeling in dextran sulfate sodium (DSS)‐induced porcine and murine colitis models alongside human UC datasets. We report that intestinal inflammation induced widespread adipocyte atrophy and triggered a convergent inflammatory response across physiologically distinct visceral and subcutaneous depots. Mechanistically, this remodeling was defined by a systemic suppression of fatty acid synthesis pathways. Importantly, the expression levels of key lipogenic enzymes were negatively correlated with the severity of colonic inflammation, indicating that intestinal injury directly dictates the magnitude of lipogenesis inhibition. Cross‐species alignment revealed a critical distinction: while murine visceral fat exhibited fatty acid metabolism activation, the porcine response mirrored the fatty acid metabolism downregulation observed in human patients. These results identify a fundamental species‐specific difference and establish the porcine model as a translational tool, which faithfully replicates the atrophy and fatty acid metabolism suppression characteristic of human inflammatory bowel disease.

ABSTRACTType I interferons (IFN-Is) play a critical role in innate immunity, modulating the host response. While dysregulated IFN-I signaling has been implicated in autoimmune and infectious disorders, its role in inflammatory bowel disease (IBD) remains unclear. In this study, we extensively assessed the function of IFN-I signaling in human IBD and murine models of colitis. Expression of IFN-I signature genes was elevated in patients with active ulcerative colitis as well as multiple murine models of colitis. Single cell RNA sequencing revealed that upregulated IFN-I signature genes were enriched in myeloid cells, which exhibited increased expression of IFN receptors during mucosal inflammation. Mice carrying gain-of-function alleles ofIfnar1, a subunit of IFN-I receptor, showed heightened IFN-I signaling and altered colonic immune homeostasis at baseline, and were more susceptible to experimental colitis. In contrast, postnatal inhibition of IFNAR1, using either an inducible transgenic mouse model or an anti-IFNAR1 blocking antibody, protected against experimental colitis. Taken together, our findings reveal a previously under-recognized pathogenic role of IFN-I in IBD and provide a rationale for therapeutic intervention targeting this pathway.

Inflammatory bowel disease (IBD), comprising Crohn's disease and ulcerative colitis, is a chronic relapsing inflammatory disorder with steadily increasing global prevalence, significantly impairing patient quality of life. Preclinical research in IBD has traditionally relied on animal models to investigate disease etiology, immunopathogenesis, and therapeutic responses. A wide range of experimental models primarily rodents, along with limited use of larger animals such as pigs and nonhuman primates, have been developed to reproduce key clinical, histological, and immunological features of human IBD. These models have played a crucial role in evaluating pharmacological agents, biologics, probiotics, dietary interventions, gene-based therapies, and microbiome-targeted strategies. Notably, accumulating evidence highlights the pivotal role of gut microbiota dysbiosis in disease initiation, progression, and therapeutic responsiveness, making host-microbiome interactions a central component of contemporary IBD research. However, despite their utility, animal models exhibit important limitations related to interspecies differences, incomplete microbiome representation, and poor translational predictability. In response, increasing regulatory pressure from agencies such as the National Institutes of Health (NIH) and FDA to reduce animal experimentation has accelerated the development of human-relevant, nonanimal platforms, including intestinal organoids, in vitro coculture systems, and gut-on-chip technologies. This review critically evaluates existing in vivo IBD models with particular emphasis on their ability to capture immune-microbiome epithelial interactions while also discussing emerging human-derived systems as complementary translational tools. Collectively, the integration of microbiome-responsive and immune-competent-advanced in vitro models represents a promising direction to bridge the gap between experimental findings and clinical application in IBD research.

Enteropathogenic bacteria are a major cause of morbidity and mortality globally. While mouse models have been indispensable in advancing our understanding of infectious enteric diseases, key differences in intestinal microbiota and immunobiology between mice and humans underscore the need for alternative mammalian models that better recapitulate human disease states. The naked mole rat (NMR), the longest-lived rodent and a model of healthy ageing, presents a unique opportunity. It possesses an exceptionally robust intestinal barrier, an abundance of goblet cells, a thicker mucin layer, and reduced gut permeability compared to mice. Additionally, the NMR gut microbiome exhibits compositional and functional features shared with human centenarians and traditional-lifestyle populations (e.g. Hadza hunter-gatherers), including an enrichment of health-associated taxa and metabolic pathways. Here, we leverage this model to show that systemic Citrobacter braakii infection is associated with colonic inflammation and epithelial injury that closely mimics human haemorrhagic colitis. Infected NMRs develop mucosal erosions, ulcerations, depletion of goblet cells, expansion of proliferative compartments, and active inflammation in the lamina propria. Without intervention, systemic inflammation associated with sepsis ensues and results in high mortality. Furthermore, we demonstrate the utility of this model for therapeutic testing by showing a strong effect of a probiotic cocktail comprising lactobacilli, bifidobacteria, streptococci, and enterococci. Treatment with this cocktail promoted mucosal healing, restored intestinal homeostasis, and exerted an anti-inflammatory effect. Taken together, we establish the NMR as a translatable model for investigating disease mechanisms in infectious colitis, including disruptions in mucosal barrier permeability, gut microbial ecology, and local and systemic immune regulation, as well as for testing functional probiotic strains as potential therapeutics. © 2026 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Paneth cells are defensive cells in the intestinal tract, which secrete niche factors and antimicrobial peptides (AMPs) to maintainthesmall intestinal stem cell niche and immune homeostasis. Here, we show that Vestigial-like family member 4 (VGLL4) plays a pivotal role in maintaining small intestinal homeostasis and in regulating Paneth cells. VGLL4 expression is downregulated in response to irradiation and DSS-induced colitis. Consistently, public datasets of human colitis show reduced VGLL4 expression. Loss of VGLL4 in the intestinal epithelium decreases Paneth cell numbers and AMPs production, and triggers gut microbiota dysbiosis, impairing intestinal regenerative capacity. Mechanistically, VGLL4 forms a complex with TEAD4 and ATOH1, stimulating GFI1 expression and promoting Paneth cell differentiation. Furthermore, VGLL4 forms a complex with TEAD4 and TCF4 to induce defensin expression, thereby maintaining microbiota composition. Collectively, our findings uncover novel roles for VGLL4 in intestinal homeostasis.

Abstract Background Intestinal microbiota, diet, and the immune system contribute to the development of inflammatory bowel diseases (IBD). The aryl hydrocarbon receptor (AhR) is a critical regulator of intestinal immunity and barrier function, activated by microbial and host tryptophan (Trp) metabolites. IBD patients in large cohorts and in our own studies show reduced bacterial Trp metabolism genes and metabolites, which is associated with downregulation of colonic AhR activation, but the contribution of microbiota to this phenotype is unclear. Enhancing Trp metabolism through diet/probiotics may offer a therapeutic strategy to reduce inflammation in IBD. Aims To investigate the impact of diet-microbe interventions on AhR activation in mice colonized with IBD and mouse microbiota with impaired Trp metabolism. Methods Germ-free C57BL/6 mice were colonized with microbiota from human healthy controls (HC) or IBD patients, and specific pathogen free (SPF) or a minimal microbiota composed of 8 species (MM). Mice were provided a high Trp (HT; 1% Trp) or control diet (CD; 0.14% Trp). Subsets of mice received probiotic Clostridium sporogenes with Trp metabolism genes. Colitis was induced in mice using dextran sulfate sodium (DSS; 2.5% w/v), 2,4,6-trinitrobenzenesulfonic acid (TNBS; 2% w/v), and IL-10-/- models. AhR antagonist (CH223191) or vehicle were provided daily during colitis. Trp metabolites quantified by LC-MS. AhR activation was quantified by AhR reporter and RT-qPCR. Histologic analysis of distal colon, clinical scores, intestinal permeability (Ussing chambers), and inflammatory genes (Nanostring) were assessed. Results Humanized mice colonized with IBD microbiota had reduced Trp metabolism vs HC. MM microbiota mice had reduced Trp metabolism vs SPF. Reduced Trp metabolism associated with exacerbated colitis severity. HT diet supplementation in IBD humanized mice increased Trp metabolism, activated AhR, and reduced DSS, TNBS, and IL-10-/- colitis. Probiotic supplementation was required with HT diet to achieve clinical colitis alleviation in mice with severe impairment of AhR signalling. Conclusions Transfer of AhR activation from humans to mice by microbiota transplant indicates that AhR activation is microbiota dependent. Impaired host/microbial Trp metabolism increases susceptibility to colitis. Trp supplementation and microbial interventions offer an approach to restore AhR signalling and reduce intestinal inflammation. Funding Agencies CAG, CCC, CIHR

The inflammatory process is a conserved and adaptive biological response to infection or tissue damage. Despite its substantial energy demands, inflammation triggers centrally regulated changes in behavior, commonly referred to as sickness behavior, which includes anorexia and consequent negative energy balance. Although these responses have been extensively modeled through infection or cytokine administration, they remain less explored in a more dynamic spectrum of clinical conditions, such as inflammatory bowel disease (IBD). In this study, we used the dextran sodium sulfate (DSS) model of colitis, which mimics key features of human IBD. We assessed food and water intake, locomotor activity, and body composition over the disease progression. We further assessed neuronal activation and transcriptional changes in metabolic-sensing brain regions at key disease stages. Acute DSS-induced disease progression was associated with metabolic alterations, including anorexia, energy conservation, reduced physical activity, and changes in body mass composition. A positive correlation between disease severity and neuronal activation in the hypothalamus and the caudal brainstem was also found. Transcriptomic analysis revealed changes in hypothalamic gene expression associated with the immune response. Furthermore, targeted colocalization studies identified the activation of hypothalamic hunger-promoting AgRP/NPY-expressing neurons as a neuronal population recruited during colitis, suggesting a role for these neurons in coordinating allostatic metabolic adaptations to intestinal inflammation. This study provides evidence that the DSS model is a clinically relevant, dynamic, and tractable tool for studying the progression of sickness-like behavior in IBD, as well as the underlying neurometabolic adaptations that extend beyond the gut.NEW & NOTEWORTHY By showing that experimental colitis induced by DSS in mice triggers metabolic adaptations and activation of brain regions regulating energy balance, this study expands the model's relevance beyond intestinal inflammation. These findings provide a framework to investigate gut-brain interactions and the neurometabolic components of sickness-like behavior in inflammatory bowel disease.

Abstract Background Inflammatory bowel disease (IBD) involves dysregulated inflammation shaped by environmental factors, including diet and the gut microbiota. IBD prevalence is rising in Western countries with diets high in animal protein, fat, and sugar. Diet-microbiota interactions are thought to play a significant role in intestinal inflammation, and microbial composition and function are known to be altered in IBD. Our lab recently showed that high protein diets (HPD) worsen colitis by activating the mechanistic target of rapamycin (mTOR) and suppressing autophagy. Branched-chain amino acids (BCAAs) and arginine, abundant in HPD, are potent mTOR activators. Therefore, we hypothesize that microbial protein metabolism is altered after inflammation and produces bioactive compounds that enhance mTOR activation. Aims Determine how gut microbiota and intestinal inflammation influence dietary protein metabolism and mTOR activation. Methods Specific-pathogen-free (SPF), altered Schaedler flora (ASF; stable murine microbiota of 8 bacterial strains), and germ-free (GF) C57BL/6 mice received a control diet (CD;14% protein; n = 5/group) or HPD (40%; n = 6/group) for two weeks to study the importance of the microbiota on protein metabolism and mTOR activation. To study the role of inflammation on microbial protein metabolism, colitis was induced in SPF C57BL/6 mice with 3% dextran sulfate sodium (DSS) for 5 days, followed by 2 days of water. mTOR activation was assessed by p-S6 expression, a downstream target of mTOR, using immunohistochemistry in colon and small intestine tissue. Fecal samples were analyzed using LC-MS metabolomics to quantify total amino acids, BCAAs, and related metabolites. Results Gut microbiota composition strongly shaped protein metabolism. On the CD, GF, ASF, and SPF mice had distinct metabolite profiles; SPF mice had higher amino acid-derived metabolites (spermine, spermidine, para-cresol). After DSS, SPF mice had increased BCAAs (valine, isoleucine) and arginine versus baseline, and higher putrescine, spermine, spermidine, and cadaverine. GF mice displayed greater colonic p-S6 expression than ASF and SPF mice on CD. Under HPD conditions, GF mice showed increased small-intestinal mTOR activation relative to ASF and SPF mice. Inflammation increased colonic mTOR activation. Conclusions The gut microbiota regulates dietary protein metabolism and modulates mTOR activity, as microbiota-defined differences influence inflammatory signalling. Intestinal inflammation elevates colonic amino acids with mTOR-activating potential. Future work will extend HPD-focused metabolomics in mouse models and human IBD cohorts. Funding Agencies CCC, CIHR