Dietary Protein Source Shapes Gut Microbial Structure and Predicted Functional Potential: a Systematic Integrative Re-Analysis Using Machine Learning

Advances in Nutrition · 2026-01-02 · 1 citations

Dietary proteins shape gut microbial ecology, yet the taxonomic and functional consequences of plant- compared with animal-based proteins remain poorly defined. Although digestibility and fermentation profiles differ by protein type, a systematic evaluation of how these differences influence microbial diversity, community structure, and metabolic capacity is lacking. This study represents a systematic integrative re-analysis of raw 16S rRNA sequencing datasets derived from independent controlled animal feeding studies. Following PRISMA guidelines, we analyzed 16S rRNA sequencing data from 10 murine studies (n = 187) comparing plant- and animal-protein diets. Alpha diversity was assessed using Shannon, Inverse Simpson, and Chao1 indices, and beta diversity with Aitchison distances. Differentially abundant taxa were identified using linear discriminant analysis, effect size, and class-weighted Random Forest (RF) models. Functional potential was inferred with phylogenetic investigation of communities by reconstruction of unobserved states, and taxon-pathway relationships were explored using correlation and network analyses. Plant-protein diets increased gut microbial diversity across all alpha diversity metrics and were associated with higher representation of saccharolytic and nitrogen-recycling genera such as Bacteroides, Muribaculaceae, and Allobaculum. Animal-protein diets favored proteolytic taxa, including Clostridium sensu stricto 1 and Colidextribacter. Microbial community structure differed significantly between diets (analysis of similarities R = 0.663, P < 0.001). RF models achieved >88% accuracy (area under the curve = 0.995) in predicting dietary groups, and linear discriminant analysis effect size identified consistent discriminating taxa. Functional profiling showed that plant-based diets enriched pathways linked to short-chain fatty acid and aromatic amino acid metabolism, whereas animal-based diets favored sulfur- and branched-chain amino acid-associated pathways. Network analysis identified Muribaculaceae as a plant-associated hub and Lactobacillus as an animal-associated hub. Dietary protein source significantly influences gut microbiota composition and functional potential in mice. Plant- and animal-based proteins generate distinct metabolic signatures with implications for nitrogen cycling, sulfur metabolism, and microbial ecology. Future controlled dietary studies that harmonize protein source with other macronutrient variables are needed to isolate protein-specific effects.

Social stress worsens colitis through β-adrenergic–driven oxidative stress in intestinal mucosal compartments

bioRxiv (Cold Spring Harbor Laboratory) · 2025-07-18 · 1 citations

Abstract Psychological stress is a known risk factor for inflammatory bowel disease (IBD), but the mechanisms linking stress to worsened disease remain unclear. Because distinct stress paradigms activate different neuroimmune circuits, it is critical to investigate model-specific effects. We examined how social stress primes the gut for heightened inflammation and whether this is mediated by specific neuroendocrine pathways, including α2-/β-adrenergic (sympathetic) or glucocorticoid/ corticotropin-releasing hormone receptor (CRHR1) (HPA axis) signaling. Mice were exposed to social disruption (SDR) stress and pre- treated with pharmacological antagonists targeting α2-adrenergic receptors (idazoxan), β-adrenergic receptor (β-AR) (propranolol), glucocorticoid receptor (mifepristone), or CRHR1 (antalarmin). Intestinal epithelial cell (IEC) gene expression and microbiota composition were assessed following SDR. To determine disease impact, SDR was combined with either Citrobacter rodentium infection or dextran sulfate sodium (DSS)-induced colitis, with interventions including the β-AR inhibitor propranolol and the NADPH oxidase inhibitor apocynin. SDR significantly upregulated expression of Dual oxidase 2 (Duox2) , Dual oxidase maturation factor 2 (Duoxa2) , and inducible nitric oxide synthase 2 (Nos2) in IECs (2- to 8- fold, p &lt; 0.0001), effects reversed by β-AR blockade but not α2-adrenergic, CRH, or glucocorticoid inhibition. SDR also induced microbial dysbiosis, characterized by reduced alpha-diversity and compositional shifts, which was rescued by propranolol. Stress exacerbated disease severity in both infectious ( C. rodentium ) and chemically induced (DSS) colitis, amplifying colonic expression of Duox2 , Nos2 , and Ccl2, especially. Apocynin mitigated stress-induced ROS/RNS production and body weight loss even prior to colitis onset, reduced colonic gene expression of key oxidative enzymes, and alleviated both chemically and infectious colitis severity. These findings provide strong evidence that social stress sensitizes the gut to inflammation through β-adrenergic and NADPH oxidase–driven oxidative stress, highlighting potential therapeutic targets for mitigating stress-exacerbated IBD. Highlights Social disruption (SDR) and restraint stress (RST) activate distinct neuroendocrine pathways, with SDR driving epithelial ROS/RNS pathways via β-adrenergic signaling. β-adrenergic blockade prevents SDR-induced epithelial priming, microbial dysbiosis, and colitis exacerbation. NADPH oxidase inhibition with apocynin mitigates stress-induced oxidative stress and disease severity across different colitis models. Findings identify β-adrenergic and redox pathways as therapeutic targets for stress-exacerbated IBD.

Ingestion of Fermented Food-Associated Microbial Aryl Metabolites Attenuates High Fat Diet-Induced Liver Lipid Accumulation

Physiology · 2025-05-01

Objective: Consumption of fermented foods is associated with lower systemic inflammation and improved metabolic health, yet the underlying mechanisms remain unclear. We investigated if microbial-derived aromatic amino acid metabolites found in fermented foods, specifically phenyllactic acid (PLA), 4-hydroxyphenyllactic acid (4HPLA), and indole-3-lactic acid (ILA), collectively termed aryl-lactates (AL), mitigate liver fat accumulation, a key marker of metabolic-associated steatotic liver disease (MASLD). We also assessed their effects on macrophage phagocytic activity ex vivo. Hypothesis: We hypothesized that AL reduce weight gain and hepatic lipid accumulation when consumed at doses found in fermented foods. Methods: 12-week-old; C57BL/6 male mice were fed either control diet (CD; 15% kcal fat) or high-fat diet (HFD; 45% kcal fat) and provided a combination of AL (4HPLA+PLA+ILA) (0 or 10 µM/d total) in drinking water (n=6/group) for 16 weeks. Body weight, food intake, and water consumption were recorded 3 times/wk. Body composition was analyzed with EchoMRI at wks 8 and 15. After 16 weeks, liver weight was recorded, and hepatic lipid accumulation was assessed via Oil Red O staining. To evaluate the effect of AL on macrophage function, we performed ex vivo assays using macrophages differentiated from human peripheral blood mononuclear cells exposed to 0 or 50 µM of either 4HPLA, ILA, or PLA. Phagocytic activity was assessed using fluorescein stained, lyophilized E. Coli particles. Results: Weight gain, fat mass, and liver weight were increased for HFD compared to CD fed mice. AL decreased weight gain, fat mass, liver weight, and increased lean mass of only HFD fed mice (p&lt;0.05). Additionally, AL attenuated HFD-induced hepatic lipid accumulation (p&lt;0.05). Given the critical role of macrophages in MASLD progression, we examined if these effects were associated with enhanced macrophage function. Ex vivo treatment of macrophages with any of the three AL increased macrophage phagocytosis (p&lt;0.05) with 4HPLA being the most effective. Result summary: AL reduced weight gain, fat mass, and hepatic lipid accumulation of HFD-fed mice. Conclusion: Ingestion of microbial-derived aryl-lactates limited fat accumulation in the liver in response to HFD. Ongoing studies aim to further elucidate the mechanistic contributions of macrophages in this process. These findings highlight the therapeutic potential of fermented food-derived microbial metabolites in mitigating MASLD severity. Funding: USDA-AFRI- 2023-67017-39053 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Social stress worsens colitis through β-adrenergic–driven oxidative stress in intestinal mucosal compartments

Brain Behavior and Immunity · 2025-12-17 · 1 citations

Psychological stress is a known risk factor for inflammatory bowel disease (IBD), but the mechanisms linking stress to worsened disease remain unclear. Because distinct stress paradigms activate different neuroimmune circuits, it is critical to investigate model-specific effects. We examined how social stress primes the gut for heightened inflammation and whether this is mediated by specific neuroendocrine pathways, including α2-/β-adrenergic (sympathetic) or glucocorticoid/ corticotropin-releasing hormone receptor (CRHR1) (HPA axis) signaling. Mice were exposed to social disruption (SDR) stress and pre-treated with pharmacological antagonists targeting α2-adrenergic receptors (idazoxan), β-adrenergic receptor (β-AR) (propranolol), glucocorticoid receptor (mifepristone), or CRHR1 (antalarmin). Intestinal epithelial cell (IEC) gene expression and microbiota composition were assessed following SDR. To determine disease impact, SDR was combined with either Citrobacter rodentium infection or dextran sulfate sodium (DSS)-induced colitis, with interventions including the β-AR inhibitors and the NADPH oxidase inhibitor apocynin. SDR significantly upregulated expression of Dual oxidase 2 (Duox2), Dual oxidase maturation factor 2 (Duoxa2), and inducible nitric oxide synthase 2 (Nos2) in IECs (2- to 8-fold, p < 0.0001), effects reversed by β-AR blockade but not α2-adrenergic, CRH, or glucocorticoid inhibition. SDR also induced microbial dysbiosis, characterized by reduced α -diversity and compositional shifts, which was rescued by propranolol. Stress exacerbated disease severity in both infectious (C. rodentium) and chemically induced (DSS) colitis, amplifying colonic expression of Duox2, Nos2, and Ccl2, especially. Apocynin mitigated stress-induced ROS/RNS production and body weight loss even prior to colitis onset, reduced colonic expression of key oxidative enzymes, especially DUOX2, and alleviated both chemically and infectious colitis severity. These findings provide strong evidence that social stress sensitizes the gut to inflammation through β-adrenergic and NADPH oxidase-driven oxidative stress, highlighting potential therapeutic targets for mitigating stress-exacerbated IBD.

Abstract 2215: Age and indoleamine 2,3-dioxygenase 1 (IDO1) interact to affect gut microbiome homeostasis in subjects with glioblastoma

Cancer Research · 2025-04-21

Objective: This study aimed to determine how age and IDO1 interact to affect the gut microbiome, microbial metabolite levels, and survival outcomes in subjects undergoing IDO1-targeted immunotherapy against glioblastoma (GBM). Methods: Serum and colon contents from younger 16-20- or older 92-118-week-old wild-type (WT), IDO1 knockout (IDO1-KO), and IDO1 enzyme null (IDO1-H350A) mice were analyzed for 16S rRNA gut microbiome composition and microbial-derived aromatic amino acid metabolites via LC/MS/MS. Fecal samples from younger 7-8 or older 81-85 week-old WT mice with intracranial syngeneic GL261 GBM cells treated with or without brain radiation (RT) and PD-1 mAb were also analyzed. Fecal and plasma samples from 12 newly diagnosed IDHwt/MGMT promoter unmethylated GBM patients who received RT plus PD-1 mAb and IDO enzyme inhibitor were also analyzed. Results: Young and old IDO1-KO and IDO1-H350A mice demonstrated a unique gut microbiome signature with an elevated abundance of Bifidobacterium and Helicobacter (p&amp;lt;0.05) and a reduced abundance of Ventriosum and Sireaum compared to WT mice (p&amp;lt;0.001). Prevotellaceae, Muribaculum, Alistipes, Enterohabdus, Clostridia, and Akkermansia were different in IDO1-KO and IDO1-H350A mice compared to WT - but only in older mice. Serum microbiome-derived aryl-lactates phenyllactate (PLA), indolelactate (ILA), and 4-hydroxyphenyllactate (4HPLA) were increased in IDO1-KO mice compared to WT and IDO1-H350A mice (p&amp;lt;0.001) - but only in older mice. Older WT mice with GL261 GBM showed lower fecal aryl-lactates (PLA, 4HPLA, and ILA) compared to younger counterparts. Treatment with RT + PD-1 mAb also showed decreased serum indole propionic acid (IPA) levels in young but not old mice with GL261 GBM (p&amp;lt;0.05). Analyses from GBM patient baseline fecal samples revealed that higher abundance of Massilioclostridium coli, GGB3819-SGB5184, Dysosmobacter welbionis, and Phocaeicola plebeius was associated with longer survival (p&amp;lt;0.004), while higher levels of fecal ILA and lower levels of plasma IPA also correlated with elongated survival (p&amp;lt;0.04 and p&amp;lt;0.01 respectively). Combined treatment of RT + PD-1 mAb + IDO1 inhibitor decreased plasma IPA when compared to baseline (p&amp;lt;0.01). Conclusions: IDO1 possesses both enzyme- and non-enzyme-dependent effects that change gut microbiota composition and bioactive microbial aromatic amino acid metabolite levels, which is also influenced by aging. Certain gut microbiota species and metabolites may serve as prognostic markers for predicting outcomes to immunotherapy for GBM. These findings warrant further investigation into how IDO1 interacts with age to affect immunotherapeutic efficacy in subjects with GBM. Citation Format: Lijie Zhai, Akriti Shrestha, Kristen L. Lauing, Manon Penco-Campillo, Oluwatomilayo Odum, Prashant Bommi, Taylor Koch, Robert H. McCusker, Douglas E. Anderson, Anand Germanwala, Vikram C. Prabhu, Jigisha P. Thakkar, Jacob M. Allen, Rimas V. Lukas, Derek A. Wainwright. Age and indoleamine 2,3-dioxygenase 1 (IDO1) interact to affect gut microbiome homeostasis in subjects with glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2215.

Indoleamine 2, 3 dioxygenase (IDO1) differentiallyModulates Microbial Aromatic Amino Acid Metabolites and Colonic Inflammation in an Age-Dependent Manner

Physiology · 2025-05-01 · 1 citations

Background and Objective: Indoleamine 2,3-dioxygenase (IDO1) metabolizes tryptophan (Trp) into kynurenine (Kyn), an immunosenescence pathway linked to immune dysfunction in aging, though its mechanism remains unclear. Kynurenine activates aryl hydrocarbon receptor (AHR) which further regulates immunity and gut homeostasis by shaping the microbiota. H350A transgenic mice that express IDO1 but are incapable of IDO1 Trp metabolism show distinct immune responses compared to constitutional IDO1 knockout (IDOKO) mice, confirming that IDO1 possesses both enzyme- and non-enzyme-dependent functions. Gut microbes also metabolize aromatic amino acids including Trp, tyrosine (Tyr), and phenylalanine (Phe) into bioactive aryl metabolites that influence IDO1-driven immune responses. We examined how age and IDO1 activity interact to influence the gut microbiome, microbrial-derived aryl metabolites, and colonic inflammation across the lifespan. Hypothesis: Microbial aryl metabolites are lower in WT mice as compared to IDOKO and H350A (IDO1 enzyme null) mice, and in-turn, intestinal inflammation is increased in WT as compared to IDOKO and H350A mice during advanced age. Methods: Serum, ileal content, and colonic content of young 12 –22-week-old WT, H350A, and IDOKO mice, (n=6-8/group) and old 114-138 week old (n=10/group) wwere analyzed for microbial aryl metabolites by LC/MS/MS. Microbe compositionof ileum and colon contents were assessed by 16S rRNA sequencing. Inflammatory markers were assessedin ileum and colon tissues by multiplex qPCR. Results: Tryptophan-derived microbial aryl metabolites, indole-lactate and indole-acetatein aged WT mice as compared to IDOKO or H350A mice (p&lt;0.05).Immune-modulating Tyr-derived 4-hydroxyphenyllactic acid (4-HPLA) and Phe-derived phenyllactate (PLA), were also increased in the serum of aged IDOKO and H350A mice (p&lt;0.05) as compared to aged WT mice. A known microbial derived anti-inflammatory aryl metabolite producer, Bifidobacterium sp, was increased in the colon of young and aged IDOKO mice and also in the ileum of H350A mice compared to WT mice. In contrast, the pro-inflammatory pathobiont, Helicobacter hepaticus, was elevated in the colon of both IDOKO and H350A aged mice compared to aged WT mice, correlating to increased colonic inflammatory markers (Lbp, Tnfrsf1b, S100a8, S100a9, and Ccr2). Conclusion: Enzymatic and non-enzymatic activity of IDO1 influences microbial-derived aryl production, microbiota composition, and colonic inflammation, with effects more pronounced in aging. IDO1 deficiency (genetic/metabolic) alters gut microbial metabolism, with protective consequences depending on microbiota composition.Ongoing studies are examining the role of AHRin mediating IDO1-dependent and independent regulation of gut microbiota and inflammation. This work was supported in part by National Institutes of Health (NIH) grants R01NS097851 (D.A.W.), K02AG068617 (D.A.W.), R01NS129835 (D.A.W.), RO1DK131133 (J.M.A) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Rediscovering the wild: MiFoDB brings fermented food microbiomes into focus

mSystems · 2025-08-19 · 1 citations

Fermented foods have sustained human societies for thousands of years, with their microbial communities subtly shaping flavor, nutrient preservation, and health. Yet despite this long-standing relationship, much of the microbial complexity within fermented foods remains unresolved. In recent work, Caffrey et al. (E. B. Caffrey, M. R. Olm, C. I. Kothe, H. C. Wastyk, et al., mSystems 10:e00141-25, 2025, https://doi.org/10.1128/msystems.00141-25) put forth a new tool, MiFoDB, a metagenomic workflow that offers a promising alternative for advancing food microbiome science. By enabling strain-level resolution, functional gene annotation, and microbial tracking across substrates and time, MiFoDB provides a clearer view into the ecological and functional landscape of the fermented food microbiota. This work also bridges gaps between food and human microbiome research and brings us closer to a mechanistic understanding of how fermented foods influence health, helping transform ancient dietary practices into actionable and targeted nutritional strategies for improving human health and well-being.

Gut microbiota dysbiosis in a novel mouse model of colitis potentially increases the risk of colorectal cancer

American Journal of Physiology-Gastrointestinal and Liver Physiology · 2025-05-05 · 5 citations

Ablation of hnRNPI in intestinal epithelia modulates gut microbiota; causing dysbiosis. Increased ratio of fecal Paraclostridium bifermentans to Dubosiella sp. is a signature of inflammation in hnRNPI knockout mice. hnRNPI knockout exacerbated colitis from dextran sodium sulfate challenge in knockout mice. Bacterial metabolites produced by P. bifermentans and Dubosiella newyorkensis could impact colon health in mice. hnRNPI gene ablation exacerbates chemically induced inflammation and colitis; potentially increasing cancer risk.

Evaluation of the Gerofit to Home Model of Care: A Remote Clinical Exercise Program for Older Adults

Journal of the American Geriatrics Society · 2025-09-26 · 1 citations

OBJECTIVE: The COVID-19 pandemic prompted a rapid shift to remote healthcare. In response, the Veteran Health Administration's (VHA) Gerofit clinical exercise program began providing supervised, group-based exercise sessions three times weekly via real-time video for older Veterans. The longer-term benefits of such remote programs, particularly their impact on mobility and physical function, remain unclear. This analysis evaluates whether the Gerofit to Home (GTH) model of care improves physical performance in older veterans with multiple chronic conditions. DESIGN: This is a retrospective, quality improvement analysis of national data collected in the VHA GTH program from April 2020 to December 2022. Physical function assessments (PFA) including the 2-min step test, 30-s chair stand test, 5 times sit to stand, and the 30-s arm curl test were completed at baseline, 3 months, 6 months, and 1 year time points. To assess functional trajectories for the PFAs, repeated measures ANCOVA, as implemented under mixed models, was applied, adjusting for age, gender, and race. RESULTS: The overall GTH sample included 626 older veterans (mean age of 75 ± 6.8 years). All PFA measures showed similar longitudinal trajectories, with significant improvements from baseline to 1 year (p < 0.01). Each follow-up differed significantly from baseline (ps < 0.01), and clinically meaningful gains were achieved and sustained across all assessments. CONCLUSION: The GTH program demonstrates that remotely delivered, medically supervised group exercise-integrated within a national healthcare system-can produce and sustain meaningful improvements in physical function for older adults. As one of the first large-scale, health care system-supported remote exercise programs for older adults, GTH offers a novel, scalable model for delivering effective, accessible care to high-risk populations.

Rational Design and Optimization of a Potent IDO1 Proteolysis Targeting Chimera (PROTAC)

Journal of Medicinal Chemistry · 2025-02-13 · 20 citations

Indoleamine 2,3-dioxygenase 1 (IDO1) is an immunosuppressive protein that inhibits antitumor immunity through both tryptophan metabolism and nonenzymatic functions. Drugs targeting IDO1 enzyme activity have failed to improve the overall survival of patients with cancer. Developing new therapeutics that neutralize both enzyme- and nonenzyme-derived immunosuppressive IDO1 effects is therefore of high interest. We previously described a novel proteolysis targeting chimera (PROTAC), NU223612, that degrades IDO1 in cultured human glioblastoma (GBM) cells, as well as in well-established brain tumors, in vivo. In this study, we rationally optimized the structure of our lead series to create NU227326, which degrades IDO1 with a DC50 of 5 nM in human GBM cells. Mechanistic studies showed that IDO1 degradation occurred through the ubiquitin–proteasome system and was sustained for at least 2 days, supporting NU227326 as a highly potent IDO1 PROTAC suitable for further studies in GBM and other human cancers.