Background:Multiple meta-analyses have evaluated the effect of Helicobacter pylori (H. pylori) infection on the risk of inflammatory bowel disease (IBD), but their findings remain inconsistent. Objectives:To critically assess the existing meta-analyses regarding the effect of H. pylori infection on the risk of IBD. Data sources and methods:We systematically searched four electronic databases and manually screened the reference lists of relevant articles. A Measurement Tool to Assess systematic Reviews 2 (AMSTAR-2), Grade of Recommendation, Assessment, Development, and Evaluation (GRADE), and Graphical Representation of Overlap for OVErviews (GROOVE) tools were used to assess the methodological quality, evidence quality, and the overall overlap of primary studies among the included meta-analyses, respectively. A second-order meta-analysis was performed on primary studies derived from the included meta-analyses with very high overlap to explore the association of H. pylori infection with IBD risk and identify the potential influencing factors. Risk ratios (RRs) were pooled. Results:Twelve meta-analyses encompassing 55 primary studies were included. The methodological quality was very low to low. The evidence quality was very low to high. Moderate- to high-quality evidence suggested that H. pylori virulence factors and antibiotic exposure influenced the association of H. pylori infection with IBD. The overall overlap of primary studies among the included meta-analyses was very high. A second-order meta-analysis demonstrated a negative association of H. pylori infection with any type of IBD (RR: 0.60), Crohn's disease (RR: 0.53), ulcerative colitis (RR: 0.67), and IBD-unclassified (RR: 0.73). Subgroup analyses demonstrated that both IBD subtypes (P interaction < 0.01), regions (P interaction < 0.01), and study designs (P interaction = 0.05) significantly influenced this association. Sensitivity analyses indicated that the results were robust across IBD subtypes and regions, but not across the study designs. Discussion:H. pylori infection seems to exhibit a protective effect against the risk of IBD. This association may be influenced by IBD subtypes, regions, H. pylori virulence factors, and antibiotic exposure. Design:This is an umbrella review. Registration:The protocol was prospectively registered with the PROSPERO database (CRD42024559958).

Background: Multiple meta-analyses have evaluated the effect of Helicobacter pylori ( H. pylori ) infection on the risk of inflammatory bowel disease (IBD), but their findings remain inconsistent. Objectives: To critically assess the existing meta-analyses regarding the effect of H. pylori infection on the risk of IBD. Data sources and methods: We systematically searched four electronic databases and manually screened the reference lists of relevant articles. A Measurement Tool to Assess systematic Reviews 2 (AMSTAR-2), Grade of Recommendation, Assessment, Development, and Evaluation (GRADE), and Graphical Representation of Overlap for OVErviews (GROOVE) tools were used to assess the methodological quality, evidence quality, and the overall overlap of primary studies among the included meta-analyses, respectively. A second-order meta-analysis was performed on primary studies derived from the included meta-analyses with very high overlap to explore the association of H. pylori infection with IBD risk and identify the potential influencing factors. Risk ratios (RRs) were pooled. Results: Twelve meta-analyses encompassing 55 primary studies were included. The methodological quality was very low to low. The evidence quality was very low to high. Moderate- to high-quality evidence suggested that H. pylori virulence factors and antibiotic exposure influenced the association of H. pylori infection with IBD. The overall overlap of primary studies among the included meta-analyses was very high. A second-order meta-analysis demonstrated a negative association of H. pylori infection with any type of IBD (RR: 0.60), Crohn’s disease (RR: 0.53), ulcerative colitis (RR: 0.67), and IBD-unclassified (RR: 0.73). Subgroup analyses demonstrated that both IBD subtypes ( P interaction < 0.01), regions ( P interaction < 0.01), and study designs ( P interaction = 0.05) significantly influenced this association. Sensitivity analyses indicated that the results were robust across IBD subtypes and regions, but not across the study designs. Discussion: H. pylori infection seems to exhibit a protective effect against the risk of IBD. This association may be influenced by IBD subtypes, regions, H. pylori virulence factors, and antibiotic exposure. Design: This is an umbrella review. Registration: The protocol was prospectively registered with the PROSPERO database (CRD42024559958).

Paneth cells (PCs) are specialized epithelial cells mainly found in the small intestine. PCs have a key role in shaping microbiome. It is considered as a site of origin for intestinal inflammation. Previous studies indicate that vitamin D3/vitamin D receptor (VDR) are implicated in IBD and infection. However, there is limited study exploring Paneth cells with VDR deficiency in ileitis and dysbiosis. We hypothesize that VDR deficiency in Paneth cells alters the microbiome and makes the host susceptible to small intestinal inflammation (e.g., ileitis). Paneth cells isolated from conditional PC KO (VDRΔPC) mice and the lox controls (VDRloxp) were analyzed by single cell RNA sequencing. Small intestinal microbiome of VDRΔPC mice were detected by Fluorescence in situ hybridization (FISH). To rule out confounding factors that may contribute to the intestinal epithelial VDR regulation, we have established enteroids derived from the VDRΔPC and VDRloxp mice. SAMP1/YitFc mice develop a spontaneous ileitis that is similar in many features to human Crohn’s Disease (CD). We then examined the expression level of VDR in the SAMP1/YitFc mice with ileitis. The Paneth cells in the ileal organoids isolated from VDRloxp and VDRΔPC mice were labeled with lysozyme via immunofluorescence staining. The number of lysozyme-positive cell, ie Paneth cell, were counted in each bud of the organoids. There were significantly reduced Paneth cells in the VDRΔPC organoids, compared to the VDRloxp organoids. Further, we observed dislocation of small intestinal microbiome and penetration to epithelial cells in the VDRΔPC mice. The mucus thickness was significantly reduced in the small intestine of the VDRΔPC mice, compared to the VDRloxp mice. Conditional VDR deletion in Paneth cell severely changed antimicrobial peptides, DEFA3, DEFA4 and DEFA21 expression decreased in the ileum of VDRΔPC mice, compared to the VDRloxp mice. Development of ileitis is accelerated by the presence of luminal bacteria and is characterized by discontinuous segmental inflammation in the ileum while sparing the proximal small intestine and colon. We found abnormal Paneth cells in the SAMP1/YitFc mice, compared with the control AKR mice. VDR and VDR target ATG16L1 were significantly reduced in the small intestine of the SAMP1/YitFc mice with ileitis via RT-PCR, WB, and IHC staining. Our current observation suggests that PC VDR deletion makes the host susceptible to small intestinal inflammation. Insights gained from understanding how VDR is integrally involved in regulating Paneth cell function may serve as a novel paradigm for understanding the mechanism of host-microbial interactions. This effort can facilitate new therapeutic targets for CD by assessing cellular and molecular responses or resistance to treatment.

Background: The association of Helicobacter pylori ( H. pylori ) infection with non-alcoholic fatty liver disease (NAFLD) remains controversial. Additionally, its associations with metabolic dysfunction-associated fatty liver disease (MAFLD), metabolic dysfunction-associated steatotic liver disease (MASLD), and Chinese MAFLD using the most recent diagnostic criteria were unclear. Objective: To analyze the associations of H. pylori infection with NAFLD, MAFLD, MASLD, and Chinese MAFLD. Design: Cross-sectional study. Methods: This study screened 1172 inpatients who underwent both H. pylori test and liver computed tomography or ultrasound examination between June 2020 and May 2024. Multivariate Logistic regression analyses were performed to evaluate the associations of H. pylori infection with the severity of hepatic steatosis and risk of hepatic fibrosis. Adjusted odds ratios (aORs) with 95% confidence intervals (CIs) were calculated after adjusting for gender, hypertension, hyperlipidemia, body mass index (BMI), fasting plasma glucose (FPG), and high-sensitivity C-reactive protein (HsCRP) in NAFLD analyses; age, gender, drinking, hypertension, diabetes, hyperlipidemia, BMI, and HsCRP in MAFLD analyses; and gender, drinking, hypertension, hyperlipidemia, BMI, FPG, and HsCRP in MASLD and Chinese MAFLD analyses. Results: Overall, 875, 982, 869, and 869 patients were included in NAFLD, MAFLD, MASLD, and Chinese MAFLD analyses, respectively. In NAFLD, MAFLD, MASLD, and Chinese MAFLD analyses, 257, 302, 257, and 257 patients had H. pylori infection, respectively. H. pylori infection was independently associated with severe hepatic steatosis in NAFLD (aOR = 3.956; 95% CI = 1.171–13.359, p = 0.027), MAFLD (aOR = 3.730; 95% CI = 1.083–12.850, p = 0.037), and MASLD (aOR = 3.962; 95% CI = 1.158–13.557, p = 0.028) analyses, but not Chinese MAFLD analyses. However, H. pylori infection was not independently associated with the risk of hepatic fibrosis. Conclusion: H. pylori infection may increase the severity of NAFLD, MAFLD, and MASLD, but not the risk of liver fibrosis.

BACKGROUND: Alzheimer’s disease (AD) is a progressive neurodegenerative condition affecting 39 million people worldwide. Although classically framed as a brain disorder, accumulating evidence also points to early disturbances in intestinal physiology. These gut changes may affect neural functions through the microbiota–gut–brain axis, where microbial and host signals engage immune, endocrine, and enteric pathways. OBJECTIVE: We hypothesize that 3xTg-AD mice exhibit early dysbiosis and disruption of intestinal barrier integrity. We tested whether intestinal microbiota composition and epithelial barrier integrity are altered in 3xTg-AD mice before and during symptomatic stages compared with age-matched controls. METHODS: Gross anatomy was assessed in 12-mo. 3xTg-AD mice and controls. Barrier integrity was evaluated by immunofluorescence for tight junction proteins (e.g. ZO-1, ZO-2) in ileum and colon. Intestinal permeability was measured by a permeability tracer assay (FITC-dextran), and serum endotoxin was quantified by chromogenic Limulus amebocyte lysate (LAL). Bacterial distribution was assessed on colonic sections by fluorescent in situ hybridization (FISH) using the universal EUB338 probe and an Akkermansia muciniphila–specific probe. Relative abundance of selected taxa was quantified by real-time PCR at 4 mo. (presymptomatic) and 12 mo. (symptomatic). RESULTS: At 12 mo., 3xTg females showed reduced body weight, increased lung weight, and shorter ileum compared to controls, whereas 3xTg males showed increased body weight and longer ileum compared to controls. Tight junction changes were region specific. In 3xTg mice, ZO-1 staining was discontinuous in both colon and ileum. ZO-2 expression was reduced at epithelial and endothelial colonic junctions in 3xTg mice, with more apical concentration in ileum compared to diffuse distribution in controls. Serum FITC-dextran did not differ significantly between groups, although 3xTg mice trended towards a higher mean. Serum endotoxin was significantly elevated in 3xTg mice compared with controls (p=0.0271). FISH with EUB 338 probe revealed predominantly luminal bacterial distribution in both groups, while A. muciniphila staining showed mucin-associated localization that appeared similar across groups. Across 4 and 12 mo., 3xTg-AD mice showed lower Firmicutes, Bacteroides spp. 2, and Lactobacillus versus controls. At 4 mo., 3xTg mice showed increased Butyrivibrio spp. 1, decreased Bacteroides spp. 1, and increased Bacteroides spp. 3, while 12 mo. 3xTg mice showed the inverse. Species-level shifts were observed for Adlercreutzia equolifaciens, A. muciniphila, and Blautia hansenii (decreased at 4 mo. and increased at 12 mo. in 3xTg-AD vs age-matched controls). CONCLUSIONS: Our studies have shown that presymptomatic species-level microbiota shifts at the species level are followed by symptomatic, region-specific barrier disruption with altered tight junctions and elevated serum endotoxin. These data suggest that dysbiosis and barrier dysfunction are potential contributors to AD and support the evaluation of intestinal and microbial changes as early biomarkers. FUNDING: NIDDK/National Institutes of Health grant R01 DK134343, R01DK114126, VA Merit Award VA 1 I01 BX004824-06, and VA Collaborative Merit Award 1 I01BX006878-01A1 to Jun Sun. This abstract was presented at the American Physiology Summit 2026 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.

Background: Vitamin D3 and its receptor vitamin D receptor (VDR) are implicated in the pathogenesis of inflammatory bowel disease and susceptibility to infection. Paneth cells (PCs) are specialized epithelial cells mainly found in the small intestine. PCs have a key role in shaping microbiome and inflammatory responses. However, there is limited study exploring PCs with VDR deficiency in ileitis and dysbiosis. Hypothesis: We hypothesize that VDR deficiency in Paneth cells alters the microbiome and makes the host susceptible to small intestinal inflammation (e.g., ileitis). Methods/Models: Small intestinal microbiome of VDRΔPC mice were detected by Fluorescence in situ hybridization. SAMP1/YitFc mice develop a spontaneous ileitis that is similar in many features to human Crohn’s Disease. We then examined the expression level of VDR in the SAMP1/YitFc mice with ileitis. To rule out confounding factors that may contribute to the intestinal epithelial VDR regulation, we have established a germ-free VDRΔIEC mouse model. We also cultured enteroids derived from the conditional PC KO (VDRΔPC) and VDRLoxp mice. Paneth cells isolated from VDRΔPC and VDRloxp mice were analyzed by single cell RNA sequencing. Results: We observed dislocation of small intestinal microbiome and penetration to epithelial cells in the VDRΔPC mice. The mucus thickness was significantly reduced in the small intestine of the VDRΔPC mice, compared to the VDRloxp mice. We found abnormal Paneth cells in the SAMP1/YitFc mice, compared with the control AKR mice. VDR and VDR target ATG16L1 were significantly reduced in the small intestine of the SAMP1/YitFc mice with ileitis via RT-PCR, WB, and IHC staining. Interestingly, the differences of PCs between specific pathogen free VDRΔIEC mice and VDRloxp controls were markedly reduced when both mice groups were germ-free. Paneth cells in the ileal organoids isolated from VDRloxp and VDRΔPC mice were labeled with lysozyme via immunofluorescence staining. The number of lysozyme-positive cell, i.e. Paneth cell, were counted in each bud of the organoids. There were significantly reduced number of Paneth cells in the VDRΔPC enteroids. At the molecular level, single cell RNA sequencing of Paneth cells isolated VDRΔPC mice and VDRloxp showed that changes of VDR target genes include AMPs and α-defensin. PC VDR involves processes in broad-spectrum humoral immune response, predominantly mediated by AMPs and a tissue repair mechanism for proliferation. Conclusion and Future Plan: PC VDR deletion makes the host susceptible to small intestinal inflammation. Insights gained from understanding how VDR is integrally involved in regulating PC function may serve as a novel paradigm for understanding host-microbial interactions and new therapeutic targets for ileitis. This abstract was presented at the American Physiology Summit 2026 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.

Background:The association of Helicobacter pylori (H. pylori) infection with non-alcoholic fatty liver disease (NAFLD) remains controversial. Additionally, its associations with metabolic dysfunction-associated fatty liver disease (MAFLD), metabolic dysfunction-associated steatotic liver disease (MASLD), and Chinese MAFLD using the most recent diagnostic criteria were unclear. Objective:To analyze the associations of H. pylori infection with NAFLD, MAFLD, MASLD, and Chinese MAFLD. Design:Cross-sectional study. Methods:This study screened 1172 inpatients who underwent both H. pylori test and liver computed tomography or ultrasound examination between June 2020 and May 2024. Multivariate Logistic regression analyses were performed to evaluate the associations of H. pylori infection with the severity of hepatic steatosis and risk of hepatic fibrosis. Adjusted odds ratios (aORs) with 95% confidence intervals (CIs) were calculated after adjusting for gender, hypertension, hyperlipidemia, body mass index (BMI), fasting plasma glucose (FPG), and high-sensitivity C-reactive protein (HsCRP) in NAFLD analyses; age, gender, drinking, hypertension, diabetes, hyperlipidemia, BMI, and HsCRP in MAFLD analyses; and gender, drinking, hypertension, hyperlipidemia, BMI, FPG, and HsCRP in MASLD and Chinese MAFLD analyses. Results:Overall, 875, 982, 869, and 869 patients were included in NAFLD, MAFLD, MASLD, and Chinese MAFLD analyses, respectively. In NAFLD, MAFLD, MASLD, and Chinese MAFLD analyses, 257, 302, 257, and 257 patients had H. pylori infection, respectively. H. pylori infection was independently associated with severe hepatic steatosis in NAFLD (aOR = 3.956; 95% CI = 1.171-13.359, p = 0.027), MAFLD (aOR = 3.730; 95% CI = 1.083-12.850, p = 0.037), and MASLD (aOR = 3.962; 95% CI = 1.158-13.557, p = 0.028) analyses, but not Chinese MAFLD analyses. However, H. pylori infection was not independently associated with the risk of hepatic fibrosis. Conclusion:H. pylori infection may increase the severity of NAFLD, MAFLD, and MASLD, but not the risk of liver fibrosis.

BACKGROUNDS AND AIMS:SNX27, member of the sorting nexin (SNX) family, carries a unique PDZ domain and mediates recycling of endocytosed transmembrane proteins. SNX27 is critical for neurodevelopmental processes, however its role in intestine remains unexplored. We aim to determine the previously unknown roles of SNX27 in regulating intestinal homeostasis, epithelial barrier integrity, and inflammatory responses. METHODS:We used available datasets to analyze SNX27 expression in human IBD. We generated a novel mouse model of SNX27 conditional deletion from intestinal epithelial cells (SNX27ΔIEC) and challenged these mice with Dextran Sulfate Sodium (DSS). RESULTS:SNX27 expression was significantly lower in human IBD, including UC and CD. SNX27ΔIEC mice had significantly lower bodyweight and exhibited increased proliferation and poor differentiation of secretory Paneth and Goblet cells. We found reduced mucin layer and downregulation of crucial epithelial barrier proteins β-catenin, E-cadherin, ZO-1, and Claudin10 in SNX27ΔIEC mice. SNX27ΔIEC mice showed high intestinal permeability and spontaneously developed intestinal inflammation. Moreover, SNX27ΔIEC mice were more susceptible towards DSS-induced colitis, compared to the SNX27Loxp mice. CONCLUSION:Overall, deletion of intestinal epithelial SNX27 weakens barrier functions and promotes inflammation. Our results indicate a novel role of SNX27 in regulating intestinal physiology and protecting against intestinal disorders. Thus, understanding the mechanisms of SNX27 downregulation in IBD will provide insights into new prevention and targets against chronic inflammation.

Background and aims:Transfer RNA (tRNA) modifications determine translation fidelity and efficiency. It occurs through the action of specific enzymes that modify the nucleotides within the tRNA molecule. Our previous study demonstrated tRNA modopathies and altered queuine-related metabolites in inflammatory bowel diseases. Queuine tRNA-ribosyltransferase catalytic subunit 1 (QTRT1) and QTRT 2 co-localize in mitochondria and form a heterodimeric TGT participating in tRNA Queuosine (tRNA-Q) modification. Human body acquires Queuine/Vitamin Q from intestinal microbiota or from diet. However, the roles of tRNA-Q modifications in the maintenance of intestinal mitochondrial homeostasis and microbiome are still unclear. Methods:We used publicly available human IBD datasets, QTRT1 knockout (KO) mice, QTRT1 intestinal epithelial conditional KO (QTRT1 ΔIEC ) mice, cultured cell lines with QTRT1-specific siRNA, and organoids from patients with IBD to investigate the mechanism of tRNA-Q modifications in intestinal mitochondrial homeostasis and therapeutic potential in anti-inflammation. Results:In single cell RNA sequencing datasets of human IBD, we identified significant reduced intestinal epithelial QTRT1 expression in the patients with Crohn's Disease. Using publicly available datasets, we identified significantly changes of Vitamin Q-associated bacteria in human IBD, compared to the healthy control. Qtrt1 -/- mice had significant reduction of Q-associated bacteria, e.g., Bacteroides . Alcian Blue and Mucin-2 staining revealed mucosal barrier damage and disrupted homeostasis, with reduced colonic cell proliferation. Intestinal tight junction integrity was impaired in QTRT1-KO mice, as evidenced by reduced ZO-1 and increased Claudin-10 expression. QTRT1 ΔIEC mice also showed dysbiosis and disrupted TJs. ATP synthesis was significantly decreased in the colon of QTRT1-KO mice, accompanied by severe mitochondrial dysfunction: reduced mitochondrial quality, Cytochrome-C release, and mitochondrial DNA (mtDNA) leakage. Mitochondrial dysfunction contributed to colonic cell death, as shown by elevated expressions of Cleaved Caspase-3 and Cleaved Caspase-1, increased BAX/Bcl-2 ratio, and positive TUNEL signals. Elevated CDC42, CD14, and CD4 levels in QTRT1-KO colon suggested mucosal immune activation and tissue repair responses. QTRT1-deficient CaCO2-BBE cells showed mitochondrial dysfunction. Cytochrome-C and mito-DNA release leading to cell death characterized by elevated expressions of Cleaved Caspase-3 and Caspase-1, increased BAX/Bcl-2 ratio, and higher apoptosis rate. Organoids isolated from patients with IBD showed reduced levels of QTRT1 and dysfunctional mitochondria. Restoring mitochondrial function leads to enhanced QTRT1. Conclusions:These findings underscore the critical role of QTRT1/Q-tRNA modification in maintaining intestinal and microbial homeostasis. Mechanistically, QTRT1 loss impacts mitochondrial integrity and mucosal homeostasis. Our study highlights the novel roles of tRNA-Q modification in maintaining mucosal barriers and innate immunity in intestinal health. What is already known about this subject?:Eukaryotes acquire queuine (q), also known as Vitamin Q, as a micronutrient factor from intestinal microbiota or from diet.Vitamin Q is needed for queuosine (Q) modification of tRNAs for the protein translation rate and fidelity.Queuine tRNA-ribosyltransferase catalytic subunit 1 (QTRT1) is reduced in human IBD.However, health consequences of disturbed availability of queuine and altered Q-tRNA modification in digestive diseases remain to be investigated. What are the new findings?:QTRT1 deficiency leads to altered microbiome and reduced Vitamin Q-associated bacteria in human IBD and a QTRT1 KO animal model.QTRT1 protects the host against losing intestinal integrity during inflammation.QTRT1 localizes in mitochondria and plays novel functions by maintaining intestinal mitochondrial function. QTRT1 loss impacts tRNA modification in the intestine, linking to mitochondrial integrity and mucosal homeostasis.Human IBD showed reduced levels of QTRT1 and dysfunctional mitochondria. Restoring mitochondrial function leads to enhanced QTRT1. How might it impact on clinical practice in the foreseeable future?:Targeting tRNA-Q modification in enhancing mitochondrial function will be a novel method to maintain intestinal health.

Dysbiosis microbiota can drive chronic inflammation by disrupting immune regulation, especially in macrophages. Mitochondria, vital for energy production, play a central role in macrophage metabolic and immune functions. The vitamin D receptor (VDR), a nuclear receptor activated by Vitamin D, modulates macrophage activity. Our prior research demonstrated that myeloid-specific VDR knockout (VDRΔLYZ) mice showed elevated levels of Candida Albicans, a phenomenon observed in human inflammatory bowel disease (IBD). However, the role of VDR in regulating macrophage mitochondrial function and the inflammatory response remains unclear. We hypothesize that mitochondrial dysfunction in VDR-deficient macrophages impairs host defense mechanisms. VDRΔLYZ mice were generated by crossing VDRLoxP mice with Lyz-cre mice. Colitis was induced using 5% Dextran Sulfate Sodium (DSS). Bone marrow-derived macrophages were generated by stimulating cells from mouse femurs and tibias with 20 ng/mL M-CSF. To assess host-microbiota interactions, VDR-deficient and wild-type macrophages were treated with Mito-Q, a mitochondria-targeted antioxidant, followed by an infection with Candida Albicans. Mitochondrial dynamics (fusion/fission), reactive oxygen species (ROS) production, cytochrome C release, and cell death were measured using Mito-track immunofluorescence, MitoSox Red staining, flow cytometry, and Western blotting. VDR-/- macrophages from VDRΔLYZ mice exhibited pronounced mitochondrial dysfunction, with increased fusion/fission, elevated mitochondrial ROS, cytochrome C and mtDNA release, and heightened apoptosis and pyroptosis (via Caspase 3 and Caspase 1). Macrophages from mice with DSS-induced colitis displayed severe mitochondrial abnormalities: elevated fusion/fission, cytochrome C and mtDNA release, decreased Tomm20 expression, and increased mitophagy. These defects led to increased cell death, confirmed by flow cytometry and increased cleaved caspase 3 and BAX/Bcl-2 ratio. Functionally, VDR-/- macrophages showed impaired fungal clearance post Candida Albicans infection. Mito-Q treatment significantly restored mitochondrial function by reducing ROS, inhibiting cytochrome C and mtDNA release, and preventing macrophage death. It also normalized immune function, decreasing the expression of CD206, CD163, and IL-10 in macrophages of VDRΔLYZ mice. VDR deficiency in macrophages results in profound mitochondrial dysfunction, increased oxidative stress, and impaired immune responses, leading to ineffective pathogen clearance and chronic inflammation. Mito-Q effectively restored mitochondrial integrity and reduced cell death. This study underscores the critical role of VDR in maintaining macrophage homeostasis and suggests therapeutic potential for addressing mitochondrial dysfunction in inflammation-related diseases.

Vitamin D / Vitamin D Receptor (VDR) signaling is regulated by several factors: environmental, host, and microbial. Dysregulation in VDR signaling increases breast cancer risk. Intestinal VDR deletion in mice alters the microbiome, increases intestinal permeability, and promotes growth and bacterial infiltration of breast tumors. Dysbiosis also occurs in breast cancer. Several depleted intestinal bacteria (ex. H. biformis) and tumor bacteria (Lactobacilli and Pseudomonas) are thought to exert anti-tumor effects. The factors that cause these bacterial shifts and their exact contribution to cancer remain unclear. We hypothesize that VDR deficiency predisposes to breast cancer by depleting tumor suppressive bacteria from the intestine and tumor. To test our hypothesis, we conditionally deleted the VDR in intestinal epithelial cells (VDRΔIEC) of mice with 7, 12-dimethylbenzanthracene (DMBA)-induced breast cancer. A separate group was administered a cocktail of probiotic bacteria to evaluate if the dysbiosis caused by VDR deletion could be reversed. The intestinal and tumor microbiomes were studied using 16S rRNA sequencing of feces (VDRLoxp and VDRΔIEC mice +/- probiotics) and tissue samples (VDRLoxp and VDRΔIEC mice). To validate our findings, we pooled three studies of the human breast microbiome and performed hypothesis testing after batch correction to identify bacteria altered in breast cancer. Many bacterial genera (51) and species (10) had significantly (q<=0.05) different abundances in the intestines of the VDRΔIEC mice relative to VDRLoxp mice, including depletion of the genera Lactobacilli and Pseudomonas. Within the probiotic control group, VDR deletion also significantly (q= 0.014) depleted F. rodentium, the mouse analog to H. biformis. Administration of probiotics did not rescue the depletion of F. rodentium or any other altered bacteria except for Candidatus Arthromitus. We further found that the breast tumor microbiomes from VDRΔIEC mice were significantly different from those of the VDRLoxp mice with 12 differentially abundant bacterial species (2 enriched / 10 depleted). These include depletion of Lactobacilli species (L. johnsonii, L. reuteri, and L. intestinalis) and Pseudomonas species (P. azotoformans, P. fluorescens, or P. synxantha) from the breast tumor microenvironment. Our analysis of public data showed that the genera Lactobacilli and Pseudomonas are both significantly (q<=0.05) depleted in human breast tumors relative to tissue from healthy patients. Intestinal VDR deficiency alters the bacteria in the intestines and breast tumors of DMBA mice. The Vitamin D / VDR signaling dependent beneficial microbiome may contribute to the prevention of breast cancer pathogenesis. Further studies are needed to validate these findings and evaluate if this mechanism can be targeted to prevent or treat breast cancer. Duncan J. Claypool, Yong-guo Zhang, Yinglin Xia, Jun Sun. Intestinal vitamin D receptor deficiency changes the tumor-associated microbiome in a breast cancer model [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 6525.

BACKGROUND:Aggressive systemic inflammation due to activation of macrophage-derived excessive immune responses is a critical cause of sepsis leading to clinical death. The effect of cycloastragenol (CAG) on cecal ligation and puncture (CLP)-induced systemic inflammation in mice with sepsis and the underlying mechanism are still unknown. PURPOSE:Here, we firstly investigated the ameliorative functions of CAG in CLP-induced systemic inflammation in sepsis and LPS-mediated inflammatory response, and the impact of Toll-like receptor 4 (TLR4) pathway on the anti-inflammatory effects of CAG. METHODS:The in vitro effect of CAG on RAW264.7 cells and THP-1-derived macrophages induced by LPS was detected with quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA), and Western blotting (WB) assays. In addition, the association of TLR4-MD2 complex with CAG was measured through molecular docking, molecular dynamics (MD) simulation, surface plasmon resonance imaging (SPRi), cellular thermal shift assay (CETSA), immunofluorescence and WB. A specific inhibitor of TLR4 receptor TAK-242 and a TLR4-encoding adenovirus were adopted for verifying the functions of CAG. Meanwhile, the in vivo effects of CAG on cardiopulmonary structure, inflammatory factors and survival of CLP-induced septic mice were analyzed through hematoxylin and eosin staining, qPCR, ELISA, and survival analysis. RESULTS:CAG hindered the LPS-induced production of inflammatory mediators like TNF-α, IL-6 and IL-1β within macrophages in vitro. It also inhibited MAPK and NF-κB pathway activation induced by binding of LPS to TLR4 receptor. As suggested by molecular docking results, the MD2-CAG binding energy was -9.53 kcal/mol. During the MD simulation, CAG could tightly bind to the binding pocket of MD2. SPRi revealed that the equilibrium dissociation constant (KD) value for CAG and TLR4 was 5.24× 10-9 M. Moreover, CAG enhanced the thermal stability of TLR4 by approximately 2.68 °C. It further inhibited the binding between LPS-488 and cell membrane receptors. These inhibitory effects of CAG could be partly reversed by TLR4 overexpression and could not increase by specifically blocking TLR4. In vivo, CAG attenuated cardiopulmonary injury and inflammation and improved survival in septic mice dose-dependently. CONCLUSION:CAG exerts its anti-inflammatory activity through suppressing MAPK and NF-κB pathway activation caused by TLR4 activation and inhibiting inflammatory factor production dose-dependently.

Emerging evidence has shown that gut-brain barrier dysfunction occurs at the early stages of ALS. Previous studies demonstrated that sodium butyrate significantly prolonged the life span of ALS mice. Riluzole is the first FDA-approved drug for ALS treatment. We hypothesize that Riluzole and sodium butyrate combined treatment further decreases aggregation of the h-SOD1G93A, restores the gut-brain barrier function, and delays ALS progression. SOD1G93A mice (9-10-week-old) were treated with Riluzole (10 mg/kg, I.P. daily), sodium butyrate (2% in drinking water), or Riluzole and sodium butyrate combination for 6 weeks. The Riluzole/butyrate combination showed a significantly longer rotarod time, increased grip strength, and enhanced intestinal barrier, as compared with Riluzole or sodium butyrate-only treatment. More reduction of h-SOD1G93A aggregation was observed in the colon, spinal cord lumbar, and brain cortex with Riluzole and sodium butyrate combination, compared with Riluzole or sodium butyrate-only treatment. Tight junction proteins (ZO-1 and Claudin-5) significantly increased in the colon, spinal cord lumbar, and brain cortex of mice with Riluzole and sodium butyrate treatment. The Riluzole and sodium butyrate combination reduced serum lipopolysaccharides and h-SOD1G93A aggregation, and inflammatory cytokines more than those in Riluzole or sodium butyrate-only treatment. Overall, Riluzole and sodium butyrate treatment is more effective than either Riluzole or sodium butyrate-only in delaying ALS progress. It provides a potential therapeutic strategy and mechanism by restoring barrier function through the gut-brain axis for ALS.