Colon Tissues Of Mice Research Articles

Extraction of Soluble Dietary Fiber from Sunflower Receptacles (Helianthus annuus L.) and Its Alleviating Effect on Constipation in Mice

Background/Objectives: Sunflower receptacles are the main by-product of the processing of Helianthus annuus L. Methods: In this study, several extraction methods of soluble dietary fiber (SDF) from sunflower receptacles were evaluated, and then, the physicochemical structure and functional properties of these SDFs were examined. Finally, a mouse constipation model was established to explore its therapeutic potential for constipation. Results: The results showed that the SDF yield of citric acid extraction and enzyme extraction was better than that of hot-water extraction. Structural characterization showed that the three SDF functional groups were similar and amorphous, while the surface distribution of the SDF obtained by the citric acid extraction method (ASDF) had more fine pores. Physicochemical analysis showed that ASDF had the best water-holding capacity, oil-holding capacity, and expansion force. Animal experiments showed that the first black stool defecation time of the model group changed significantly (p < 0.001), indicating that the model was successful. Compared with the model group, the middle- and high-dose groups reduced the first black stool defecation time (p < 0.05 or p < 0.01) and increased the fecal water content (p < 0.05). The high-dose group significantly promoted the intestinal peristalsis of mice (p < 0.05). From hematoxylin–eosin (H&E) staining, it can be seen that the three dose groups of ASDF can improve the damage of mouse colon tissue induced by loperamide hydrochloride to a certain extent. Conclusions: Our results show that ASDF has good physical and chemical properties and laxative properties and has broad development space in the field of health food.

Investigation of the chemical composition and biological activities of Eremurus spectabilis M. Bieb through antioxidant, enzyme inhibition, COX-2 and iNOS assessment.

Eremurus spectabilis is widespread and used primarily for medicinal and culinary purposes. This study aimed to evaluate the chemical composition, antiradical and antioxidant activities, enzyme inhibitory activities, and anti-inflammatory properties of various extracts from the aerial parts of E. spectabilis. Various assays were used to investigate the antioxidant and enzyme inhibitory properties. The chemical composition of the tested extracts was analyzed using High-Performance Liquid Chromatography-Electrospray Ionization Tandem Mass Spectrometry (HPLC-ESI-MS/MS). Additionally, the extracts were tested on isolated mouse colon tissue challenged with E. coli lipopolysaccharide (LPS) to replicate the inflammation and oxidative stress burden characteristic of inflammatory bowel diseases. In the chemical composition, vanillic, ferulic, 4-hydroxybenzoic acids were the prominent compounds. The greatest antioxidant activity was observed in the methanol and water extracts from the aerial parts. Enzyme inhibition tests showed that the ethyl acetate extract had the highest anti-acetylcholinesterase activity. The gene expression of pro-inflammatory cyclooxygenase-2 (COX-2) and pro-oxidant inducible nitric oxide synthase (iNOS) biomarkers were assayed. Among the extracts, the methanol extract was the most effective in blunting LPS-induced gene expression of COX-2. E. spectabilis may serve as a valuable source of phytochemicals for combating oxidative stress and inflammation-driven diseases, with a particular emphasis on colon inflammatory condition.

Scutellarein ameliorates dextran sulfate sodium-induced ulcerative colitis by inhibiting colonic epithelial cell proinflammation and barrier disruption

IntroductionScutellarein (Scu) is a natural occurring flavonoid found in multiple traditional Chinese medicines such as Oroxylum indicum (L.) Kurz and Scutellaria baicalensis, with various pharmacological activities including anti-inflammation, anti-oxidation and myocardial protection. Here, we investigated the therapeutic efficacy of Scu on ulcerative colitis (UC) and the underlying mechanism.MethodsEfficacy of Scu on UC was evaluated in dextran sulfate sodium (DSS) induced colitis mouse model. Inflammation in colonic tissues was assessed by myeloperoxidase activity assay and RT-qPCR. Barrier proteins expression was examined using immunostaining and Western blot. IL-1β-treated HT-29 cells was used for mechanical investigation.ResultsGavage of Scu significantly decreased the DAI score, improved colon shortening, ameliorated the pathological score in DSS-treated mice with better efficacy than the positive drug, 5-aminosalicylic acid. Scu also inhibited the expression levels of cytokines (Il-1β, Tnf-α, Il-1α, Il-6, and Cxcl1) as well as barrier proteins (E-cadherin, Occludin, and ZO-1) in colon tissues of DSS mice. In intestinal epithelial HT-29 cells, Scu attenuated the IL-1β-downregulated expression levels of E-cadherin, occludin, and ZO-1, while reduced IL-1β-upregulated IL-6 and IL-8 mRNA levels. Moreover, Scu inhibited the phosphorylation and nuclear translocation of NF-κB and suppression of NF-κB phosphorylation abolished IL-1β-disrupted epithelial barrier integrity and IL-1β-upregulated proinflammatory mediators expression in HT-29 cells.ConclusionThese data demonstrate that Scu is an efficacious therapeutic agent to treat UC. Inhibition of inflammatory responses and maintenance of epithelial barrier integrity through NF-κB signaling pathway underlines Scu therapeutic effect on UC.

N4BP3 facilitates NOD2-MAPK/NF-κB pathway in inflammatory bowel disease through mediating K63-linked RIPK2 ubiquitination

The NOD2 signaling pathway, which plays an important role in the mechanisms of inflammatory bowel disease (IBD) development, has been closely associated with ubiquitination. It was revealed in this study that NOD2 receptor activation could obviously affect the expression of 19 ubiquitination-related genes, with N4BP3 being the most prominently expressed and upregulated. In addition, N4BP3 knockdown was found to reduce the mRNA levels of MDP-induced inflammatory factors, while N4BP3 overexpression elevated their mRNA levels as well as the levels of phospho-ERK1/2, phospho-JNK, phospho-P38 and phospho-NF-κB P65 proteins. Immunoprecipitation tests showed that N4BP3 could pull down RIPK2 and promote its K63-linked ubiquitination. In human tissue specimen assays and mouse experiments, we found that the expression of N4BP3 was significantly elevated in Crohn’s disease (CD) patients and IBD mice, and N4BP3 knockdown reduced the dextran sulfate sodium-induced pathological score and the expression of inflammatory factors in the mouse colon tissue. In conclusion, N4BP3 is able to interact with RIPK2 and promote its K63-linked ubiquitination, to further promote the NOD2-MAPK/NF-κB pathway, thereby increasing promoting the release of inflammation factors and the degree of IBD inflammation.

Real meat shows good performance in maintaining colonic barrier function at a recommended amount of intake in mice

AbstractThis study aimed to investigate the effects and underlying mechanisms of real meat and plant‐based meat analogs on intestinal barrier function. Methods including PacBio sequencing, HPLC, histological observation, western blotting, and RT‐qPCR were jointly applied. The results indicated that the 20‐day intake of real meat and plant‐based meat analogs did not induce pathological changes in the colon of mice. However, when the feeding period was extended to 68 days, inflammatory infiltration was observed in the colon tissue of mice fed plant‐based meat analogs. In contrast, the intake of real meat for 68 days performed better in maintaining colonic barrier function by reducing cell apoptosis, increasing the number of goblet cells and mucin secretion, and upregulating the expression of tight junction proteins. High‐throughput sequencing revealed that the intake of real meat significantly altered the composition of the gut microbiota, reducing pro‐inflammatory bacteria (such as Bacteroides, Desulfovibrio, or Colidextribacter) or pathogenic bacteria (such as Mucispirillum and Helicobacter) while increasing beneficial bacteria (such as Lactobacillus, Bifidobacterium, or Roseburia). Additionally, the consumption of real meat markedly decreased protein fermentation in the colon, leading to lower levels of cadaverine, putrescine, and spermine. These changes associated with real meat consumption reduced the activation of the pro‐inflammatory pathway (lipopolysaccharide/Toll‐like receptor 4/nuclear factor kappa B), ultimately maintaining colonic homeostasis in mice. Collectively, the findings suggest that the proper intake of real meat is more beneficial for colon health in mice compared to plant‐based meat analogs.

Dipeptidyl peptidase 4: A predictor of ferroptosis in ulcerative colitis.

With its rapidly increasing incidence and prevalence, ulcerative colitis (UC) has become a major global health challenge. Recent evidence suggests that ferroptosis plays a significant role in the development of UC. However, the relationship between ferroptosis and the progression of UC needs to be extensively studied. The differentially expressed genes in UC patients were screened from the GEO database. The ferroptosis-related genes were obtained from FErrDB and GeneCards. The UC subtypes were identified with the R package "CancerSubtype" and evaluated with consensus clustering (CC) to identify gene expression patterns in patients with UC. The key genes were detected with qRT-PCR, Western blot, and immunohistochemistry in vitro and in vivo models. Ferroptosis was identified with western blotting on ferrotic-associated proteins and staining on Fe2+ with commercial FerroOrange kits. Dipeptidyl peptidase 4 (DPP4), also known as CD26, is a potential biomarker for ferroptosis in UC patients. Transcriptome sequencing data showed a positive correlation between decreased DPP4 expression and proinflammatory cytokines such as TNF-α, IL-6, and IL-β, as well as immune cell infiltration in the colon tissues of UC patients. Furthermore, DPP4 was strongly associated with ferroptosis biomarkers, particularly in Subtype 2 of UC. Interestingly, our study also found that DPP4 expression was significantly reduced in RSL3-treated ferroptotic intestinal epithelial cells, more so than in LPS-treated cell models. Inhibition of DPP4 had a significant impact on the expression of ferroptotic biomarkers. Additionally, DPP4 expression was decreased in the colon tissues of DSS-treated mice, and the ferroptosis inhibitor Ferritin-1 effectively counteracted the effects of DSS on immune cell infiltration, colon length, and DPP4 expression. DPP4 can serve as a biomarker for ferroptosis in the diagnosis and management of UC.

Demethylzeylasteral alleviates inflammation and colitis via dual suppression of NF-κB and STAT3/5 by targeting IKKα/β and JAK2

BackgroundUlcerative colitis (UC) is a common inflammatory bowel disease and a risk factor of colorectal cancer. Demethylzeylasteral (DZT), a bioactive component mainly isolated from Tripterygium wilfordii, has been shown to inhibit inflammation and cancer. However, its anti-UC function and molecular mechanisms have not been well characterized. This study aims to explore the therapeutic effect and functional targets of demethylzeylasteral against UC. MethodsRT-qPCR, Western blot and ELISA were used to detect the generation of pro-inflammatory cytokines and chemokines in murine macrophage cells. Luciferase reporter gene, Western blot, pull-down, CETSA, DARTS, and virtual docking were employed to detect the anti-inflammatory targets and molecular mechanisms of demethylzeylasteral. The anti-inflammatory and anti-colitis effects of demethylzeylasteral were further determined in DSS-challenged mice. ResultsIn vitro, demethylzeylasteral inhibited NO and PGE2 production by suppressing the mRNA and protein expression of iNOS and COX-2, and suppressed the mRNA expression of TNF-α, IL-1β, IL-6, MCP-1, CXCL9, and CXCL10 in RAW264.7 macrophages stimulated by LPS/IFNγ. Furthermore, demethylzeylasteral was not only capable of inhibiting IKKα/β-NF-κB activation, but also able to block JAKs-STAT3/5 activation in LPS/INFγ-incubated RAW264.7 cells or DSS-exposed colon tissues of mice. Mechanistically, demethylzeylasteral was found to directly bind to IKKα/β and JAK2 kinases, leading to inactivation of pro-inflammatory signaling cascades and reduced generation of cytokines and chemokines. In vivo, oral administration of demethylzeylasteral significantly attenuated DSS-induced colitis, which was mainly manifested as mitigated symptoms of colitis, colonic mucosal barrier damage, and colonic inflammation. ConclusionWe demonstrated that demethylzeylasteral alleviated UC pathology by blocking NF-κB and STAT3/5 pathways via targeting IKKα/β and JAK2 kinases, raising the possibility that demethylzeylasteral could act as a candidate for the treatment of UC.

Novel mechanisms of intestinal flora regulation in high-altitude hypoxia

BackgroundThis study investigates the molecular mechanisms behind firmicutes-mediated macrophage (Mψ) polarization and glycolytic metabolic reprogramming through HIF-1α in response to intrinsic mucosal barrier injury induced by high-altitude hypoxia. MethodsEstablishing a hypoxia mouse model of high altitude, we utilized single-cell transcriptome sequencing to identify key cell types involved in regulating intestinal mucosal barrier damage caused by high-altitude hypoxia. Through proteomic analysis of colonic tissue Mψ and metabolomic analysis of Mψ metabolites, we determined crucial proteins and metabolic pathways influencing intestinal mucosal barrier damage induced by high-altitude hypoxia. Mechanistic validation was conducted using RAW264.7 Mψ in vitro by assessing cell viability with CCK-8 assay following treatment with different metabolites. The hypoxia mouse model was further validated in vivo by transplanting gut microbiota of Firmicutes. Histological examinations through H&E staining assessed colonic cell morphology and structure, while the FITC-dextran assay evaluated intestinal tissue permeability. Hypoxia probe signal intensity in mouse colonic tissue was assessed via metronidazole staining. Various experimental techniques, including flow cytometry, immunofluorescence, ELISA, Western blot, and RT-qPCR, were employed to study the impact of HIF-1α/glycolysis pathway and different gut microbiota metabolites on Mψ polarization. ResultsBioinformatics analysis revealed that single-cell transcriptomics identified Mψ as a key cell type, with their polarization pattern playing a crucial role in the intestinal mucosal barrier damage induced by high-altitude hypoxia. Proteomics combined with metabolomics analysis indicated that HIF-1α and the glycolytic pathway are pivotal proteins and signaling pathways in the intestinal mucosal barrier damage caused by high-altitude hypoxia. In vitro cell experiments demonstrated that activation of the glycolytic pathway by HIF-1α led to a significant upregulation of mRNA levels of IL-1β, IL-6, and TNFα while downregulating mRNA levels of IL-10 and TGFβ, thereby promoting M1 Mψ activation and inhibiting M2 Mψ polarization. Further mechanistic validation experiments revealed that the metabolite butyric acid from Firmicutes bacteria significantly downregulated the protein expression of HIF-1α, GCK, PFK, PKM, and LDH, thus inhibiting the HIF-1α/glycolytic pathway that suppresses M1 Mψ and activates M2 Mψ, consequently alleviating the hypoxic symptoms in RAW264.7 cells. Subsequent animal experiments confirmed that Firmicutes bacteria inhibited the HIF-1α/glycolytic pathway to modulate Mψ polarization, thereby mitigating intestinal mucosal barrier damage in high-altitude hypoxic mice. ConclusionThe study reveals that firmicutes, through the inhibition of the HIF-1α/glycolysis pathway, mitigate Mψ polarization, thereby alleviating intrinsic mucosal barrier injury in high-altitude hypoxia.

In vivo absorption, in vitro digestion, and fecal fermentation properties of non-starch polysaccharides from Chinese chestnut kernels and their effects on human gut microbiota

Non-starch polysaccharides are major bioactive components in chestnuts, and can serve as water-soluble polysaccharides with potential prebiotic properties. This study aims to establish an in vitro digestion and fermentation model to reveal the digestive and fermentative characteristics of Non-starch polysaccharides from chestnut kernels (NSPCK). The results indicated that under simulated digestion, NSPCK was partially digested in gastric juice but remained significantly undigested in saliva and intestinal juice, demonstrating considerable resilience against hydrolysis. After digestion, NSPCK still exhibited stable rough, lamellar, and porous structure and maintained strong antioxidant capacity. Animal experiments revealed positive effects of NSPCK on blood lipid level, and colon tissue of mice. Moreover, NSPCK enhanced the accumulation of short-chain fatty acids during fermentation, particularly acetic acid, propionic acid, and butyric acid. Furthermore, NSPCK intervention increased the abundance of beneficial bacteria such as Lactobacillus and Bifidobacterium, and at the same time reduced that of harmful bacteria such as Enterococcus.

Oral food-derived whey protein isolate-Tremella fuciformis polysaccharides pickering emulsions with adhesive ability to delivery magnolol for targeted treatment of ulcerative colitis

Magnolol (Mag) is a promising natural compound with therapeutic potential for ulcerative colitis (UC). Here we designed and fabricated an oral food-grade whey protein isolate-Tremella fuciformis polysaccharides (WPI-TFPS) stabilized pickering emulsions to encapsulate Mag (Mag-WPI-TFPS) for targeted treatment of UC. With the assistance of the WPI-TFPS, pickering emulsions were well encapsulated and formed stable microparticles with a particle size of approximately 9.49 ± 0.047 μm, a 93.63 ± 0.21 % encapsulation efficiency and a loading efficiency of 21.53 ± 0.01 %. In vitro, the formulation exhibited sustained-release properties in simulated colon fluid with a cumulative release rate of 60.78 % at 48 h. In vivo, the Mag-WPI-TFPS specifically accumulated in the colon tissue for 24 h with stronger fluorescence intensity, which demonstrated that TFPS and WPI had a good adherence ability to inflamed mucosa by electrostatic attraction and ligand-receptor interactions. As expected, compared with Free-Mag, the oral administration of Mag-WPI-TFPS remarkably alleviated the symptoms of UC and protected the colon tissue in DSS-induced UC mice. More importantly, WPI-TFPS enhanced gut microbiota balance by increasing the diversity and relative abundances of Lactobacillaceae and Firmicutes. Overall, this study presents a convenient, eco-friendly, food-derived oral formulation with potential as a dietary supplement for targeted UC treatment.

Long-term exposure to polystyrene microplastics reduces macrophages and affects the microbiota–gut–brain axis in mice

The remarkably increase in plastic use has led to worldwide pollution involving microplastics (MPs), which have been shown to be potentially hazardous substances. Although several studies have focused on the effects of small MPs on the brain and behavior of aquatic species, their effects on the mouse brain and the underlying mechanisms remain unclear. Our study’s aim was to investigate the effects of long-term oral ingestion of different sizes of MPs (0.1, 5, and 50 μm) on mouse colon tissue. Of these sizes, the smallest (0.1 μm) had the greatest effect. Pre-administration of MP promotes colitis but reduces tumor growth in a colitis-associated colorectal cancer (CAC) mouse mode. MPs can increase inflammation in mice via activation of the very late antigen 4–vascular cell adhesion molecule 1 (VLA4-VCAM1) signaling pathway in macrophages, while also inducing macrophage reduction in the late phase of inflammation. In the microbiota–gut–brain axis, polystyrene MP treatment altered bile acid and carbohydrate metabolism in the intestine, inhibited intestinal motility, reduced water reabsorption, and led to a certain degree of depression in mice. These findings suggest that small MPs can induce macrophage reduction, thereby affecting the physical and mental health by modulating the microbiota–gut–brain axis.

The highly stabilized biologically derived peptide VIESPPEI alleviates DSS-induced colitis in mice by preventing colonic atrophy and modulating gut microbiota

Ulcerative colitis (UC) is a recurrent gastrointestinal infection within the spectrum of inflammatory bowel disease, posing risks associated with long-term medication. Consequently, this study aimed to investigate the effects of novel duck liver protein-derived bioactive peptides, characterized by a higher safety profile, on dextran sulfate sodium (DSS)-induced colitis in mice. The results indicated that the retention rate of VIESPPEI remained above 80% following simulated gastrointestinal digestion in vitro, demonstrating strong stability and practical application potential. The peptide VIESPPEI was found to alleviate weight loss in colitis-afflicted mice, inhibit the elevation of the disease activity index (DAI), and significantly improve symptoms such as atrophy and shortening of the colon. Histological examination further confirmed that VIESPPEI intake promoted the recovery of acute colitis in mice. Additionally, it significantly reduced myeloperoxidase activity and increased superoxide dismutase and catalase activities in the colon tissues of UC mice. Notably, a substantial increase in beneficial gut microbiota and the populations of three beneficial bacteria species (Anaplasma spp., Tannabacterium spp., and Bifidobacterium spp.) that produce short-chain fatty acids, along with a decrease in pathogenic groups such as Streptococcus and Turicibacter, may account for the improvement in colitis observed in UC mice treated with VIESPPEI. This study provides new evidence that VIESPPEI alleviates UC and provides a theoretical basis for the development of functional peptide beverages.

2-(3,4-dihydroxyphenyl)ethyl 3-hydroxybutanoate (HTHB) ameliorates cognitive dysfunction via modulating gut microbiota in aged senescence-accelerated mouse prone 8 (SAMP8) mice.

Numerous studies have indicated a close association between gut microbiota dysbiosis, inflammation, and cognitive impairment, highlighting their crucial role in the aging process. 2-(3,4-Dihydroxyphenyl)ethyl 3-hydroxybutanoate (HTHB), a novel derivative of hydroxytryrosol (HT), known for its metabolic and anti-inflammatory properties, was investigated for its effects on memory, inflammation, and gut microbiota in senescence-accelerated mouse prone 8 (SAMP8) mice. The study employed behavioral testing, biochemical detection and 16S RNA analysis. Results revealed that HTHB mitigated memory decline and lymphocyte aberrance, reduced inflammation in the brain cortex, intestine and peripheral system, and modulated gut microbiota dysbiosis. Interestingly, the cognitive function and serum inflammation of mice significantly correlated with differences in gut microbiota in SAMP8 mice. Furthermore, HTHB treatment exhibited an enhancement of gut barrier integrity in colon tissue in SAMP8 mice. In vitro experiments using HCT116 and DLD1 cells further evidenced that HTHB rescued the tight junction protein levels impaired by lipopolysaccharide (LPS). These finding demonstrate that HTHB effectively ameliorates cognitive dysfunction in aged mice, might by modulating gut microbiota, suppressing inflammation and promoting intestinal barrier integrity. This highlights the potential of HTHB as a therapeutic agent for age-related cognitive loss.

Human placental mesenchymal stromal cells alleviate intestinal oxidative damage in mice with graft-versus-host disease via CD73/adenosine/PI3K/Akt/GSK-3β axis

BackgroundIntestinal damage is a common and serious complication in patients with graft-versus-host disease (GVHD). Human placental mesenchymal stromal cells (hPMSCs) ameliorate GVHD tissue damage by exerting anti-oxidative effects; however, the underlying mechanisms remain not fully clear. MethodsA GVHD mouse model and tumor necrosis factor-α (TNF-α)-stimulated human colon epithelial cell lines NCM460 and HT-29 cells were used to investigate the mechanisms of hPMSCs alleviating GVHD-induced intestinal oxidative damage. ResultshPMSCs reduced TNF-α concentrations and the number of CD3+TNF-α+ T-cells, which were negatively correlated with the expression of claudin-1, occludin, and ZO-1, through CD73 in the colon tissue of GVHD mice. Meanwhile, hPMSCs reduced the mean fluorescence intensity (MFI) of reactive oxygen species (ROS) and the concentration of malondialdehyde (MDA), promoted superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) activities, as well as claudin-1, occludin, and ZO-1 expression, in colonic epithelial cells of GVHD mice and TNF-α-stimulated cells via CD73. Moreover, hPMSCs upregulated adenosine (ADO) concentrations in GVHD mice and TNF-α-stimulated cells and mitigated the loss of tight junction proteins via the CD73/ADO/ADO receptors. Further analysis showed that hPMSCs diminished Fyn expression and enhanced Nrf2, GCLC, and HO-1 expression in both TNF-α-stimulated cells and colonic epithelial cells of GVHD mice by activating PI3K/Akt/GSK-3β pathway. ConclusionsThe results suggested that hPMSC-mediated redox metabolism balance and promoted tight junction protein expression were achieved via CD73/ADO/PI3K/Akt/GSK-3β/Fyn/Nrf2 axis, by which alleviating intestinal oxidative injury in GVHD mice.

Moxibustion with seed-size moxa cones alleviates colonic injury in mice with ulcerative colitis by regulating TLR4/MyD88/NF-κB signaling pathway.

To observe the effect of moxibustion with seed-size moxa cones on the Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear transcription factor-κB(NF-κB) signaling pathway in mice with ulcerative colitis(UC), so as to explore the therapeutic mechanism of moxibustion with seed-size moxa cones on colonic injury in UC. Forty male C57BL/6 mice were randomly divided into blank group, model group, moxibustion group, and western medicine group, with 10 mice in each group. The UC mouse model was established by 3% DSS solution by free drinking for 7 consecutive days. Mice in the moxibustion group were treated with seed-size moxa cones at "Zhongwan"(CV12), "Tianshu"(ST25) and "Shangjuxu"(ST37), 3 moxa cones per point, with each cone applied for approximately 30 s, while mice in the western medicine group were orally administered with 300 mg/kg mesalazine solution, which were both conducted once a day for 7 consecutive days. The general condition of mice was observed every 2 days, and the disease activity index (DAI) score was calculated. HE staining was used to observe the morphology of colonic tissue in mice. ELISA was used to detect the serum interleukin(IL)-1β, tumor necrosis factor(TNF)-α, IL-6, and IL-8 contents. Immunohistochemistry was used to detect the positive expression of TLR4 and MyD88 in colonic tissue of mice. Real-time fluorescence quantitative PCR was used to detect the expression levels of TLR4, MyD88, and NF-κB p65 mRNAs in colonic tissue. Compared with the blank group, varying degrees of soft or watery stools were observed, colon length and body weight were decreased(P<0.01) in mice of the model group, while DAI score, colon weight index, mucosal damage score, colonic pathological score, serum IL-1β, TNF-α, IL-6, and IL-8 contents, positive expressions of TLR4 and MyD88, and TLR4, MyD88, and NF-κB p65 mRNA expressions in colonic tissue were increased(P<0.01). Compared with the model group, improved fecal characteristics were observed, colon length and body weight were increased(P<0.01) in mice of the moxibustion group and western medicine group, while DAI scores, colon weight indexes, mucosal damage scores, colonic pathological score, serum contents of IL-1β, TNF-α, IL-6, and IL-8, positive expressions of TLR4 and MyD88, and TLR4, MyD88, and NF-κB p65 mRNA expressions in colonic tissue were decreased(P<0.01, P<0.05). There was no significant difference in the above indicators between the moxibustion group and the western medicine group. Moxibustion with seed-size moxa cones may alleviate colonic injury in UC mice by regulating the TLR4/MyD88/NF-κB signaling pathway and reducing the release of inflammatory factors.

Role of gut microbiota in Crohn's disease pathogenesis: Insights from fecal microbiota transplantation in mouse model.

Inflammatory bowel disease, particularly Crohn's disease (CD), has been associated with alterations in mesenteric adipose tissue (MAT) and the phenomenon termed "creeping fat". Histopathological evaluations showed that MAT and intestinal tissues were significantly altered in patients with CD, with these tissues characterized by inflammation and fibrosis. To evaluate the complex interplay among MAT, creeping fat, inflammation, and gut microbiota in CD. Intestinal tissue and MAT were collected from 12 patients with CD. Histological manifestations and protein expression levels were analyzed to determine lesion characteristics. Fecal samples were collected from five recently treated CD patients and five control subjects and transplanted into mice. The intestinal and mesenteric lesions in these mice, as well as their systemic inflammatory status, were assessed and compared in mice transplanted with fecal samples from CD patients and control subjects. Pathological examination of MAT showed significant differences between CD-affected and unaffected colons, including significant differences in gut microbiota structure. Fetal microbiota transplantation (FMT) from clinically healthy donors into mice with 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced CD ameliorated CD symptoms, whereas FMT from CD patients into these mice exacerbated CD symptoms. Notably, FMT influenced intestinal permeability, barrier function, and levels of proinflammatory factors and adipokines. Furthermore, FMT from CD patients intensified fibrotic changes in the colon tissues of mice with TNBS-induced CD. Gut microbiota play a critical role in the histopathology of CD. Targeting MAT and creeping fat may therefore have potential in the treatment of patients with CD.

Sanguinarine alleviates ulcerative colitis in mice by regulating the Nrf2/NF-κB pathway

To investigate the mechanism of sanguinarine (SA) for alleviating ulcerative colitis (UC) induced by dextran sodium sulfate (DSS) in mice. Male C57BL/6 mouse models of 3.5% DSS-induced UC were randomized for treatment with 1, 5 and 10 mg/kg SA by gavage, 400 mg/kg sulfasalazine by gavage, or 10 mg/kg SA combined with intraperitoneal injection of 30 mg/kg ML385 (a Nrf2 inhibitor). The changes in intestinal inflammation was assessed by monitoring weight changes, disease activity index (DAI) score, colon length measurement, and HE staining. After the treatments, the colon tissues were collected for detection of malondialdehyde (MDA) content using colorimetry, mRNA expressions of inflammatory factors using RT-qPCR, and the expressions of Nrf2, HO-1, Keap-1, p-p65, p65, occludin, and ZO-1 proteins were detected using Western blotting. SA treatment obviously alleviated weight loss, colon length shortening and DAI score increase and ameliorated structural destruction of the colon glands and colonic crypts in mice with DSSinduced UC. SA intervention significantly decreased the levels of TNF-α, IL-1β and IL-6 mRNA and lowered ROS and MDA levels in the colon tissue of UC mice. The mouse models receiving SA treatment showed significantly increased expressions of Nrf2, HO-1, occludin and ZO-1 and lowered expressions of Keap-1 and P-P65 in the colon tissue without significant changes of p65 expression, and these changes were SA dose-dependent. Treatment with ML385 obviously attenuated the effect of highdose SA for improving UC in the mouse models. SA can improve UC-like enteritis in mice possibly by activating the Nrf2 pathway and inhibiting the NF-κB pathway in the colon tissue.

Effects of the N-Butanol Extract of Pulsatilla Decoction on Neutrophils in a Mouse Model of Ulcerative Colitis.

Ulcerative colitis (UC) is a chronic inflammatory disease, the incidence of which is increasing worldwide. However, the etiology and pathogenesis of UC remains unclear. The n-butanol extract of Pulsatilla decoction (BEPD), a traditional Chinese medicine, has been shown to be effective in treating UC. This study aimed to explore the molecular mechanism underlying the effects of BEPD on UC, in particular its effects on neutrophil extracellular trap (NET) formation by neutrophils. High-performance liquid chromatography was used to determine the principal compounds of BEPD. UC was induced in mice using dextran sodium sulfate, and mice were treated with 20, 40, or 80 mg/kg BEPD daily for seven days. Colonic inflammation was determined by assessing the disease activity index, histopathology, colonic mucosal damage index, colonic mucosal permeability, and pro- and anti-inflammatory cytokine levels. The infiltration and activation status of neutrophils in the colon were determined by analyzing the levels of chemokine (C-X-C motif) ligand (CXCL) 1 and CXCL2, reactive oxygen species, Ly6G, and numerous NET proteins. The findings suggest that BEPD improved the disease activity index, histopathology, and colonic mucosal damage index scores of mice with UC, and restored colonic mucosal permeability compared with untreated mice. The expression levels of the pro-inflammatory cytokines interleukin-1β, interleukin-6, and tumor necrosis factor-α in colon tissues were significantly decreased, while the expression levels of anti-inflammatory cytokines in colon tissues were significantly increased, exceeding those of control mice. In addition, BEPD reduced the expression of the neutrophil chemokines CXCL1 and CXCL2 in the colon tissue of mice with UC, reduced neutrophil infiltration, reduced reactive oxygen species levels, and significantly reduced the expression of NET proteins. BEPD also significantly reduced NET formation. The results of this study suggest that BEPD exerts therapeutic effects in a murine model of UC by inhibiting neutrophil infiltration and activation in the colon, as well as by inhibiting the expression of key proteins involved in NET formation and reducing NET formation, thereby alleviating local tissue damage and disease manifestations.

Rapid Analysis of Colonic Metabolomics in High-Fat Diet Mice by Extraction Electrospray Ionization Mass Spectrometry (EESI-MS).

Propolis exhibits significant anti-inflammatory, antidiabetic, and antiobesity properties in both mouse models and clinical applications. However, the underlying metabolic mechanisms remain poorly understood. Traditional metabolomic methods that rely on chromatographic separation require complex preprocessing steps and extended detection periods. In this study, we employed extraction electrospray ionization mass spectrometry combined with multivariate analysis to directly profile metabolites in the colon tissue of mice. Our findings demonstrate the efficacy of ethanol extract of propolis (EEP) in mitigating weight gain, reducing inflammatory cytokines, and improving insulin resistance induced by a high-fat diet. Additionally, EEP enhanced glucose tolerance. Through collision-induced dissociation experiments, we identified 26 metabolites, with 4-hydroxyphenylacetic acid, protocatechuic acid, caffeic acid, ferulic acid, hippuric acid, histidine, and tryptophan emerging as potential biomarkers. Notably, tryptophan exhibited the highest content at 8.25 mg/g. Our research facilitates rapid profiling of colon metabolites, underscoring its significant potential for broader applications in animal metabolomic studies.

Expression and pathogenesis of insulin-like growth factor-1 and insulin-like growth factor binding protein 3 in a mouse model of ulcerative colitis

Background and aimInsulin-like growth factor-1 may be involved in the epithelial-to-mesenchymal transition process. It can mitigate adverse effects when interacting with insulin-like growth factor binding protein 3. This study aimed to explore alterations in the expression of these two factors in the colonic tissue of mice with ulcerative colitis. MethodThis study utilized animal models. Mice were randomly allocated into three distinct groups. Disease activity index assessment was performed first, followed by histological grading of colitis. Protein and mRNA expression levels were determined using Western blotting and RT-qPCR. Immunohistochemical detection was used to determine histochemistry scores. Pearson correlation and SPSS 25.0 software were used for data analysis. ResultsThe findings indicated a reduction in the expression of the two investigated factors as well as in epithelial-to-mesenchymal transition epithelial markers during inflammation, while the expression of noninflammatory factors increased. These effects were notably amplified following treatment. Interestingly, the changes in epithelial-to-mesenchymal transition-inducing factors and mesenchymal markers contradicted this trend. Pearson correlation analysis revealed a correlation between molecular indicators of change and epithelial-to-mesenchymal transition. ConclusionInsulin-like growth factor-1 and insulin-like growth factor binding protein 3 may play a protective role in the development and progression of ulcerative colitis, potentially through their inhibition of the epithelial-to-mesenchymal transition. These factors hold promise as targets for the clinical diagnosis and treatment of ulcerative colitis.