Search Results (126)

Background: As a well-known environmental hazard, ambient fine particulate matter (PM_2.5, aerodynamic diameter ≤ 2.5 µm) has been positively correlated with an increased risk of digestive system diseases, including appendicitis, inflammatory bowel disease, and gastrointestinal cancer. Additionally, PM_2.5 exposure has been shown to alter microbiota composition and diversity in human and animal models. However, its impact on goblet cells and gut mucus barrier integrity remains unclear. Methods: To address this, 8-week-old male and female interleukin-10 knockout (IL10^−/−) mice, serving as a spontaneous colitis model, were exposed to concentrated ambient PM_2.5 or filtered air (FA) in a whole-body exposure system for 17 weeks. Colon tissues from the PM_2.5-exposed mice and LS174T goblet cells were analyzed using H&E staining, transmission electron microscopy (TEM), and transcriptomic profiling. Results: The average PM_2.5 concentration in the exposure chamber was 100.20 ± 13.79 µg/m³. PM_2.5 exposure in the IL10^−/− mice led to pronounced colon shortening, increased inflammatory infiltration, ragged villi brush borders, dense goblet cells with sparse enterocytes, and lipid droplet accumulation in mitochondria. Similar ultrastructure changes were exhibited in the LS174T goblet cells after PM_2.5 exposure. Transcriptomic analysis revealed a predominantly upregulated gene expression spectrum, indicating an overall enhancement rather than suppression of metabolic activity after PM_2.5 exposure. Integrated enrichment analyses, including GO, KEGG, and GSEA, showed enrichment in pathways related to oxidative stress, xenobiotic (exogenous compound) metabolism, and energy metabolism. METAFlux, a metabolic activity analysis, further substantiated that PM_2.5 exposure induces a shift in cellular energy metabolism preference and disrupts redox homeostasis. Conclusions: The findings of exacerbated gut barrier impairment and goblet cell dysfunction following PM_2.5 exposure provide new evidence of environmental factors contributing to colitis, highlighting new perspectives on its role in the pathogenesis of colitis. Full article

The development of reversible animal models for the study of intestinal pathologies is essential to reduce the number of animals used in research and to better understand disease mechanisms. In this study, we present a reversible model of intestinal malabsorption through the administration of sublethal doses of ebulin-f, a ribosome-inactivating protein, and validate its usefulness by monitoring vitamin C absorption. The scientific community increasingly recognizes the importance of rationalizing experimental designs, optimizing treatment protocols, and minimizing the use of animals in research models. Thus, new methodologies are needed to minimize invasive sampling and to develop reversible animal models that recover physiologically post-study. Such models are essential for in vivo studies of human pathologies. Sublethal doses of ebulin-f (2.5 mg/kg) administered intraperitoneally to female Swiss CD1 mice (n = 6 per group) can cause reversible intestinal alterations in the small intestine, which offer the possibility of having a valuable reversible study model of malabsorption for the investigation of this syndrome. To verify whether nutrient absorption is altered, we used vitamin C as a traceable nutrient that can be quantified in the blood. Peripheral blood samples were collected through the retro-orbital area at 30, 80, 120, 180, and 1440 min post-administration, treated with DTT and MPA, and analyzed using a validated UV/Vis–HPLC method to indirectly determine vitamin C absorption by enterocytes. Pharmacokinetic analysis revealed significantly increased vitamin C absorption on days 1 and 3 post-treatment (AUC values of 3.65 × 10⁴ and 7.10 × 10⁴, respectively) compared to control (0.94 × 10⁴), with partial recovery by day 22 (3.27 × 10⁴). Blood concentration profiles indicate that intestinal damage peaks at day 3, followed by significant regeneration by day 22, establishing this as a viable reversible model for inflammatory bowel disease research. Full article

Epithelial linings are crucial for the maintenance of physiological barriers. The intestinal epithelial barrier (IEB) consists of enterocytes through tight junctions and mucus-secreting cells and can undergo physiological modifications throughout life. To reproduce as closely as possible the IEB main features over time, in vitro co-cultures of Caco2/HT-29 70/30 formed by parental Caco2 and HT-29 cells sub-cultivated for more than 40 passages were set up. The measurements of the transepithelial electrical resistance (TEER) identified two populations: physiological TEER co-cultures (PC) with values > 50 Ωcm² formed by parental cells with fewer than 40 passages, and leaky TEER co-cultures (LC) with values < 50 Ωcm² formed by parental cells with more than 40 passages. In LC, paracellular permeability increased in parallel. By immunofluorescence and Western blot analysis, an increase in claudin 2 was observed in LC vs. PC, with no differences in occludin expression. MUC-2 immunoreactivity was stronger in PC than in LC. LC also showed an enhanced vulnerability to TNFα+IFN-γ. These results reproduce the main morpho-functional modifications reported in the human leaky/aged gut and support the usefulness of our in vitro cell model for studying the molecular processes underlying these modifications and testing drug/nutraceutical treatments to ameliorate leaky gut aging. Full article

Objective: Cholesin is a recently discovered gut-derived hormone secreted by enterocytes upon dietary cholesterol uptake via the transmembrane sterol transporter Niemann–Pick disease C1-like intracellular cholesterol transporter 1 (NPC1L1). In the liver, cholesin activates G protein-coupled receptor 146 (GPR146), causing reduced cholesterol synthesis. In this exploratory, hypothesis-generating study based on post hoc analysis, human data on the cholesin system are presented. Methods: Mucosal biopsies were collected throughout the intestinal tract from 12 individuals with type 2 diabetes (T2D) and 12 healthy, matched controls. Upper small intestinal mucosal biopsies were collected from 20 individuals before and after Roux-en-Y gastric bypass (RYGB) surgery. Liver biopsies were collected from 12 men with obesity and 15 matched controls without obesity. Subcutaneous abdominal adipose tissue biopsies were collected from 20 men with type 1 diabetes (T1D). All biopsies underwent full mRNA sequencing. Results: Cholesin mRNA expression was observed throughout the intestinal tracts of the individuals with T2D and the controls, in the livers of men with and without obesity, and in adipose tissue of men with T1D. NPC1L1 mRNA expression was robust throughout the small intestines but negligible in the large intestines of both individuals with and without T2D. RYGB surgery induced the expression of NPC1L1 mRNA in the upper small intestine. GPR146 mRNA was expressed in the livers of men, both with and without obesity, and in the adipose tissue of men with T1D, but not in the intestines. Conclusions: Our results suggest a role of the cholesin system in human physiology, but whether it is perturbed in metabolic diseases remains unknown. Clinical trial registration numbers: NCT03044860, NCT03093298, NCT02337660, NCT03734718. Full article

Celiac disease (CeD) is a chronic, lifelong, multifactorial, polygenic, and autoimmune disorder, characteristically triggered by exposure to the exogenous factor “gluten” in genetically predisposed individuals, with resulting duodenal inflammation and enteropathy, as well as heterogeneous multisystemic and extraintestinal manifestations. The immunopathogenesis of CeD is complex, favored by a peculiar human leukocyte antigen (HLA) genetic predisposition, leading to gluten presentation by antigen-presenting cells to CD4+ T helper (Th) cells, T cell–B cell interactions, and production of specific antibodies, resulting in the immune-mediated killing of enterocytes and, macroscopically, in duodenal inflammation. Here, the most relevant correlations between cellular and molecular aspects and clinical manifestations of this complex disease are reviewed, with final considerations on nutritional aspects for disease management. Full article

Background/Objectives: The barrier properties of the human small intestine play a crucial role in regulating digestion, nutrient absorption and drug metabolism. Current in vitro organotypic models consist only of an epithelium, which does not take into account the possible role of stromal cells such as fibroblasts or the extracellular matrix (ECM) which could contribute to epithelial barrier properties. Therefore, the aim of this study was to determine whether these stromal cells or ECM were beneficial or detrimental to barrier function when incorporated into an organotypic human small intestine model. Methods: Intestinal epithelial cell lines or primary cell organoids derived from the epithelial stem cells of the small intestine were cultivated either on a porous Transwell membrane (epithelial model) or on a primary small intestinal stromal cell-populated collagen-fibrin hydrogel (full thickness model). Results: Both models expressed villin (enterocytes), lysozyme (Paneth cells), Ki67 (proliferative cells) and zonula occludens-1 (tight junctions). The polarized epithelial barriers of the full thickness models demonstrated a significant decrease in transepithelial electrical resistance (TEER) with values comparable to that found in the native small intestine in contrast to the higher TEER values observed in the epithelial models. This correlated to an increase in secreted zonulin, a regulator of intestine permeability, in the full thickness models. The decreased TEER values were due to both the stromal cells and the choice of the hydrogel versus the Transwell membrane. Moreover, erythropoietin and epithelial growth factor secretion, which have roles in regulating barrier integrity, directly correlated with the changes in TEER and permeability. Conclusions: This study emphasizes the importance of different cell types being incorporated into small intestine models and, also, the influence of the scaffold or matrix used. Full article

Gut health is crucial in many ways, such as in improving human health in general and enhancing production in agricultural animals. To maximize the effect of a healthy gastrointestinal tract (GIT), an understanding of the regulation of intestinal functions is needed. Proper intestinal functions depend on the activity, composition, and behavior of intestinal epithelial cells (IECs). There are various types of IECs, including enterocytes, Paneth cells, enteroendocrine cells (EECs), goblet cells, tuft cells, M cells, and intestinal epithelial stem cells (IESCs), each with unique 3D structures and IEC distributions. Although the communication between IECs and other cell types, such as immune cells and neurons, has been intensively reviewed, communication between different IECs has rarely been addressed. The present paper overviews the networks among IECs that influence intestinal functions. Intestinal absorption is regulated by incretins derived from EECs that induce nutrient transporter activity in enterocytes. EECs, Paneth cells, tuft cells, and enterocytes release signals to activate Notch signaling, which modulates IESC activity and intestinal homeostasis, including proliferation and differentiation. Intestinal immunity can be altered via EECs, goblet cells, tuft cells, and cytokines derived from IECs. Finally, tools for investigating IEC communication have been discussed, including the novel 3D intestinal cell model utilizing enteroids that can be considered a powerful tool for IEC communication research. Overall, the importance of IEC communication, especially EECs and Paneth cells, which cover most intestinal functional regulating pathways, are overviewed in this paper. Such a compilation will be helpful in developing strategies for maintaining gut health. Full article

Background/Objectives: Campylobacter jejuni (CJ) is the etiological agent of the world’s most common intestinal infectious food-borne disease, ranging from mild symptoms to fatal outcomes. The development of innovative synbiotics that inhibit the adhesion and reproduction of multidrug-resistant (MDR) CJ in animals and humans, thereby preserving intestinal homeostasis, is relevant. We have created a synbiotic based on the consortium of Lactobacillus crispatus 2029 (LC2029), Ligilactobacillus salivarius 7247 (LS7247), and a mannan-rich prebiotic (Actigen^®). The purpose of this work was to study the in vitro anti-adhesive and antagonistic activities of the created synbiotic against MDR CJ strains, along with its role in preventing intestinal barrier dysfunction, which disrupts intestinal homeostasis. Methods: A complex of microbiological, immunological, and molecular biological methods was used. The ability of the LC2029 and LS7247 consortium to promote intestinal homeostasis in vitro was assessed by the effectiveness of controlling CJ-induced TLR4 activation, secretion of pro-inflammatory cytokines, development of intestinal barrier dysfunction, and production of intestinal alkaline phosphatase (IAP). Results: All MDR CJ strains showed marked adhesion to human Caco-2, pig IPEC-J2, chicken CPCE, and bovine BPCE enterocytes. For the first time, we found that the prebiotic and cell-free culture supernatant (CFS) from the consortium of LC2029 and LS7247 strains exhibit an additive effect in inhibiting the adhesion of MDR strains of CJ to human and animal enterocytes. CFS from the LC2029 and LS7247 consortium increased the permeability of the outer and inner membranes of CJ cells, which led to extracellular leakage of ATP and provided access to the peptidoglycan of the pathogen for the peptidoglycan-degrading bacteriocins nisin and enterolysin A produced by LS7247. The LC2029 and LS7247 consortium showed a bactericidal effect on CJ strains. Co-cultivation of the consortium with CJ strains resulted in a decrease in the viability of the pathogen by 6 log. CFS from the LC2029 and LS7247 consortium prevented the growth of CJ-induced TLR4 mRNA expression in enterocytes. The LC2029 and LS7247 consortium inhibited a CJ-induced increase in IL-8 and TNF-α production in enterocytes, prevented CJ-induced intestinal barrier dysfunction, maintained the transepithelial electrical resistance of the enterocyte monolayers, and prevented an increase in intestinal paracellular permeability and zonulin secretion. CFS from the consortium stimulated IAP mRNA expression in enterocytes. The LC2029 and LS7247 consortium and the prebiotic Actigen represent a new synergistic synbiotic with anti-CJ properties that prevents intestinal barrier dysfunction and preserves intestinal homeostasis. Conclusions: These data highlight the potential of using a synergistic synbiotic as a preventive strategy for creating feed additives and functional nutrition products based on it to combat the prevalence of campylobacteriosis caused by MDR strains in animals and humans. Full article

Damage to intestinal epithelial cells is present in obesity and other diseases because of inflammatory and oxidative processes. This damage compromises the gastrointestinal barrier, killing enterocytes, altering intestinal permeability, and eliciting abnormal immune responses that promote chronic inflammation. Recent evidence shows that spirulina is a potent natural agent with antioxidant and anti-inflammatory properties. Objectives: This study was conducted to evaluate the effect of spirulina aqueous extract (SPAE) on the alterations of the intestinal epithelium induced by lipid micelles (LMs) and/or inflammation induced by lipopolysaccharides (LPSs) in the Caco-2 cell line. Methods: In the current research, we assessed the protective actions of SPAE against cytotoxicity, oxidative stress, inflammation, and epithelial barrier perturbation by using an in vitro model, the intestinal Caco-2 cells, treated with LPSs and/or LMs. We also performed an in silico molecular docking analysis with spirulina’s bioactive compound, phycocyanobilin. Results: Our results showed that SPAE has no cytotoxic effect on Caco-2 cells. On the contrary, it improved cell viability and exhibited anti-inflammatory and antioxidant actions. SPAE also protected against endoplasmic reticulum stress and tight junction proteins, thus improving the epithelial barrier. The in silico study revealed a strong binding affinity of the phycocyanobilin compound with human SOD and NADPH oxidase and a good binding affinity towards COX-2 and iNOS. Conclusions: Taken together, these findings demonstrate the beneficial actions of SPAE on Caco-2 cells, suggesting it may be useful in preserving the epithelial intestinal barrier in human conditions involving oxidative stress and inflammation such as obesity. Full article

Human milk (HM) is rich in bioactive factors promoting postnatal small intestinal development and maturation of the microbiome. HM is also protective against necrotizing enterocolitis (NEC), a devastating inflammatory condition predominantly affecting preterm infants. The HM glycosaminoglycan, hyaluronan (HA), is present at high levels in colostrum and early milk. Our group has demonstrated that HA with a molecular weight of 35 kDa (HA35) promotes maturation of the murine neonatal intestine and protects against two distinct models of NEC. However, the molecular mechanisms underpinning HA35-induced changes in the developing ileum are unclear. CD-1 mouse pups were treated with HA35 or vehicle control daily, from P7 to P14, and we used network and functional analyses of bulk RNA-seq ileal transcriptomes to further characterize molecular mechanisms through which HA35 likely influences intestinal maturation. HA35-treated pups separated well by principal component analysis, and cell deconvolution revealed increases in stromal, Paneth, and mature enterocyte and progenitor cells in HA35-treated pups. Gene set enrichment and pathway analyses demonstrated upregulation in key processes related to antioxidant and growth pathways, such as nuclear factor erythroid 2-related factor-mediated oxidative stress response, hypoxia inducible factor-1 alpha, mechanistic target of rapamycin, and downregulation of apoptotic signaling. Collectively, pro-growth and differentiation signals induced by HA35 may present novel mechanisms by which this HM bioactive factor may protect against NEC. Full article

In the small intestine, sodium (Na) absorption occurs primarily via two apical transporters, Na-hydrogen exchanger 3 (NHE3) and Na-glucose cotransporter 1 (SGLT1). The two primary Na-absorptive pathways were previously shown to compensatorily regulate each other in rabbit and rat intestinal epithelial cells. However, whether NHE3 and SGLT1 regulate one another in normal human enterocytes is unknown, mainly due to a lack of appropriate experimental models. To investigate this, we generated 2D enterocyte monolayers from human jejunal 3D organoids and used small interfering RNAs (siRNAs) to knock down NHE3 or SGLT1. Molecular and uptake studies were performed to determine the effects on NHE3 and SGLT1 expression and activity. Knockdown of NHE3 by siRNA in enterocyte monolayers was verified by qPCR and Western blot analysis and resulted in reduced NHE3 activity. However, in NHE3 siRNA-transfected cells, SGLT1 activity was significantly increased. siRNA knockdown of SGLT1 was confirmed by qPCR and Western blot analysis and resulted in reduced SGLT1 activity. However, in SGLT1 siRNA-transfected cells, NHE3 activity was significantly increased. These results demonstrate for the first time the functionality of siRNA in patient-derived organoid monolayers. Furthermore, they show that the two primary Na absorptive pathways in human enterocytes reciprocally regulate one another. Full article

Background/Objectives: The global emergence of antibiotic-resistant zooanthroponotic Escherichia coli strains, producing extended-spectrum beta-lactamases (ESBL-E) and persisting in the intestines of farm animals, has now led to the development of a pandemic of extra-intestinal infectious diseases in humans. The search for innovative probiotic microorganisms that eliminate ESBL-E from the intestines of humans and animals is relevant. Previously, we received three isolates of bifidobacteria: from milk of a calved cow (BLLT1), feces of a newborn calf (BLLT2) and feces of a three-year-old child who received fresh milk from this calved cow (BLLT3). Our goal was to evaluate the genetic identity of BLLT1, BLLT2, BLLT3 isolates using genomic DNA fingerprinting (GDF), to study the tolerance, adhesion, homeostatic and antibacterial activity of BLLT1 against ESBL-E. Methods: We used a complex of microbiological, molecular biological, and immunological methods, including next generation sequencing (NGS). Results: GDF showed that DNA fragments of BLLT2 and BLLT3 isolates were identical in number and size to DNA fragments of BLLT1. These data show for the first time the possibility of natural horizontal transmission of BLLT1 through with the milk of a calved cow into the intestines of a calf and the intestines of a child. BLLT1 was resistant to gastric and intestinal stresses and exhibited high adhesive activity to calf, pig, chicken, and human enterocytes. This indicates the unique ability of BLLT1 to inhabit the intestines of animals and humans. We are the first to show that BLLT1 has antibacterial activity against ESBL-E strains that persist in humans and animals. BLLT1 produced 145 ± 8 mM of acetic acid, which reduced the pH of the nutrient medium from 6.8 to 5.2. This had an antibacterial effect on ESBL-E. The genome of BLLT1 contains ABC-type carbohydrate transporter gene clusters responsible for the synthesis of acetic acid with its antibacterial activity against ESBL-E. BLLT1 inhibited TLR4 mRNA expression induced by ESBL-E in HT-29 enterocytes, and protected the enterocyte monolayers used in this study as a bio-model of the intestinal barrier. BLLT1 increased intestinal alkaline phosphatase (IAP) as one of the main molecular factors providing intestinal homeostasis. Conclusions: BLLT1 shows promise for the creation of innovative functional nutritional products for humans and feed additives for farm animals that will reduce the spread of ESBL-E strains in the food chain. Full article

Pollution by microplastics and nanoplastics (MNPs) raises concerns, not only regarding their environmental effects, but also their potential impact on human health by internalization via the small intestine. However, the detailed pathways of MNP internalization and their toxicities to the human intestine have not sufficiently been understood, thus, further investigations are required. This work aimed to understand the behavior of MNPs, using in vitro human intestine models, tri-culture models composed of enterocyte Caco-2 cells, goblet-like HT29-MTX-E12 cells, and microfold cells (M cells) induced by the lymphoblast cell line Raji B. Three sizes (50, 100, and 500 nm) of polystyrene (PS) particles were exposed as MNPs on the culture model, and size-dependent translocation of the MNPs and the contributions of each cell were clarified, emphasizing the significance of the tri-culture model. In addition, potential concerns of MNPs were suggested when they invaded the circulatory system of the human body. Full article

The objective of this review was to identify the characteristics and functional roles of milk coproducts from human, bovine, and porcine sources and their impacts on the intestinal microbiota and intestinal immunity of suckling and nursery pigs. Modern pig production weans piglets at 3 to 4 weeks of age, which is earlier than pigs would naturally be weaned outside of artificial rearing. As a result, the immature intestines of suckling and nursery pigs face many challenges associated with intestinal dysbiosis, which can be caused by weaning stress or the colonization of the intestines by enteric pathogens. Milk oligosaccharides are found in sow milk and function as a prebiotic in the intestines of pigs as they cannot be degraded by mammalian enzymes and are thus utilized by intestinal microbial populations. The consumption of milk oligosaccharides during suckling and through the nursery phase can provide benefits to young pigs by encouraging the proliferation of beneficial microbial populations, preventing pathogen adhesion to enterocytes, and through directly modulating immune responses. Therefore, this review aims to summarize the specific functional components of milk oligosaccharides from human, bovine, and porcine sources, and identify potential strategies to utilize milk oligosaccharides to benefit young pigs through the suckling and nursery periods. Full article

PepT1, a proton-coupled oligopeptide transporter, is crucial for intestinal homeostasis. It is mainly expressed in small intestine enterocytes, facilitating the absorption of di/tri-peptides from dietary proteins. In the colon, PepT1 expression is minimal to prevent excessive responses to proinflammatory peptides from the gut microbiota. However, increased colonic PepT1 is linked to chronic inflammatory diseases and colitis-associated cancer. Despite promising results from animal studies on the benefits of extracellular vesicles (EVs) from beneficial gut commensals in treating IBD, applying probiotic EVs as a postbiotic strategy in humans requires a thorough understanding of their mechanisms. Here, we investigate the potential of EVs of the probiotic Nissle 1917 (EcN) and the commensal EcoR12 in preventing altered PepT1 expression under inflammatory conditions, using an interleukin (IL)-1-induced inflammation model in Caco-2 cells. The effects are evaluated by analyzing the expression of PepT1 (mRNA and protein) and miR-193a-3p and miR-92b, which regulate, respectively, PepT1 mRNA translation and degradation. The influence of microbiota EVs on PepT1 expression is also analyzed in the presence of bacterial peptides that are natural substrates of colonic PepT1 to clarify how the regulatory mechanisms function under both physiological and pathological conditions. The main finding is that EcN EVs significantly decreases PepT1 protein via upregulation of miR-193a-3p. Importantly, this regulatory effect is strain-specific and only activates in cells exposed to IL-1β, suggesting that EcN EVs does not control PepT1 expression under basal conditions but can play a pivotal role in response to inflammation as a stressor. By this mechanism, EcN EVs may reduce inflammation in response to microbiota in chronic intestinal disorders by limiting the uptake of bacterial proinflammatory peptides. Full article