Epithelial Cells Of Large Intestine Research Articles

Bacterial lipopolysaccharide inhibits free thiamin uptake along the intestinal tract via interference with membrane expression of thiamin transporters 1 and 2.

This study examined the effect of exposure of small and large intestinal epithelial cells to the bacterial lipopolysaccharide (LPS) on uptake of free form of vitamin B1, i.e., thiamin. The intestinal tract encounters two sources of thiamin: diet and the gut microbiota. Absorption of thiamin in both the small and large intestine occurs via a carrier-mediated process that involves thiamin transporters 1 and 2 (THTR-1 and -2). Complementary in vitro (human duodenal epithelial HuTu-80 cells and human colonic epithelial NCM460 cells), in vivo (mice), and ex vivo (human primary differentiated enteroid and colonoid monolayers) models were used. The results showed that exposure to LPS causes a significant inhibition in carrier-mediated [3H]-thiamin uptake by small and large intestinal epithelia, with no change in the levels of expression of THTR-1 and -2 mRNAs and their total cellular proteins. However, a significant decrease in the fractions of the THTR-1 and -2 proteins that are expressed at the cell membranes of these epithelial cells was observed. These effects of LPS appeared to involve a protein kinase A (PKA) signaling pathway as activating this pathway caused a reversal in the inhibition of thiamin uptake and level of expression of its transporters at the cell membrane. These findings demonstrate that exposure of gut epithelia to LPS (a situation that occurs under different pathological conditions) leads to inhibition in thiamin uptake due to a decrease in level of expression of its transporters at the cell membrane that is likely mediated via a PKA signaling pathway. NEW & NOTEWORTHY This study shows that the exposure of gut epithelial cells to bacterial LPS negatively impact the uptake process of the free form of vitamin B1 (i.e., thiamin). This appears to be mediated via suppression in the level of thiamin transporters 1 and 2 (THTR-1 and -2) expression at the cell membrane and involves a protein kinase A (PKA) signaling pathway.

Effects of Adding Sphingomonas Z392 to Drinking Water on Growth Performance, Intestinal Histological Structure, and Microbial Community of Broiler Chickens.

Probiotics are a prominent alternative to antibiotics in antimicrobial-free broiler farming. To assess the effect of Sphingomonas sp. Z392 (isolated and identified) on broiler growth, 600 one-day-old Kebao broiler chickens were randomly divided into a control group and an experimental group. Each group had three replicates, with 100 broiler chickens being raised in each replicate. Regarding the experimental group of broiler chickens, 4.0 × 105 CFU/mL of Sphingomonas Z392 was added to their drinking water. Then, the changes in broiler body weight, the EPI, intestinal histological structure, and gut microbiota were examined. The results show that the supplementation of the broilers' drinking water with 4 × 105 CFU/mL of Sphingomonas Z392 resulted in an increase in the relative abundance of Lactobacillus, Bacteroides, Lachnospiraceae, Aminobacterium, Oribacterium, Christensenellaceae, Faecalibacterium, Barnesiella, Ruminococcus, Parabacteroides, Phascolarctobacterium, Butyricicoccaceae, and Caproiciproducens, which have been reported to be positively correlated with the improved digestion and absorption of broiler chickens. The relative abundance of Odoribacter, Alistipes, Parabacteroides, and Rikenellaceae increased, and these have been reported to be negatively correlated with the occurrence of intestinal diseases. The relative abundance of Campylobacter, Shigella Castellani, Bilophila, Campylobacter, Clostridia, and Anaerotruncus decreased, and these have been reported to be positively correlated with the occurrence of intestinal diseases. At the same time, the following also increased: the integrity of small intestinal villus morphology; the number of goblet cells in small intestinal epithelial cells; the health of the mitochondria in the cytoplasm of jejunal villous epithelial cells; the number of lysosomes in the cytoplasm of goblet cells in the small intestinal epithelium, ileal villous epithelial cells, and mitochondria in the cytoplasm of large intestinal villous epithelial cells; the VH/CD of the ileum; and digestive, absorption, and defense capabilities. In particular, the final weight increased by 4.33%, and the EPI increased by 10.10%. Therefore, the supplementation of broiler drinking water with Sphingomonas generated better economic benefits from the broiler chickens.

Research Progress on Anti-Inflammatory Effects and Related Mechanisms of Astragalin.

Chronic inflammation is a significant contributor to the development of cancer, cardiovascular disease, diabetes, obesity, autoimmune disease, inflammatory bowel disease, and other illnesses. In the academic field, there is a constant demand for effective methods to alleviate inflammation. Astragalin (AST), a type of flavonoid glycoside that is the primary component in several widely used traditional Chinese anti-inflammatory medications in clinical practice, has garnered attention from numerous experts and scholars. This article focuses on the anti-inflammatory effects of AST and conducts research on relevant literature from 2003 to 2023. The findings indicate that AST demonstrates promising anti-inflammatory potential in various models of inflammatory diseases. Specifically, AST is believed to possess inhibitory effects on inflammation-related factors and protein levels in various in vitro cell models, such as macrophages, microglia, and epithelial cells. In vivo studies have shown that AST effectively alleviates neuroinflammation and brain damage while also exhibiting potential for treating moderate diseases such as depression and stroke; it also demonstrates significant anti-inflammatory effects on both large and small intestinal epithelial cells. Animal experiments have further demonstrated that AST exerts therapeutic effects on colitis mice. Molecular biology studies have revealed that AST regulates complex signaling networks, including NF-κB, MAPK, JAK/STAT pathways, etc. In conclusion, this review will provide insights and references for the development of AST as an anti-inflammatory agent as well as for related drug development.

The UDP-glucose/P2Y14 receptor axis promotes eosinophil-dependent large intestinal inflammation.

Ulcerative colitis (UC) is a chronic disorder of the large intestine with inflammation and ulceration. The incidence and prevalence of UC have been rapidly increasing worldwide, but its etiology remains unknown. In patients with UC, the accumulation of eosinophils in the large intestinal mucosa is associated with increased disease activity. However, the molecular mechanism underlying the promotion of intestinal eosinophilia in patients with UC remains poorly understood. Here, we show that uridine diphosphate (UDP)-glucose mediates the eosinophil-dependent promotion of colonic inflammation via the purinergic receptor P2Y14. The expression of P2RY14 mRNA was upregulated in the large intestinal mucosa of patients with UC. The P2Y14 receptor ligand UDP-glucose was increased in the large intestinal tissue of mice administered dextran sodium sulfate (DSS). In addition, P2ry14 deficiency and P2Y14 receptor blockade mitigated DSS-induced colitis. Among the large intestinal immune cells and epithelial cells, eosinophils highly expressed P2ry14 mRNA. P2ry14-/- mice transplanted with wild-type bone marrow eosinophils developed more severe DSS-induced colitis compared with P2ry14-/- mice that received P2ry14-deficient eosinophils. UDP-glucose prolonged the lifespan of eosinophils and promoted gene transcription in the cells through P2Y14 receptor-mediated activation of ERK1/2 signaling. Thus, the UDP-glucose/P2Y14 receptor axis aggravates large intestinal inflammation by accelerating the accumulation and activation of eosinophils.

Design of 8-mer peptides that block Clostridioides difficile toxin A in intestinal cells

Infections by Clostridioides difficile, a bacterium that targets the large intestine (colon), impact a large number of people worldwide. Bacterial colonization is mediated by two exotoxins: toxins A and B. Short peptides that can be delivered to the gut and inhibit the biocatalytic activity of these toxins represent a promising therapeutic strategy to prevent and treat C. diff. infection. We describe an approach that combines a Peptide Binding Design (PepBD) algorithm, molecular-level simulations, a rapid screening assay to evaluate peptide:toxin binding, a primary human cell-based assay, and surface plasmon resonance (SPR) measurements to develop peptide inhibitors that block Toxin A in colon epithelial cells. One peptide, SA1, is found to block TcdA toxicity in primary-derived human colon (large intestinal) epithelial cells. SA1 binds TcdA with a KD of 56.1 ± 29.8 nM as measured by surface plasmon resonance (SPR).

Intestinal bacteria associated with irritable bowel syndrome and chronic fatigue.

The etiology of irritable bowel syndrome (IBS) is unknown. Abnormal intestinal bacterial profiles and low bacterial diversity appear to play important roles in the pathophysiology of IBS. This narrative review was designed to present recent observations made relating to fecal microbiota transplantation (FMT), which implicate possible roles of 11 intestinal bacteria in the pathophysiology of IBS. The intestinal abundances of nine of these bacteria increased after FMT in patients with IBS, and these increases were inversely correlated with IBS symptoms and fatigue severity. These bacteria were Alistipes spp., Faecalibacterium prausnitzii, Eubacterium biforme, Holdemanella biformis, Prevotella spp., Bacteroides stercoris, Parabacteroides johnsonii, Bacteroides zoogleoformans, and Lactobacillus spp. The intestinal abundances of two bacteria were decreased in patients with IBS after FMT and were correlated with the severity of IBS symptoms and fatigue (Streptococcus thermophilus and Coprobacillus cateniformis). Ten of these bacteria are anaerobic and one (Streptococcus thermophilus) is facultative anaerobic. Several of these bacteria produce short-chain fatty acids, especially butyrate, which is used as an energy source by large intestine epithelial cells. Moreover, it modulates the immune response and hypersensitivity of the large intestine and decreases intestinal cell permeability and intestinal motility. These bacteria could be used as probiotics to improve these conditions. Protein-rich diets could increase the intestinal abundance of Alistipes, and plant-rich diet could increase the intestinal abundance of Prevotella spp., and consequently improve IBS and fatigue.

Non-steroidal anti-inflammatory drugs and biomarkers: A new paradigm in colorectal cancer.

Colorectal cancer is a sporadic, hereditary, or familial based disease in its origin, caused due to diverse set of mutations in large intestinal epithelial cells. Colorectal cancer (CRC) is a common and deadly disease that accounts for the 4th worldwide highly variable malignancy. For the early detection of CRC, the most common predictive biomarker found endogenously are KRAS and ctDNA/cfDNA along with SEPT9 methylated DNA. Early detection and screening for CRC are necessary and multiple methods can be employed to screen and perform early diagnosis of CRC. Colonoscopy, an invasive method is most prevalent for diagnosing CRC or confirming the positive result as compared to other screening methods whereas several non-invasive techniques such as molecular analysis of breath, urine, blood, and stool can also be performed for early detection. Interestingly, widely used medicines known as non-steroidal anti-inflammatory drugs (NSAIDs) to reduce pain and inflammation have reported chemopreventive impact on gastrointestinal malignancies, especially CRC in several epidemiological and preclinical types of research. NSAID acts by inhibiting two cyclooxygenase enzymes, thereby preventing the synthesis of prostaglandins (PGs) and causing NSAID-induced apoptosis and growth inhibition in CRC cells. This review paper majorly focuses on the diversity of natural and synthetic biomarkers and various techniques for the early detection of CRC. An approach toward current advancement in CRC detection techniques and the role of NSAIDs in CRC chemoprevention has been explored systematically. Several prominent governing mechanisms of the anti-cancer effects of NSAIDs and their synergistic effect with statins for an effective chemopreventive measure have also been discussed in this review paper.

The ATP-hydrolyzing ectoenzyme E-NTPD8 attenuates colitis through modulation of P2X4 receptor–dependent metabolism in myeloid cells

Extracellular adenosine triphosphate (ATP) released by mucosal immune cells and by microbiota in the intestinal lumen elicits diverse immune responses that mediate the intestinal homeostasis via P2 purinergic receptors, while overactivation of ATP signaling leads to mucosal immune system disruption, which leads to pathogenesis of intestinal inflammation. In the small intestine, hydrolysis of luminal ATP by ectonucleoside triphosphate diphosphohydrolase (E-NTPD)7 in epithelial cells is essential for control of the number of T helper 17 (Th17) cells. However, the molecular mechanism by which microbiota-derived ATP in the colon is regulated remains poorly understood. Here, we show that E-NTPD8 is highly expressed in large-intestinal epithelial cells and hydrolyzes microbiota-derived luminal ATP. Compared with wild-type mice, Entpd8-/- mice develop more severe dextran sodium sulfate-induced colitis, which can be ameliorated by either the depletion of neutrophils and monocytes by injecting with anti-Gr-1 antibody or the introduction of P2rx4 deficiency into hematopoietic cells. An increased level of luminal ATP in the colon of Entpd8-/- mice promotes glycolysis in neutrophils through P2x4 receptor-dependent Ca2+ influx, which is linked to prolonged survival and elevated reactive oxygen species production in these cells. Thus, E-NTPD8 limits intestinal inflammation by controlling metabolic alteration toward glycolysis via the P2X4 receptor in myeloid cells.

Distinct Age-Specific miRegulome Profiling of Isolated Small and Large Intestinal Epithelial Cells in Mice.

The intestinal epithelium serves as a dynamic barrier to protect the host tissue from exposure to a myriad of inflammatory stimuli in the luminal environment. Intestinal epithelial cells (IECs) encompass differentiated and specialized cell types that are equipped with regulatory genes, which allow for sensing of the luminal environment. Potential inflammatory cues can instruct IECs to undergo a diverse set of phenotypic alterations. Aging is a primary risk factor for a variety of diseases; it is now well-documented that aging itself reduces the barrier function and turnover of the intestinal epithelium, resulting in pathogen translocation and immune priming with increased systemic inflammation. In this study, we aimed to provide an effective epigenetic and regulatory outlook that examines age-associated alterations in the intestines through the profiling of microRNAs (miRNAs) on isolated mouse IECs. Our microarray analysis revealed that with aging, there is dysregulation of distinct clusters of miRNAs that was present to a greater degree in small IECs (22 miRNAs) compared to large IECs (three miRNAs). Further, miRNA–mRNA interaction network and pathway analyses indicated that aging differentially regulates key pathways between small IECs (e.g., toll-like receptor-related cascades) and large IECs (e.g., cell cycle, Notch signaling and small ubiquitin-related modifier pathway). Taken together, current findings suggest novel gene regulation pathways by epithelial miRNAs in aging within the gastrointestinal tissues.

IL-21 Enhances the Development of Colitis-Associated Colon Cancer: Possible Involvement of Activation-Induced Cytidine Deaminase Expression.

Inflammatory bowel diseases are known to be the origin of colitis-associated colon cancer (CAC). We previously reported that dextran sulfate sodium (DSS)-induced colitis is exacerbated in mouse-IL-21-isoform transgenic (Tg) mice. In this study, we assessed the CAC development induced by azoxymethane (AOM) and DSS in our Tg mice. AOM-DSS-induced tumor development was dramatically increased in the Tg mice compared with wild-type mice. IL-21 is known to enhance activation-induced cytidine deaminase (AID) expression in B cells and induce Ab class switching. In contrast, the AID expression in cells other than B cells initiates tumor development in many tissues. Therefore, we investigated whether IL-21 induces the AID expression in the large intestinal epithelial cells (IECs) during CAC development. AID gene and protein expression was increased in the IECs of AOM-DSS- or DSS-treated Tg mice compared with those of wild-type mice. Furthermore, we confirmed IL-21 induced AID gene expression in the purified IECs ex vivo. The present study also showed IL-21R gene expression in unstimulated wild-type mouse IECs, and this gene expression was augmented by TNF-α stimulation. The IL-21R expression and IL-21-induced AID gene activation were further confirmed in the Colon-38 cell line. Taken together, IL-21 may be involved in increasing the risk of CAC by enhancing the AID expression in IECs.

Berberine Improves Mitochondrial Function in Colon Epithelial Cells to Protect L-Cells from Necrosis in Preservation of GLP-1 Secretion

Berberine (BBR) was reported to preserve the colon function in GLP-1 secretion in obese rodents through the butyrate-GPR43 pathway. However, the mechanism remains largely unknown except the pathway. Here, we tested BBR activity in the inhibition of clone epithelial necrosis in diet-induced obese mice. BBR was administrated in DIO mice through dietary supplementation for 8 weeks. Energy balance, insulin sensitivity and intestine mucosal function were monitored. Mitochondria were examined in the large intestine epithelial cells for structure and function to understand cell death. Mitochondrial structure was examined under the transmission electron microscopy and the function was determined using Seahorse XF24. Gut microflora was examined by Miseq sequencing and the fecal short chain fatty acids (SCFAs) were examined using gas chromatograph (GC). An increase in epithelial necrosis was observed in the colon mucosa together with a reduction in GLP-1 secretion and expression in DIO mice. Cristae loss and membrane rupture were observed in the mitochondrial structure of the colon epithelial cells. A disbalance of mitochondrial complexes was observed with an elevation in the complex I activity, and a fall in the complex II and IV activities. The abundance of short chain fatty acids were decreased along with the dysbiosis in the DIO mice. All of those alterations were corrected in DIO mice together with glucose disorder by the BBR treatment. The data suggest that BBR may protect the colon epithelial cells from necrosis in DIO mice to preserve GLP-1 secretion function. The effect was mediated by restoration of mitochondrial function from the disbalance among the complex I, II and IV. The direct and indirect effects of BBR are discussed. Disclosure J. Ye: None. J. Le: None. Y. Sun: None.

Ultra-structural, histo-chemical and histo-enzymic activities in the large intestine of developing Guinea fowl

The ultrastructural, histochemical and histoenzymic activities in the large intestine of the post-hatch Guinea fowl were studied. Guinea fowl of days 1, 7, 15, 30, 90 and 180 were used. The distribution of histochemically active compounds and enzymes were correlated with age. The distributions of enzymes and histochemically reactive substance within the cytoplasm of intestinal epithelial cells have shown that the cytochemical pattern of the mature intestinal absorptive cell is the reflection of both its structural arrangement and its highly specialized function. The brush border showed a mild to moderate reaction with PAS which was resistant to diastase. The apical microvillus coating of glycocalyx, was very thin but it stained strongly with the Alcian Blue thus revealing the presence of acid mucopolysaccharides. The activity of AKPase, ACPase and lipase were observed in the surface epithelium and crypts which increased with the advancement of the age. Columnar cells of the villi and glycocalyx coat of the brush border of all segments of the large intestine showed a mild positive reaction for lipase activity. On ultra-structural study, the epithelial cells of large intestine were tall columnar in shape. They possessed well developed striated border and terminal web. The presence of numerous long micro-villi covered with glycocalyx.

Profile of Turbidity and Glucose Formation from Underutilised Wild, Edible Bean during <i>In-Vitro</i> Gastro Intestinal Digestion and Fermentation

Fermentation takes place throughout the gastrointestinal tract of all animals, but the intensity and products of fermentation depend on number and types microbes, which are generally highest in the large bowel. Large intestinal epithelial cells do not produce digestive enzymes, but contain huge numbers of bacteria which have the enzymes to digest and utilize many substrates. The seeds of beans (Otili, Feregede, Pakala and Oloyin) analyzed in this present study contain indigestible fraction called dietary fiber which helps to maintain functioning of the digestive system. Fermentation of indigestible fraction (IF) of these beans was mimicked through in-vitro method which leads to biochemical changes in the samples. During this experiment, increase in acidity and turbidity was observed. The glucose concentration decreases with some exceptions, such as Pakala fermented by Lactobacillus acidophilus which had the value of 6.260 mmol/L at 6 hr and increased to 6.616 mmol/L after 18 hours fermentation, Otili fermented by various microorganisms which had its turbidity increased by 50%. Lactobacillus acidophilus fermenting Pakala had the highest glucose concentration during the fermentation period. The increase in turbidity could be as a result of increase in microbial flora or production of metabolites, such as glucose. The approach followed here may be used as a predictive model to assess the metabolic implications of food substrates present in the traditional Nigerian orphan beans.

Porcine deltacoronavirus induces apoptosis in swine testicular and LLC porcine kidney cell lines in vitro but not in infected intestinal enterocytes in vivo

We compared the mechanisms of porcine delatacoronavirus (PDCoV) induced death of infected enterocytes in vivo and infected LLC porcine kidney (LLC-PK) and swine testicular (ST) cells in vitro. We conducted histologic analysis and immunofluorescence (IF) staining for the detection of PDCoV antigens, and TUNEL assay in singly or serially cut tissue sections from the small and large intestines of four, 11- to 14-day-old gnotobiotic pigs, inoculated orally with 8.8–11.0log10 genomic equivalents (GE) of US PDCoV strains OH-FD22 or OH-FD100 (n=3), or mock (n=1). Similar comparative assays were done on LLC-PK and ST cells inoculated with the cell-adapted PDCoV strain OH-FD22-P44 (passage 44) in cell culture medium with 2.5–10μg/ml of trypsin and 1% pancreatin, respectively. At post-inoculation days 3–4, infected pigs showed severe watery diarrhea and/or vomiting and mainly, diffuse, severe atrophic enteritis, with mild to moderate cytoplasmic vacuolation of the enteroctyes lining the atrophied villous epithelium. By IF, PDCoV antigens were evident in villous or crypt epithelial cells. No PDCoV antigen-positive, small and large intestinal villous or crypt epithelial cells, of which cytoplasm was also either vacuolated or morphologically normal, showed positive TUNEL staining. In contrast, by double IF and TUNEL staining, most of the TUNEL-positive signals (apoptotic nuclear fragmentation) were found in PDCoV antigen-positive LLC-PK and ST cells that also showed cytopathic effects, such as cell rounding, detachment and clumping in clusters. Secondary annexin V/propidium iodide (PI) staining revealed increased numbers of annexin V- or PI-positive LLC-PK and ST cells at 21h after inoculation, compared to the negative controls. Thus, PDCoV does not induce apoptosis in the infected intestinal enterocytes in vivo, but in two infected cell lines of swine origin, LLC-PK and ST cells.

Naturally Occurring Deletion Mutants of the Pig-Specific, Intestinal Crypt Epithelial Cell Protein CLCA4b without Apparent Phenotype.

The human CLCA4 (chloride channel regulator, calcium-activated) modulates the intestinal phenotype of cystic fibrosis (CF) patients via an as yet unknown pathway. With the generation of new porcine CF models, species-specific differences between human modifiers of CF and their porcine orthologs are considered critical for the translation of experimental data. Specifically, the porcine ortholog to the human CF modulator gene CLCA4 has recently been shown to be duplicated into two separate genes, CLCA4a and CLCA4b. Here, we characterize the duplication product, CLCA4b, in terms of its genomic structure, tissue and cellular expression patterns as well as its in vitro electrophysiological properties. The CLCA4b gene is a pig-specific duplication product of the CLCA4 ancestor and its protein is exclusively expressed in small and large intestinal crypt epithelial cells, a niche specifically occupied by no other porcine CLCA family member. Surprisingly, a unique deleterious mutation of the CLCA4b gene is spread among modern and ancient breeds in the pig population, but this mutation did not result in an apparent phenotype in homozygously affected animals. Electrophysiologically, neither the products of the wild type nor of the mutated CLCA4b genes were able to evoke a calcium-activated anion conductance, a consensus feature of other CLCA proteins. The apparently pig-specific duplication of the CLCA4 gene with unique expression of the CLCA4b protein variant in intestinal crypt epithelial cells where the porcine CFTR is also present raises the question of whether it may modulate the porcine CF phenotype. Moreover, the naturally occurring null variant of CLCA4b will be valuable for the understanding of CLCA protein function and their relevance in modulating the CF phenotype.

Distinct human stem cell populations in small and large intestine.

The intestine is composed of an epithelial layer containing rapidly proliferating cells that mature into two regions, the small and the large intestine. Although previous studies have identified stem cells as the cell-of-origin for intestinal epithelial cells, no studies have directly compared stem cells derived from these anatomically distinct regions. Here, we examine intrinsic differences between primary epithelial cells isolated from human fetal small and large intestine, after in vitro expansion, using the Wnt agonist R-spondin 2. We utilized flow cytometry, fluorescence-activated cell sorting, gene expression analysis and a three-dimensional in vitro differentiation assay to characterize their stem cell properties. We identified stem cell markers that separate subpopulations of colony-forming cells in the small and large intestine and revealed important differences in differentiation, proliferation and disease pathways using gene expression analysis. Single cells from small and large intestine cultures formed organoids that reflect the distinct cellular hierarchy found in vivo and respond differently to identical exogenous cues. Our characterization identified numerous differences between small and large intestine epithelial stem cells suggesting possible connections to intestinal disease.

Toll-like receptor expression in crypt epithelial cells, putative stem cells and intestinal myofibroblasts isolated from controls and patients with inflammatory bowel disease.

The aim of our studies was to investigate the expression of Toll-like receptor (TLR)-2 and TLR-4 (and in some studies TLR-5) in myofibroblasts and small and large intestinal crypt epithelial cells from control patients and those affected by Crohn's disease and ulcerative colitis. Isolated and disaggregated crypt epithelial cells and monolayers of myofibroblasts were used for studies by reverse transcription–polymerase chain reaction (RT–PCR), real-time RT–PCR, flow cytometry, immunocytochemistry and Western blot analysis. Compared to control cells, crypt epithelial cells isolated from active ulcerative colitis and Crohn's disease colonic mucosal samples showed significantly higher expression of TLR-2 and TLR-4 transcripts and protein (on the cell surface). There was also enhanced expression of TLR-4 in crypt cells from ileal Crohn's disease. Expression of TLR-2 and TLR-4 transcripts in crypt epithelial cells isolated from inflamed mucosa of distal ulcerative colitis did not differ significantly from such cells obtained from the normal proximal colon. Crypt epithelial cells with side population characteristics (putative stem cells) also expressed transcripts and protein for TLR-2, TLR-4 and TLR-5. Colonic myofibroblast expression of these TLRs was much weaker than in crypt epithelial cells. In conclusion, enhanced TLR-2 and TLR-4 expression by crypt epithelial cells in active inflammatory bowel disease likely reflects greater ability to respond to microbial products. Results from our studies using mucosal samples from patients with distal ulcerative colitis suggest that the enhanced expression of these TLRs could be constitutive. TLR-2, TLR-4 and TLR-5 expression by stem cells imply ability to respond to distinct bacterial products.

Alteration of Predominant Gastrointestinal Flora and Oxidative Damage of Large Intestine Under Simulated Hypobaric Hypoxia

Hypobaric hypoxia is an immediate and crucial starting mechanism of acute mountain sickness included with some non-specific gastrointestinal (GI) complications. To study the effect of hypoxia on GI microflora and its upshot to this system, male albino rats were exposed to 55 kPa (air pressure ~ 4872.9 m altitude) consecutively 30 days for 8 hours/day. The different indicator group of large intestinal microbial populations were enumerated and correlated with the levels of antioxidant indicators like catalase (CAT), superoxide dismutase (SOD), malondialdehyde (MDA), reduced glutathione (GSH) and oxidized glutathione (GSSG) of large intestinal epithelial cells. In addition, the histological study was performed by haematoxylin eosin (HE), periodic acid schiff staining (PAS) and scanning electron microscopy (SEM). It was observed that the density of total aerobes (104 folds) significantly (p < 0.05) decreased but the population of total anaerobes (209 folds) and Escherichia coli (125 folds) elevated after 30 days of hypoxic stress. The strict anaerobes like Bifidobacterium spp. (3 folds), Bacteroides spp. (134 folds), Lactobacillus spp. (7 folds) and other selected obligate anaerobes like Clostridium perfringens (40 folds), Peptostreptococcus spp. (21 folds) increased in respect to their control population. The growth direction index (GDI) of anaerobic populations was positive and correlated with gas formation aptitude. The activities of CAT and SOD in the large intestinal epithelia decreased significantly (p < 0.05) and GSH/GSSG pool turned into oxidized state with higher MDA (p < 0.05) formation. Histological study revealed the necrotized epithelial layer with higher lymphocytes infiltration in lamina propia accompanied by reduction of acidic mucins secreting goblet cells. From this experiment, it can be hypothesized that high altitude induced hypoxia manipulated the bacterial imprint and damaged the epithelial barrier of the large intestine which may cause systemic infection.

Essential contribution of IRF3 to intestinal homeostasis and microbiota-mediated Tslp gene induction

The large intestinal epithelial cells and immune cells are exposed to a variety of molecules derived from commensal microbiota that can activate innate receptors, such as Toll-like receptors (TLRs) and retinoic acid-inducible gene-I-like receptors (RLRs). Although the activation of these receptors is known to be critical for homeostasis of the large intestine, the underlying gene regulatory mechanisms are not well understood. Here, we show that IFN regulatory factor (IRF)3 is critical for the suppression of dextran sulfate sodium-induced colitis. IRF3-deficient mice exhibited lethal defects in the inflammatory and recovery phases of the colitis, accompanied by marked defects in the gene induction for thymic stromal lymphopoietin (TSLP), a cytokine known to be essential for protection of the large intestine. We further provide evidence that DNA and RNA of the large intestinal contents are critical for Tslp gene induction via IRF3 activation by cytosolic nucleic acid receptors. We also demonstrate that IRF3 indeed activates the gene promoter of Tslp via IRF-binding sequences. This newly identified intestinal gene regulatory mechanism, wherein IRF3 activated by microbiota-derived nucleic acids plays a critical role in intestinal homeostasis, may have clinical implication in colonic inflammatory disorders.

Norovirus GII.4 virus-like particles recognize galactosylceramides in domains of planar supported lipid bilayers.

A sticky situation: Domain-dependent recognition of the glycosphingolipid galactosylceramide by norovirus-like particles (see picture; red/yellow) is shown using supported lipid bilayers (purple) as model membranes. Optimal ligand presentation is found to promote strong binding to GalCer. This presentation can be found at the edges of the glycosphingolipid-enriched domains (green) and binding is repressed in the absence of these domains.