Intestinal Tight Junction Proteins Occludin Research Articles

Effects of vitamin D deficiency on chronic alcoholic liver injury

Vitamin D deficiency (VDD) has been found among alcoholics. However, little is known about the effect of VDD on alcoholic liver disease and the molecular mechanisms remain unclear. The aim of the current study was to evaluate whether vitamin D was deficient among patients with alcoholic fatty liver disease (AFLD) and the effect of VDD on chronic alcoholic liver injury and possible molecular mechanisms in mice. Our results found that lower 25-hydroxyvitamin D [25(OH)D] concentrations in patients with AFLD. And further analysis found that 25(OH)D is a protective factor in patients with AFLD. Mice experiments indicated that VDD can alter the composition of gut microbiota, down-regulate the protein levels of intestinal tight junction protein Occludin and E-cadherin, up-regulate the expression of inflammatory cytokines (tnf-α, il-1β, il-6, il-8, ccl2, il-10) in liver and colon tissue. And further exacerbated the protein levels of p65,P-IκB,P-p65 in alcoholic liver injury mice. In conclusion, VDD aggravates chronic alcoholic liver injury by activating NF-κB signaling pathway.

Increased intestinal permeability and lipopolysaccharide contribute to swainsonine-induced systemic inflammation

Long-term consumption of swainsonine could be poisonous to livestock, including facilitating apoptosis by impairing lysosomal function and inhibiting autophagic degradation, leading to liver inflammation and even death in livestock. However, the mechanism by swainsonine induced systemic inflammatory responses remained unclear, especially the effects of swainsonine on intestinal permeability, lipopolysaccharide (LPS) level and oxidative stress response were unknown. In this study, swainsonine increased intestinal permeability as evidenced by the significant down-regulation of colonic goblet cells, Akkermansia muciniphila and intestinal tight junction protein Occludin, Claudin 1 and ZO-1, and the significant up-regulation of mRNA expression level of the intestinal permeability indicator protein tyrosine phosphatase receptor type H (Ptprh) in the ileum of mice. Simultaneously, the elevated LPS biosynthetic genes in intestinal microbiota and increased intestinal permeability facilitated more bacterial endotoxin LPS to enter the blood. High concentration of free-form LPS induced high levels of proinflammatory cytokines and oxidative stress response, thereby causing the systemic inflammation. These findings provided a new perspective on swainsonine-induced systemic inflammation, suggesting that intestinal permeability and free-form LPS level may be the potential trigger factors.

Kaempferol Ameliorated Alcoholic Hepatitis through Improving Intestinal Barrier Function by Targeting miRNA-155 Signaling

Introduction: The aim of this study was to investigate the effect and mechanism of kaempferol on alcoholic steatohepatitis. Methods: C57BL/6 N mice were utilized to establish Binge-on-Chronic alcohol exposure mice model. Kaempferol was given as the interventional drug to chronic alcohol-fed mice for 6 weeks to assess its effects. In vitro, intestinal epithelial Caco-2 cells were stimulated by alcohol, and miRNA-155 mimics were used to further study the effect of kaempferol to miRNA-155 signaling in intestinal epithelial cells. HE staining and oil red O staining were used to observe the liver and intestinal tissue damage in each group of mice, and ALT, AST, IL-1β, and TNF-α were detected by kits; lipopolysaccharide (LPS) expression was detected by ELISA kit, and the expression of IL-1β and TNF-α was assessed by qRT-PCR; Western blot was utilized to assess the excessive inflammatory response of liver and colon tissue and the related signaling pathway activation. Results: Kaempferol treatment significantly improved pathological changes such as steatosis and vacuolated lesions in liver tissue of the alcohol diet model group, and reduced serum ALT and AST enzyme activities and liver tissue interleukin-1β and tumor necrosis factor-α mRNA expression levels. Kaempferol significantly reduced the expression of miRNA-155 in the intestinal tissue of alcohol-fed mice, significantly increased their cytokine suppressor signaling 1 (SOCS1) protein expression, inhibited the activation of nuclear factor kappa-B and significantly increased the production of the intestinal tight junction proteins occludin and zonula occludens-1. More importantly, kaempferol significantly reduced serum LPS levels in alcoholic steatohepatitis mice. In vitro experiments showed that compared with the control group, kaempferol significantly inhibited the expression level of miRNA-155 in Caco-2 cells under ethanol exposure, decreased the activation of nuclear factor kappa-B, led to an increase in the expression of SOCS1 protein, and increased the production level of occludin protein in Caco-2 cells under the effect of alcohol. In contrast, overexpression of miRNA-155 significantly decreased occludin and SOCS1 protein production and increased nuclear factor kappa-B activation levels in Caco-2 cells, and the administration of kaempferol significantly inhibited this effect. Conclusion: Kaempferol improved the stability of gut barrier function to ameliorate hepatic injury induced by alcohol intake through enhancing occludin protein expression, by targeting miR-155 to inhibit the excessive inflammatory response in the intestine.

Traditional Chinese Medicine formula Dai-Zong-Fang alleviating hepatic steatosis in db/db mice via gut microbiota modulation.

Introduction: Hepatic steatosis is a hepatic pathological change closely associated with metabolic disorders, commonly observed in various metabolic diseases such as metabolic syndrome (MetS), with a high global prevalence. Dai-Zong-Fang (DZF), a traditional Chinese herbal formula, is widely used in clinical treatment for MetS, exhibiting multifaceted effects in reducing obesity and regulating blood glucose and lipids. This study aims to explore the mechanism by which DZF modulates the gut microbiota and reduces hepatic steatosis based on the gut-liver axis. Methods: This study utilized db/db mice as a disease model for drug intervention. Body weight and fasting blood glucose were monitored. Serum lipid and transaminase levels were measured. Insulin tolerance test was conducted to assess insulin sensitivity. Hematoxylin and eosin (HE) staining was employed to observe morphological changes in the liver and intestine. The degree of hepatic steatosis was evaluated through Oil Red O staining and hepatic lipid determination. Changes in gut microbiota were assessed using 16S rRNA gene sequencing. Serum lipopolysaccharide (LPS) levels were measured by ELISA. The expression levels of intestinal tight junction proteins, intestinal lipid absorption-related proteins, and key proteins in hepatic lipid metabolism were examined through Western blot and RT-qPCR. Results: After DZF intervention, there was a decrease in body weight, alleviation of glucose and lipid metabolism disorders, reduction in serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, and mitigation of insulin resistance in mice. DZF significantly modulated the diversity of the gut microbiota, with a notable increase in the abundance of the Bacteroidetes phylum. PICRUSt indicated that DZF influenced various functions in gut microbiota, including carbohydrate and amino acid metabolism. Following DZF intervention, serum LPS levels decreased, intestinal pathological damage was reduced, and the expression of intestinal tight junction protein occludin was increased, while the expression of intestinal lipid absorption-related proteins cluster of differentiation 36 (CD36) and apolipoprotein B48 (ApoB48) were decreased. In the liver, DZF intervention resulted in a reduction in hepatic steatosis and lipid droplets, accompanied by a decrease fatty acid synthase (FASN) and stearoyl-CoA desaturase 1 (SCD1) and fatty acid transport protein 2 (FATP2). Conversely, there was an increase in the expression of the fatty acid oxidation-related enzyme carnitine palmitoyltransferase-1𝛂 (CPT-1𝛂). Conclusion: DZF can regulate the structure and function of the intestinal microbiota in db/db mice. This ameliorates intestinal barrier damage and the detrimental effects of endotoxemia on hepatic metabolism. DZF not only inhibits intestinal lipid absorption but also improves hepatic lipid metabolism from various aspects, including de novo lipogenesis, fatty acid uptake, and fatty acid oxidation. This suggests that DZF may act on the liver and intestine as target organs, exerting its effects by improving the intestinal microbiota and related barrier and lipid absorption functions, ultimately ameliorating hepatic steatosis and enhancing overall glucose and lipid metabolism.

Establishment of Epithelial Inflammatory Injury Model Using Intestinal Organoid Cultures.

Intestinal epithelial dysfunction is critical in the development of inflammatory bowel disease (IBD). However, most cellular experiments related to epithelial barrier studies in IBD have been based on tumor cell line that lack a variety of intestinal epithelial cell types. Thus, intestinal organoids can present the three-dimensional structure and better simulate the physiological structure and function of the intestinal epithelium in vitro. Here, the crypts were isolated from the small intestine of mice; with the participation of major cytokines (EGF, Noggin, and R-Spondin 1 included), the intestinal organoids were established at a density of 100 crypts per well, containing intestinal stem cells (ISC), Paneth cells, goblet cells, and intestinal endocrine cells. We found that tumor necrosis factor-alpha (TNF-α) could induce the inflammatory response of intestinal organoids, and a dose of 10 ng/mL could maintain stable passaging of organoids for dynamic observation. After stimulation with TNF-α, the intestinal organoid cultures showed lower expression of the cell proliferation-related protein identified by monoclonal antibody Ki 67 (Ki67), the ISC marker leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5), and the intestinal tight junction proteins occludin (Ocln) and claudin-1 (Cldn1) while higher expression of the inflammatory cytokine interleukin- (IL-) 15 and the chemokines C-X-C motif ligand 2 (Cxcl2) and Cxcl10 significantly. In this study, we successfully established an epithelial inflammatory injury model of intestinal organoids, which provides an effective in vitro model for studying the pathogenesis and treatment of IBD.

Butyrate ameliorates chronic alcoholic central nervous damage by suppressing microglia-mediated neuroinflammation and modulating the microbiome-gut-brain axis

BackgroundAlcohol abuse triggers neuroinflammation, leading to neuronal damage and further memory and cognitive impairment. Few satisfactory advances have been made in the management of alcoholic central nervous impairment. Therefore, novel and more practical treatment options are urgently needed. Butyrate, a crucial metabolite of short-chain fatty acids (SCFAs), has been increasingly demonstrated to protect against numerous metabolic diseases. However, the impact of butyrate on chronic alcohol consumption-induced central nervous system (CNS) lesions remains unknown. MethodsIn this study, we assessed the possible effects and underlying mechanisms of butyrate on the attenuation of alcohol-induced CNS injury in mice. Firstly, sixty female C57BL/6 J mice were randomly divided into 4 groups: pair-fed (PF) group (PF/CON), alcohol-fed (AF) group (AF/CON), PF with sodium butyrate (NaB) group (PF/NaB) and AF with NaB group (AF/NaB). Each group was fed a modified Lieber-DeCarli liquid diet with or without alcohol. After six weeks of feeding, the mice were euthanized and the associated indicators were investigated. ResultsAs indicated by the behavioral tests and brain morphology, dietary NaB administration significantly ameliorated aberrant behaviors, including locomotor hypoactivity, anxiety disorder, depressive behavior, impaired learning, spatial recognition memory, and effectively reduced chronic alcoholic central nervous system damage. To further understand the underlying mechanisms, microglia-mediated inflammation and the associated M1/M2 polarization were measured separately. Firstly, pro-inflammatory TNF-α, IL-1β, and IL-6 in brain and peripheral blood circulation were decreased, but IL-10 were increased in the AF/NaB group compared with the AF/CON group. Consistently, the abnormal proportions of activated and resting microglial cells in the hippocampus and cortex regions after excessive alcohol consumption were significantly reduced with NaB treatment. Moreover, the rectification of microglia polarization (M1/M2) imbalance was found after NaB administration via binding GPR109A, up-regulating the expression of PPAR-γ and down-regulating TLR4/NF-κB activation. In addition to the direct suppression of neuroinflammation, intriguingly, dietary NaB intervention remarkably increased the levels of intestinal tight junction protein occludin and gut morphological barrier, attenuated the levels of serum lipopolysaccharide (LPS) and dysbiosis of gut microbiota, suggesting that NaB supplementation effectively improved the integrity and permeability of gut microecology. Finally, the neurotransmitters including differential Tryptophan (Trp) and Kynurenine (Kyn) were found with dietary NaB administration, which showed significantly altered and closely correlated with the gut microbiota composition, demonstrating the complex interactions in the microbiome-gut-brain axis involved in the efficacy of dietary NaB therapy for alcoholic CNS lesions. ConclusionDietary microbial metabolite butyrate supplementation ameliorates chronic alcoholic central nervous damage and improves related memory and cognitive functions through suppressing microglia-mediated neuroinflammation by GPR109A/PPAR-γ/TLR4-NF-κB signaling pathway and modulating microbiota−gut−brain axis.

Huang Qin Decoction inhibits the initiation of experimental colitis associated carcinogenesis by controlling the PAD4 dependent NETs

BackgroundColorectal cancer is associated with ulcerative colitis (UC). The infiltration of neutrophils is the main cause of DNA damage produced by inflammation in the intestinal epithelium. Under the action of peptidyl arginine deaminase 4 (PAD4), neutrophils dissociate chromatin and form neutrophil extracellular traps (NETs), which can aggravate tissue inflammation and encourage tumor development. Although Huang Qin Decoction (HQD) was found to be useful in treating UC and was used to gradually prevent and treat digestive tract cancers, the underlying reasons were unclear. MethodsTo demonstrate HQD could inhibits the initiation of colitis associated carcinogenesis by controlling NETs related inflammation, we first performed an AOM/DSS-generated colitis-associated carcinogenesis model to assess the efficacy of HQD in reducing neutrophil infiltration and anti-tumor activity. Then, using network pharmacology research, we investigated the potential mechanisms underlying those medicinal effects, as demonstrated by the detection of NETs aggregation and PAD4 expression changes in the colon. ResultsHQD substantially reduced the number of colon cancers and the expression of Ki67, restored the level of intestinal tight junction protein occludin and ZO-1, and relieved the intestinal inflammation caused by TNF-α, IL-1β. At the same time, it inhibited neutrophil infiltration in the colon and improved the immunosurveillance of CD8+T cells. The potential mechanisms of HQD intervention against UC and UC with neoplasia (UCN) were studied using network pharmacology, and 156 conjunct genes as well as numerous inflammation-related pathways were identified. Protein-protein interaction (PPI) analysis indicated that HQD inhibition of intestinal tumors might be related to the deactivation of PAD4, which was verified by the down-regulation of NETs, MPO-DNA complex levels, and PAD4 expression after HQD treatment. ConclusionHuang Qin Decoction inhibits the initiation of colitis associated carcinogenesis by controlling PAD4-dependent neutrophil extracellular traps.

Modulating effects of a functional food containing Dendrobium officinale on immune response and gut microbiota in mice treated with cyclophosphamide

National Herbal Drink (NHD) is a functional food approved by the State Administration for Market Regulation for immune enhancement. In this study, the effects of NHD on Cyclophosphamide (CTX)-induced immune regulation and gut microbiota dysregulation in mice were investigated to elucidate whether the attenuation of immunosuppression was related to the regulation of the gut microbiota by NHD. Our study showed that CTX induced immune organ damage in immunosuppressed mice, with reduced levels of serum immunoglobulins IgG, IgM, and complement C3 and disruption of bone marrow hematopoiesis, accompanied by changes in liver toxicity and shortened colonic length. However, all these changes were reversed to varying degrees by NHD intervention. Delayed-type hypersensitivity (DTH) and serum hemolysins in mice also indicated that NHD could effectively regulate cellular and humoral immunity in the body. In addition, it is noteworthy that NHD can also improve intestinal pathology, elevate the expression of intestinal tight junction proteins Occludin and Claudin-1 to restore intestinal barrier function. Furthermore, α-diversity and β-diversity analysis showed that NHD has a positive regulatory effect on the gut microbiota. At the phylum level, NHD could regulate the abundance of three major phyla, namely, Bacteroidetes, Firmicutes, and Actinobacteriota. At the species level, the abundance of the beneficial genus uncultured_bacterium_g__norank_f__ Muribaculaceae increased and the abundance of the harmful genus unclassified_f__Lachnospiraceae decreased after the NHD intervention. These results provide evidence that NHD modulates the immune response in cyclophosphamide-induced immunocompromised mice by restoring intestinal barrier function and regulating gut microbiota disorders.

Lactiplantibacillus plantarum J-15 reduced calcium oxalate kidney stones by regulating intestinal microbiota, metabolism, and inflammation in rats.

The prevention role of Lactiplantibacillus plantarum against the formation of kidney stones has been increasingly recognized; its mechanism, however, has mainly been focused on inhibiting the inflammation in the colon in the gastrointestinal (GI) system, and the intestinal metabolites from microflora have not been revealed fully with regarding to the stone formation. In this study, we investigated the effect of L. plantarum J-15 on kidney stone formation in renal calcium oxalate (CaOx) rats induced by ethylene glycol and monitored the changes of intestinal microflora and their metabolites detected by 16S rRNA sequencing and widely targeted analysis, followed by the evaluation of the intestinal barrier function and inflammation levels in the colon, blood and kidney. The results showed that L. plantarum J-15 effectively reduced renal crystallization and urinary oxalic acid. Ten microbial genera, including anti-inflammatory and SCFAs-related Faecalibaculum, were enriched in the J-15 treatment group. There are 136metabolites from 11 categories significantly different in the J-15supplementation group compared with CaOx model rats, most of which were enriched in the amino acid metabolic and secondary bile acid pathways. The expression of intestinal tight junction protein Occludin and the concentration of pro-inflammatory cytokines and prostaglandin were decreased in the intestine, which further reduced the translocated lipopolysaccharide and inflammation levels in the blood upon J-15 treatment. Thus, the inflammation and injury in the kidney might be alleviated by downregulating TLR4/NF-κB/COX-2signaling pathway. It suggested that L. plantarum J-15might reduce kidney stone formation by restoring intestinal microflora and metabolic disorder, protecting intestinal barrier function, and alleviating inflammation. This finding provides new insights into the therapies for renal stones.

Mechanism and therapeutic effect of umbilical cord mesenchymal stem cells in inflammatory bowel disease

Inflammatory bowel disease (IBD) is a persistent and chronic disease that is characterized by destructive gastrointestinal (GI) inflammation. Researchers are trying to identify and develop new and more effective treatments with no side effects. Acute and chronic mouse models of IBD were established using dextran sulfate sodium (DSS) solution. To evaluate the efficacy and mechanism, umbilical cord mesenchymal stem cells (UCMSCs) were obtained from Kunming (KM) mice and humans. In the chronic IBD study, the survival rates of the normal control, model, mouse UCMSC (mUCMSC) and human UCMSC (hUCMSC) groups were 100%, 40%, 86.7%, and 100%, respectively. The histopathological scores of the normal control, intraperitoneal injection, intravenous treatment, and model groups were 0.5 ± 0.30, 5.9 ± 1.10, 8.7 ± 1.39, and 8.8 ± 1.33 (p = 0.021). UCMSCs promoted the expression of the intestinal tight junction protein occludin, downregulated the protein expression of the autophagy marker LC3A/B in colon tissue, and upregulated the expression of VEGF-A and VEGFR-1 at the injured site. This study provides an experimental model for elucidating the therapeutic effects of UCMSCs in IBD. We provide a theoretical basis and method for the clinical treatment of IBD using UCMSCs.

Paneth cell ablation increases the small intestinal injury during acute necrotizing pancreatitis in rats.

The present work aimed to investigate the role of Paneth cells in small intestinal injury during acute necrotizing pancreatitis (ANP) using rat models established by injection of dithizone, a metal chelator of zinc with the ability to selectively ablate Paneth cells. Sprague-Dawley rats were randomly divided into four groups: Sham-operated group, ANP group (3.5% sodium taurocholate solution, 1 ml/kg body weight), dithizone group (100 mg/kg of body weight) and ANP + dithizone group (sodium taurocholate solution was administered 6 h after dithizone injection). Each group was further divided into five subgroups (6, 12, 24, 36 and 48 h) based on the time period between induction of the model and sample collection. The present results suggested the number of Paneth cells was gradually decreased in the ANP group in a time-dependent manner. Most of the Paneth cells were ablated in the ANP + dithizone group at 6 h, but a subset of Paneth cells recovered after 24–48 h. Compared with the ANP group, combination of dithizone and ANP significantly induced more severe histopathological injuries in the pancreas and distal ileum, with higher Schmidt and Chiu's scores, respectively. Additionally, increased expression levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-17A were detected in the ileum, causing an increase in intestinal permeability, as assessed by a decrease in the expression level of the intestinal tight junction protein occludin and high plasma levels of diamine oxidase and D-lactate. The increase in intestinal permeability led to the translocation of bacteria to the bloodstream, triggering systemic inflammation, as assessed by the increased plasma levels of TNF-α, IL-1β and IL-17A, reducing the survival rates of rats, which was 66.7% and 83.3% in the ANP + dithizone and the ANP group, respectively. The increase in intestinal endoplasmic reticulum stress, as assessed by high expression levels of binding-immunoglobulin protein and activating transcription factor 6, may be one mechanism associated with Paneth cells loss and intestinal barrier impairment during ANP. Collectively, the present study suggested that the absence of Paneth cells may be an important factor involved in intestinal injury, promoting the progression of ANP.

Volatile Oil of Amomum villosum Inhibits Nonalcoholic Fatty Liver Disease via the Gut-Liver Axis.

Background The dried mature fruit of Amomum villosum has been historically used in China as food and in the auxiliary treatment of digestive system disorders. Numerous studies have shown that gastrointestinal function is closely related to the development of nonalcoholic fatty liver disease via the “gut-liver” axis. Objective The present study aimed to explore whether the mechanism underlying the regulation of lipid accumulation in nonalcoholic fatty liver disease (NAFLD) may affect related disorders using the active ingredients in A. villosum. Design Male Sprague-Dawley rats on a high-fat diet (HFD) to induce NAFLD were administered water extract of A. villosum (WEAV), volatile oil of A. villosum (VOAV), or bornyl acetate. After treatment, serum and liver total cholesterol (TC), triglyceride (TG), free fatty acid (FFA), aspartate aminotransferase (AST), alanine aminotransferase (ALT), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) levels were measured. The regulatory role of A. villosum in the microecology of the intestines was assessed using the V4 region of the 16S rDNA sequencing. The expression of the intestinal tight junction proteins occludin and ZO-1 was also measured. The influence of A. villosum on TLR4-mediated chronic low-grade inflammation was evaluated based on the concentrations of key proteins of the TLR4/NF-кB signaling pathway. Results. A. villosum effectively inhibited endogenous lipid synthesis, reduced TG, TC, and FFA accumulation, regulated the expression of LDL-C, and decreased lipid accumulation in liver tissues. VOAV effectively regulated the intestinal microflora, improved chronic low-grade inflammation by promoting ZO-1 and occludin protein expressions, and inhibited the TLR4/NF-кB signaling pathway. Conclusion The present study provides scientific basis for the potential application of A. villosum in NAFLD prevention and treatment. Additional chemical constituents other than bornyl acetate also contributed to the preventive effects of A. villosum on NAFLD.

MicroRNA-21 is upregulated during intestinal barrier dysfunction induced by ischemia reperfusion

This study aimed to investigate the expression of miRNA-21 during intestinal barrier dysfunction induced by intestinal ischemia reperfusion. Forty SPF SD rats were divided into 5 groups randomly. Intestinal ischemia-reperfusion injury (IRI) was induced by mesenteric artery occlusion for 1 h and reperfusion for 1 h, and the rats were sacrificed at 1, 3, 6 and 12 h after reperfusion. Fresh intestine tissues were immediately isolated for the measurement of transepithelial electrical resistance (TER). The levels of cytokines, ICAM-1, DAO, iFABP and MPO in serum were determined by ELISA. Intestinal tight junction proteins occludin and claudin-1 were detected by immunofluorescence analysis and Western blot analysis. miR-21 expression in intestinal tissues was measured by RT-PCR. Compared with sham group, the levels of pro-inflammatory cytokines TNF-α and IL-6 and ICAM-1, DAO, iFABP and MPO increased while IL-10 level decreased in intestinal ischemia-reperfusion group. In addition, the levels of intestinal tight junction proteins occludin and claudin-1 decreased while miR-21 level increased in intestinal ischemia-reperfusion group, compared with sham group. In conclusion, miR-21 expression is upregulated during intestinal barrier dysfunction induced by IRI. miR-21 may play an important role in the regulation of intestinal barrier function.

Expression of NF-κB and Occludin in Intestinal Mucosal Barrier During Severe Acute Pancreatitis and Its Clinical Significance

Objective to investigate the expression of NF-κB and occludin in intestinal mucosal barrier during severe acute pancreatitis (SAP) and its clinical significance. Method Twenty-four Wistar rats were randomly divided into a normal control group, an SAP group, and a PDTC group. The rats were sacrificed at 24h after modeling, and general appearance of the abdominal cavity, pathological changes of the intestinal mucosa, changes in intestinal mucosal permeability were observed, serum inflammatory factor IL-1 was measured, and expression of tight junction protein occludin in the intestinal epithelial cells were determined by immunohistochemistry. Results Rats of the SAP group showed obvious inflammatory reaction in the abdominal cavity, manifested by enlargement of intestinal tract, intestinal edema, and increased permeability of intestinal barrier, whereas NF-κB inhibitor alleviated intestinal damage and intestinal mucosa edema, upregulated intestinal epithelial tight junction protein occludin, decreased intestinal permeability, and lowered the level of inflammatory factors. Conclusion Impairment of intestinal barrier function during SAP may be associated with increased NF-κB and decreased intestinal tight junction protein occludin; inhibiting NF-kB may alleviate intestinal mucosal barrier dysfunction during SAP.

Targeting α-7 Nicotinic Acetylcholine Receptor in the Enteric Nervous System: A Cholinergic Agonist Prevents Gut Barrier Failure after Severe Burn Injury

We have previously shown that vagal nerve stimulation prevents intestinal barrier loss in a model of severe burn injury in which injury was associated with decreased expression and altered localization of intestinal tight junction proteins. α-7 Nicotinic acetylcholine receptor (α-7 nAchR) has been shown to be necessary for the vagus nerve to modulate the systemic inflammatory response, but the role of α-7 nAchR in mediating gut protection remained unknown. We hypothesized that α-7 nAchR would be present in the gastrointestinal tract and that treatment with a pharmacological agonist of α-7 nAchR would protect against burn-induced gut barrier injury. The effects of a pharmacological cholinergic agonist on gut barrier integrity were studied using an intraperitoneal injection of nicotine 30 minutes after injury. Intestinal barrier integrity was examined by measuring permeability to 4-kDa fluorescein isothiocyanate-dextran and by examining changes in expression and localization of the intestinal tight junction proteins occludin and ZO-1. Nicotine injection after injury prevented burn-induced intestinal permeability and limited histological gut injury. Treatment with nicotine prevented decreased expression and altered localization of occludin and ZO-1, as seen in animals undergoing burn alone. Defining the interactions among the vagus nerve, the enteric nervous system, and the intestinal epithelium may lead to development of targeted therapeutics aimed at reducing gut barrier failure and intestinal inflammation after severe injury.

Protective effect of galactooligosaccharide-supplemented enteral nutrition on intestinal barrier function in rats with severe acute pancreatitis

Intestinal barrier dysfunction may play an important role in the secondary infectious complications associated with severe acute pancreatitis (SAP). We aimed to investigate the protective effect of enteral nutrition supplemented with prebiotics, namely, galactooligosaccharides (GOS), on intestinal barrier function in SAP rats. We randomly divided 60 Sprague-Dawley (SD) rats into 6 groups: sham operation 4-d and 7-d groups, SAP with standard enteral nutrition (EN) 4-d and 7-d groups, and SAP with GOS-supplemented EN (PRE-EN) 4-d and 7-d groups (n = 10 in each group). The intestinal barrier function was assessed on the basis of several aspects, including the number of colonic bifidobacteria and lactobacilli, sIgA concentration in intestinal mucus, extent of apoptosis in intestinal epithelial cells, and the protein expression and mRNA of the intestinal tight junction protein occludin. The results for groups at the same time points were compared with each other. We observed that the intestinal barrier function was impaired after the establishment of the SAP model, in comparison with the sham control group. In the PRE-EN groups, the number of fecal bifidobacteria, sIgA level in intestinal mucus, intestinal occludin mRNA level at both day 4 and day 7, and extent of intestinal epithelial apoptosis at day 7 were significantly higher than the corresponding values in the standard EN groups (P < 0.05). Supplementation of the prebiotic GOS in EN can significantly improve intestinal barrier function in SAP rats.

Phosphodiesterase inhibition attenuates alterations to the tight junction proteins occludin and ZO-1 in immunostimulated Caco-2 intestinal monolayers

Aims Under normal conditions, the intestinal mucosa acts as a local barrier to prevent the influx of luminal contents. The intestinal epithelial tight junction is comprised of several membrane associated proteins, including zonula occludens-1 (ZO-1) and occludin. Disruption of this barrier can lead to the production of pro-inflammatory mediators and ultimately multiple organ failure. We have previously shown that Pentoxifylline (PTX) decreases histologic gut injury and pro-inflammatory mediator synthesis. We hypothesize that PTX prevents the breakdown of ZO-1 and occludin in an in vitro model of immunostimulated intestinal cell monolayers. Main methods Caco-2 human enterocytes were grown as confluent monolayers and incubated under control conditions, or with PTX (2 mM), Cytomix (TNF-α, IFN-γ, IL-1), or Cytomix + PTX for 24 h. Occludin and ZO-1 protein levels were analyzed by Western blot. Confocal microscopy was used to assess the cytoplasmic localization of ZO-1 and occludin. Key findings Cytomix stimulation of Caco-2 cells resulted in a 50% decrease in both occludin and ZO-1 protein. Treatment with Cytomix + PTX restored both occludin and ZO-1 protein to control levels. Confocal microscopy images show that Cytomix caused an irregular, undulating appearance of ZO-1 and occludin at the cell junctions. Treatment with PTX prevented the Cytomix-induced changes in ZO-1 and occludin localization. Significance Treatment with PTX decreases the pro-inflammatory cytokine induced changes in the intestinal tight junction proteins occludin and ZO-1. Pentoxifylline may be a useful adjunct in the treatment of sepsis and shock by attenuating intestinal barrier breakdown.