Disruption Of Intestinal Tight Junctions Research Articles

The Predictive Value of Serum DAO, HDC, and MMP8 for the Gastrointestinal Injury in the Early Stage of Acute Pancreatitis in an Animal Model and a Clinical Study.

This study was aimed at exploring the use of the acute gastrointestinal injury (AGI) grade and sensitive biomarkers to investigate gastrointestinal (GI) injury in early stage of acute pancreatitis (AP). The AGI grade was used to evaluate intestinal function. Any GI injury above grade I (grades II-IV) was considered as severe. An AP rat model was created by retrograde injection of 4% sodium taurocholate. The pancreatic and intestinal histopathology scores were calculated by hematoxylin-eosin staining. Human and rat sera were assessed using ELISA. Tight junction (TJ) proteins were detected by Western blotting. In clinical study, the GI injury rate in mild acute pancreatitis (MAP), moderate severe acute pancreatitis (MSAP), and severe acute pancreatitis (SAP) groups was 26.8%, 78.4%, and 94.8%, respectively (P < 0.05). Diamine oxidase (DAO), histidine decarboxylase (HDC), and matrix metalloproteinase 8 (MMP8) serum levels were higher in AP patients than in healthy people (P < 0.05). Patients with GI injury had higher serum levels of DAO, HDC, and MMP8 than those without GI injury (P < 0.05). In animal experiments, the serum levels of DAO, HDC, and MMP8 were higher in the AP group than in normal and sham-operated (SO) groups (P < 0.05). The expressions of tricellulin, claudin-1, ZO-1, and occludin were significantly lower in the AP group than in normal and SO groups (P < 0.05). The serum levels of DAO, HDC, and MMP8 are novel biomarkers of GI injury in the early stage of AP; their elevation indicates the development of GI injury in AP. The intestinal TJ disruption may be a primary mechanism of GI injury and requires more in-depth research.

Microfluidic Model of Necrotizing Enterocolitis Incorporating Human Neonatal Intestinal Enteroids and a Dysbiotic Microbiome.

Necrotizing enterocolitis (NEC) is a severe and potentially fatal intestinal disease that has been difficult to study due to its complex pathogenesis, which remains incompletely understood. The pathophysiology of NEC includes disruption of intestinal tight junctions, increased gut barrier permeability, epithelial cell death, microbial dysbiosis, and dysregulated inflammation. Traditional tools to study NEC include animal models, cell lines, and human or mouse intestinal organoids. While studies using those model systems have improved the field's understanding of disease pathophysiology, their ability to recapitulate the complexity of human NEC is limited. An improved in vitro model of NEC using microfluidic technology, named NEC-on-a-chip, has now been developed. The NEC-on-a-chip model consists of a microfluidic device seeded with intestinal enteroids derived from a preterm neonate, co-cultured with human endothelial cells and the microbiome from an infant with severe NEC. This model is a valuable tool for mechanistic studies into the pathophysiology of NEC and a new resource for drug discovery testing for neonatal intestinal diseases. In this manuscript, a detailed description of the NEC-on-a-chip model will be provided.

Matrix metalloproteinase-7 and claudin-7 as novel identified therapeutic targets for restoration of intestinal epithelial barrier in inflammatory bowel diseases

ABSTRACT Intestinal tight junction disruption and mucosal immune dysregulation contribute to pathogenesis and progression of inflammatory bowel diseases (IBD). A proteolytic enzyme matrix metalloproteinase 7 (MMP-7), which is highly expressed in intestinal tissue, is implicated to IBD and other immune overactivation-associated diseases. In the issue of the Frontiers in Immunology, Ying Xiao and colleagues demonstrate that MMP-7-mediated claudin-7 degradation promotes IBD pathogenesis and disease progression. Therefore, inhibition of MMP-7 enzymatic activity can be a therapeutic strategy for the treatment of IBD.

Cholecystokinin and glucagon-like peptide-1 analogues regulate intestinal tight junction, inflammation, dopaminergic neurons and α-synuclein accumulation in the colon of two Parkinson's disease mouse models

Parkinson's disease (PD) is the second most common neurodegenerative disease, and no treatment is available to stop its progression. Studies have shown that the colonic pathology of PD precedes that of the brain. The 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model and the human A53T α-synuclein (α-syn) transgenic PD mouse model show colonic pathology and intestinal dopaminergic neuronal damage, which is comparable to the intestinal pathology of PD. Cholecystokinin (CCK) and glucagon-like peptide-1 (GLP-1), which are brain-gut peptides, have neurotrophic and anti-inflammatory properties. Two GLP-1R agonists have already shown robust effects in phase II trials in PD patients. However, whether they have beneficial effects on colonic pathology in PD remains unclear. In this study, MPTP-treated mice and human A53T α-syn transgenic mice were intraperitoneally injected with a CCK analogue or Liraglutide, a GLP-1 analogue, once a day for 5 weeks. Levels of colonic epithelial tight junction proteins including occludin and zonula occludens-1 (ZO-1), inflammatory biomarkers including inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-α), brain-derived neurotrophic factor (BDNF), tyrosine hydroxylase (TH) and α-syn were analyzed. The results show that the CCK analogue and Liraglutide both restored the disruption of intestinal tight junction, reduced colonic inflammation, inhibited colonic dopaminergic neurons reduction and the accumulation of α-syn oligomers in the colon of both PD mice models. This study suggested that CCK or GLP-1 analogues could be beneficial to the improvement of leaky gut barrier, inflammation, dopaminergic neuron impairment and accumulation of α-syn in the colon of PD patients.

Protective Effects of the Novel Phytonutrient Spectra7 Against Intestinal Tight Junction Disruption: Composition Matters

The prevalence of intestinal inflammatory diseases is increasing, and pharmacologic agents for intervention are currently limited. Preserving epithelial tight junction (TJ) integrity and preventing underlying immune cell activation by intestinal bacteria are key targets for abrogating the perpetual inflammatory cycle that plagues these diseases. Phytonutrients have shown promise for their ability to reduce cellular inflammation, but the extent of their efficacy in an intestinal model of inflammation is not well understood. Here we hypothesized that Spectra7 (S7), a novel phytonutrient derived from extracts rich in curcuminoids and catechins, would reduce immune cell inflammation and preserve TJ integrity in an in vitro co‐culture model of intestinal inflammation. We further hypothesized that a curcumin containing formulation (S7‐C) would be more effective at preserving TJ integrity than that of its metabolite, tetrahydrocurcumin (S7‐THC).An in vitro intestinal co‐culture model was established by seeding Caco‐2 epithelial cells on semi‐permeable transwell inserts 21 days prior to the addition of RAW264.7 macrophages in the basolateral chamber. Macrophages were next stimulated with 10 ng/ml lipopolysaccharide (LPS) to induce inflammation, and subsequent TJ disruption in the co‐cultured Caco‐2 cells was assessed by transepithelial electrical resistance (TEER) using epithelial ohmmeter chopstick electrodes. We found that administration of S7‐THC containing 1–5 μM THC produced dose dependent mitigation of LPS‐induced decreases in TEER and approached the efficacy of the pharmacologic agent budesonide. However, to our surprise, S7‐C at 5 μM curcumin was unable to preserve TEER, suggesting that the specific combination of phytonutrients is important for preventing inflammation‐induced TJ disruption. We also found that, though apical application (Caco‐2 only) of budesonide was sufficient for preserving TEER in our model, S7‐THC required both apical (Caco‐2) and basolateral (RAW264.7) treatment, suggesting that reducing macrophage inflammation is important for limiting epithelial TJ disruption in this context. Interestingly, S7‐C was more effective than S7‐THC or budesonide at reducing inflammatory basolateral nitric oxide (NO) production as determined by the Griess assay. This suggests that, though S7‐C more effectively reduces this aspect of inflammation, another inflammatory mediator is responsible for conveying TJ disruption and is governed differentially by S7‐THC. Further support of this comes from our finding that S7‐THC, but not S7‐C, ameliorated the LPS‐induced increase in myosin light chain kinase (MLCK) expression in Caco‐2 cells as determined by Western blot. Together, these findings suggest that phytonutrients such as S7‐THC have prophylactic potential in the preservation of TJ integrity, and the specific composition of these phytonutrients matters.Support or Funding InformationOlivet Nazarene University Honors Program, Pence‐Boyce Undergraduate Summer Research Experience, and the Hippenhammer Faculty Grant

Occludin endocytosis is involved in the disruption of the intestinal epithelial barrier in a mouse model of alcoholic steatohepatitis.

We aimed to investigate the involvement of the endocytosis of occludin, a key component of tight junction (TJ), in the ethanol-induced disassembly of TJ in a model of alcoholic steatohepatitis. Wild-type mice were fed an ethanol-containing or isocaloric liquid diet for 8 weeks and then assessed for liver injury (histopathology and measurement of serum enzymes), gut permeability (in vivo lactulose/mannitol and ex vivo dye leakage assays), intestinal epithelium ultrastructure (transmission electron microscopy), and intestinal occludin localization (immunofluorescence microscopy). The human intestinal epithelial cell line Caco-2 was also analyzed in vitro for the effects of ethanol on the barrier function (transepithelial electrical resistance), occludin localization (immunofluorescence microscopy and Western blotting), and endocytosis pathways (double-labeling immunofluorescence microscopy with selective pathway inhibitors). The ethanol-fed mice developed steatohepatitis and displayed intestinal barrier dysfunction, the disruption of intestinal TJ, and enhanced intestinal endocytosis of occluding compared with the control mice. In the Caco-2 monolayers, ethanol treatment decreased transepithelial electrical resistance, disrupted TJ formation, and enhanced occludin endocytosis in a dose- and time-dependent manner. These deleterious events were reversed by pretreating the Caco-2 cells with a selective pharmacological inhibitor of macropinocytosis, but not with the inhibitors of clathrin or caveolin-mediated endocytic pathways. Chronic ethanol exposure may increase intestinal permeability by inducing the micropinocytosis of occludin, resulting in the disruption of intestinal TJ.

Severe Burn-Induced Intestinal Epithelial Barrier Dysfunction Is Associated With Endoplasmic Reticulum Stress and Autophagy in Mice.

The disruption of intestinal barrier plays a vital role in the pathophysiological changes after severe burn injury, however, the underlying mechanisms are poorly understood. Severe burn causes the disruption of intestinal tight junction (TJ) barrier. Previous studies have shown that endoplasmic reticulum (ER) stress and autophagy are closely associated with the impairment of intestinal mucosa. Thus, we hypothesize that ER stress and autophagy are likely involved in burn injury-induced intestinal epithelial barrier dysfunction. Mice received a 30% total body surface area (TBSA) full-thickness burn, and were sacrificed at 0, 1, 2, 6, 12 and 24 h postburn. The results showed that intestinal permeability was increased significantly after burn injury, accompanied by the damage of mucosa and the alteration of TJ proteins. Severe burn induced ER stress, as indicated by increased intraluminal chaperone binding protein (BIP), CCAAT/enhancer-binding protein homologous protein (CHOP) and inositol-requiring enzyme 1(IRE1)/X-box binding protein 1 splicing (XBP1). Autophagy was activated after burn injury, as evidenced by the increase of autophagy related protein 5 (ATG5), Beclin 1 and LC3II/LC3I ratio and the decrease of p62. Besides, the number of autophagosomes was also increased after burn injury. The levels of p-PI3K(Ser191), p-PI3K(Ser262), p-AKT(Ser473), and p-mTOR were decreased postburn, suggesting that autophagy-related PI3K/AKT/mTOR pathway is involved in the intestinal epithelial barrier dysfunction following severe burn. In summary, severe burn injury induces the ER stress and autophagy in intestinal epithelia, leading to the disruption of intestinal barrier.

Berberine Ameliorates Ethanol-Induced Disruption of Intestinal Epithelial Tight Junction

Endotoxemia is a crucial factor contributes to alcoholic liver disease (ALD). The disruption of intestinal tight junction (TJ) caused by ethanol is one of the main underlying mechanisms. Berberine is used widely to treat gastrointestinal disorders and reportedly has beneficial effects on TJ integrity. The aims of the present study were to examine the effects of berberine on ethanol-induced intestinal epithelial TJ damage and explore the underlying mechanisms. Caco-2 cell monolayer was treated with ethanol to induce barrier damage. The effect of berberine on TJ was analyzed by measuring the transepithelial electrical resistance (TEER). Subcellular localization of TJ protein zonula occludens-1 (ZO-1) was determined by immunofluorescence microscopy. The expression levels of ZO-1, myosin light chain kinase (MLCK) and MLCK activation were analyzed by Real-time PCR and western blot. Intracellular calcium influx was measured by flow cytometry. The results showed that berberine treatment could significantly improve intestinal barrier function by decreasing intracellular calcium influx, inhibiting MLCK activation., and then reversing ZO-1 distribution. In conclusion, given that berberine also has antimicrobial and anti-inflammatory properties, the effects of berberine on endotoxins translocation are likely to be multifactorial, therefore, berberine should be a promising therapeutic option for ALD. Keywords: Alcoholic liver disease, zonula occludens-1, intracellular calcium influx, myosin light chain kinase. Read more →

The bacterial virulence factor NleA's involvement in intestinal tight junction disruption during Enteropathogenic E. coli infection is independent of its putative PDZ binding domain

Enteropathogenic Escherichia coli (EPEC) is an enteric pathogen able to cause severe diarrhea. Once adhered to the small intestine, EPEC disrupts tight junctions that are important for intestinal barrier function. This disruption is dependent on the bacterial type III secretion system, as well as the translocated effectors EspF and Map. Recently we have shown that a third type III translocated bacterial effector protein, NleA, is also involved in tight junction disruption during EPEC infection. NleA has a predicted PDZ-binding domain at its C-terminus which is proposed to be involved in protein interactions with PDZ domain containing proteins. Since several PDZ-domain-containing proteins localize to tight junctions, we hypothesized that the PDZ-binding domain of NleA might be important for its role in tight junction disruption. However, here we show that a molecular variant of NleA lacking the PDZ-binding domain behaves indistinguishably from the wild-type protein with respect to disruption of tight junctions.

The bacterial virulence factor NleA is required for the disruption of intestinal tight junctions by enteropathogenicEscherichia coli

Enteropathogenic Escherichia coli (EPEC) is a diarrhoeal pathogen that adheres to epithelial cells of the small intestine and uses a type III secretion system to inject effector proteins into host cells. EPEC infection leads to disruption of host intestinal tight junctions that are important for maintaining intestinal barrier function. This disruption is dependent on the bacterial type III secretion system, as well as the translocated effectors EspF and Map. Here we show that a third type III translocated bacterial effector protein, NleA, is also involved in tight junction disruption during EPEC infection. Using the drug Brefeldin A, we demonstrate that the effect of NleA on tight junction integrity is related to its inhibition of host cell protein trafficking through COPII-dependent pathways. These results suggest that NleA's striking effect on virulence is mediated, at least in part, via its role in disruption of intestinal barrier function.