Epithelial Permeability Research Articles

Molecular Mechanisms of N-Acetylcysteine in RSV Infections and Air Pollution-Induced Alterations: A Scoping Review.

N-acetylcysteine (NAC) is a mucolytic agent with antioxidant and anti-inflammatory properties. The respiratory syncytial virus (RSV) is one of the most important etiological factors of lower respiratory tract infections, and exposure to air pollution appears to be additionally associated with higher RSV incidence and disease severity. We aimed to systematically review the existing literature to determine which molecular mechanisms mediate the effects of NAC in an RSV infection and air pollution, and to identify the knowledge gaps in this field. A search for original studies was carried out in three databases and a calibrated extraction grid was used to extract data on the NAC treatment (dose, timing), the air pollutant type, and the most significant mechanisms. We identified only 28 studies conducted in human cellular models (n = 18), animal models (n = 7), and mixed models (n = 3). NAC treatment improves the barrier function of the epithelium damaged by RSV and air pollution, and reduces the epithelial permeability, protecting against viral entry. NAC may also block RSV-activated phosphorylation of the epidermal growth factor receptor (EGFR), which promotes endocytosis and facilitates cell entry. EGFR also enhances the release of a mucin gene, MUC5AC, which increases mucus viscosity and causes goblet cell metaplasia; the effects are abrogated by NAC. NAC blocks virus release from the infected cells, attenuates the cigarette smoke-induced shift from necrosis to apoptosis, and reverses the block in IFN-γ-induced antiviral gene expression caused by the inhibited Stat1 phosphorylation. Increased synthesis of pro-inflammatory cytokines and chemokines is induced by both RSV and air pollutants and is mediated by the nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways that are activated in response to oxidative stress. MCP-1 (monocyte chemoattractant protein-1) and RANTES (regulated upon activation, expressed and secreted by normal T cells) partially mediate airway hyperresponsiveness (AHR), and therapeutic (but not preventive) NAC administration reduces the inflammatory response and has been shown to reduce ozone-induced AHR. Oxidative stress-induced DNA damage and cellular senescence, observed during RSV infection and exposure to air pollution, can be partially reversed by NAC administration, while data on the emphysema formation are disputed. The review identified potential common molecular mechanisms of interest that are affected by NAC and may alleviate both the RSV infection and the effects of air pollution. Data are limited and gaps in knowledge include the optimal timing or dosage of NAC administration, therefore future studies should clarify these uncertainties and verify its practical use.

Colonic Epithelial Permeability to Ions Is Restored after Vedolizumab Treatment and May Predict Clinical Response in Inflammatory Bowel Disease Patients.

Vedolizumab (VDZ) is used for treating inflammatory bowel disease (IBD) patients. A study investigating colonic epithelial barrier function ex vivo following VDZ is lacking. This work aims to evaluate ex vivo the colonic epithelial barrier function in IBD patients at baseline and during VDZ treatment, and to investigate the relationships between barrier function and clinical parameters. Colonic specimens were obtained from 23 IBD patients before, and at 24 and 52 weeks after VDZ treatment, and from 26 healthy volunteers (HV). Transepithelial electrical resistance (TEER, permeability to ions) and paracellular permeability were measured in Ussing chambers. IBD patients showed increased epithelial permeability to ions (TEER, 13.80 ± 1.04 Ω × cm2 vs. HV 20.70 ± 1.52 Ω × cm2, p < 0.001) without changes in paracellular permeability of a 4 kDa probe. VDZ increased TEER (18.09 ± 1.44 Ω × cm2, p < 0.001) after 52 weeks. A clinical response was observed in 58% and 25% of patients at week 24, and in 62% and 50% at week 52, in ulcerative colitis and Crohn's disease, respectively. Clinical and endoscopic scores were strongly associated with TEER. TEER < 14.65 Ω × cm2 predicted response to VDZ (OR 11; CI 2-59). VDZ reduces the increased permeability to ions observed in the colonic epithelium of IBD patients before treatment, in parallel to a clinical, histological (inflammatory infiltrate), and endoscopic improvement. A low TEER predicts clinical response to VDZ therapy.

The New Paradigm: The Role of Proteins and Triggers in the Evolution of Allergic Asthma.

Epithelial barrier damage plays a central role in the development and maintenance of allergic inflammation. Rises in the epithelial barrier permeability of airways alter tissue homeostasis and allow the penetration of allergens and other external agents. Different factors contribute to barrier impairment, such as eosinophilic infiltration and allergen protease action-eosinophilic cationic proteins' effects and allergens' proteolytic activity both contribute significantly to epithelial damage. In the airways, allergen proteases degrade the epithelial junctional proteins, allowing allergen penetration and its uptake by dendritic cells. This increase in allergen-immune system interaction induces the release of alarmins and the activation of type 2 inflammatory pathways, causing or worsening the main symptoms at the skin, bowel, and respiratory levels. We aim to highlight the molecular mechanisms underlying allergenic protease-induced epithelial barrier damage and the role of immune response in allergic asthma onset, maintenance, and progression. Moreover, we will explore potential clinical and radiological biomarkers of airway remodeling in allergic asthma patients.

Gut Epithelial Barrier Function is Impacted by Hyperglycemia and Secondary Bile Acids in Vitro: Possible Rescuing Effects of Specific Pectins.

Gut epithelial barrier disruption is commonly observed in Western diseases like diabetes and inflammatory bowel disease (IBD). Enhanced epithelial permeability triggers inflammatory responses and gut microbiota dysbiosis. Reduced bacterial diversity in IBD affects gut microbiota metabolism, altering microbial products such as secondary bile acids (BAs), which potentially play a role in gut barrier regulation and immunity. Dietary fibers such as pectin may substitute effects of these BAs. The study examines transepithelial electrical resistance of gut epithelial T84 cells and the gene expression of tight junctions after exposure to (un)sulfated secondary BAs. This is compared to the impact of the dietary fiber pectin with different degrees of methylation (DM) and blockiness (DB), with disruption induced by calcium ionophore A23187 under both normal and hyperglycemic conditions. Unsulfated lithocholic acid (LCA) and deoxycholic acid (DCA) show a stronger rescuing effect, particularly evident under 20mM glucose levels. DM19 with high DB (HB) and DM43HB pectin exhibit rescuing effects under both glucose conditions. Notably, DM19HB and DM43HB display higher rescue effects under 20mM glucose compared to 5mM glucose. The study demonstrates that specific pectins such as DM19HB and DM43HB may serve as alternatives for preventing barrier disruption in the case of disturbed DCA metabolism.

Cryptosporidium parvum disrupts intestinal epithelial barrier in neonatal mice through downregulation of cell junction molecules.

Cryptosporidium spp. cause watery diarrhea in humans and animals, especially in infants and neonates. They parasitize the apical surface of the epithelial cells in the intestinal lumen. However, the pathogenesis of Cryptosporidium-induced diarrhea is not fully understood yet. In this study, we infected C57BL/6j neonatal mice with C. parvum IIa and IId subtypes, and examined oocyst burden, pathological changes, and intestinal epithelial permeability during the infection. In addition, transcriptomic analyses were used to study the mechanism of diarrhea induced by the C. parvum IId subtype. The neonatal mice were sensitive to both C. parvum IIa and IId infection, but the IId subtype caused a wide oocyst shedding window and maintained the high oocyst burden in the mice compared with the IIa subtype. In addition, the mice infected with C. parvum IId resulted in severe intestinal damage at the peak of infection, leading to increased permeability of the epithelial barrier. The KEGG, GO and GSEA analyses revealed that the downregulation of adherens junction and cell junction molecules at 11 dpi. Meanwhile, E-cadherin, which is associated with adherens junction, was reduced at the protein level in mouse ileum at peak and late infection. C. parvum IId infection causes more severe pathological damage than C. parvum IIa infection in neonatal mice. Furthermore, the impairment of the epithelial barrier during C. parvum IId infection results from the downregulation of intestinal junction proteins.

Phospholipids of inhaled liposomes determine the in vivo fate and therapeutic effects of salvianolic acid B on idiopathic pulmonary fibrosis

Since phospholipids have an important effect on the size, surface potential and hardness of liposomes that decide their in vivo fate after inhalation, this research has systematically evaluated the effect of phospholipids on pulmonary drug delivery by liposomes. In this study, liposomes composed of neutral saturated/unsaturated phospholipids, anionic and cationic phospholipids were constructed to investigate how surface potential and the degree of saturation of fatty acid chains determined their mucus and epithelium permeability both in vitro and in vivo. Our results clearly indicated that liposomes composed of saturated neutral and anionic phospholipids possessed high stability and permeability, compared to that of liposomes composed of unsaturated phospholipids and cationic phospholipids. Furthermore, both in vivo imaging of fluorescence-labeled liposomes and biodistribution of salvianolic acid B (SAB) that encapsulated in liposomes were performed to estimate the effect of phospholipids on the lung exposure and retention of inhaled liposomes. Finally, inhaled SAB-loaded liposomes exhibited enhanced therapeutic effects in a bleomycin-induced idiopathic pulmonary fibrosis mice model via inhibition of inflammation and regulation on coagulation-fibrinolytic system. Such findings will be beneficial to the development of inhalable lipid-based nanodrug delivery systems for the treatment of respiratory diseases where inhalation is the preferred route of administration.

Systematic-Narrative Hybrid Literature Review: Crosstalk between Gastrointestinal Renin-Angiotensin and Dopaminergic Systems in the Regulation of Intestinal Permeability by Tight Junctions.

In the first part of this article, the role of intestinal epithelial tight junctions (TJs), together with gastrointestinal dopaminergic and renin-angiotensin systems, are narratively reviewed to provide sufficient background. In the second part, the current experimental data on the interplay between gastrointestinal (GI) dopaminergic and renin-angiotensin systems in the regulation of intestinal epithelial permeability are reviewed in a systematic manner using the PRISMA methodology. Experimental data confirmed the copresence of DOPA decarboxylase (DDC) and angiotensin converting enzyme 2 (ACE2) in human and rodent enterocytes. The intestinal barrier structure and integrity can be altered by angiotensin (1-7) and dopamine (DA). Both renin-angiotensin and dopaminergic systems influence intestinal Na+/K+-ATPase activity, thus maintaining electrolyte and nutritional homeostasis. The colocalization of B0AT1 and ACE2 indicates the direct role of the renin-angiotensin system in amino acid absorption. Yet, more studies are needed to thoroughly define the structural and functional interaction between TJ-associated proteins and GI renin-angiotensin and dopaminergic systems.

Interplay of hypoxia and host cells in the translocation, invasion and dissemination of Taenia solium in neurocysticercosis

Neurocysticercosis, caused by the invasion of the central nervous system (CNS) by Taenia solium larvae, poses a significant global health burden. Despite its prevalence and severe neurological consequences, understanding of the molecular and signalling pathways facilitating parasite dissemination and CNS invasion is limited. The lack of comprehensive knowledge of host-parasite interactions and associated proteins involved in T. solium infection hinders the development of targeted interventions to mitigate its ability to cross the epithelia barrier. This is complicated by reduced oxygen availability in the intestine, a phenomenon called hypoxia. Hypoxia can result in epithelial barrier disruption and cell damage, thereby promoting the translocation and dissemination of T. solium. This study aims to establish the role of hypoxia in T. solium invasion and disseminated infections. The effect of hypoxia on the migration, viability and morphological characteristics of T. solium would be determined using transwell invasion assays, flow cytometry and microscopy. T. solium oncosphere development and dissemination under hypoxic and normoxic conditions will be monitored using animal models. Also, host-parasite transcriptome and proteome profiling will be performed to determine pathways triggered under hypoxic conditions. It is expected that hypoxia would promote the invasion and dissemination of T. solium by enhancing epithelial and endothelial cell permeability. , hypoxia will induce the expression of binding and adhesion proteins and other virulence markers such as enolase, serpin, and glutathione transferases that are involved in host invasion. Understanding the role of hypoxia in the translocation mechanism of T. solium can be leveraged to provide insights into host tissues dissemination and the development of appropriate interventions.

Prevotella melaninogenica disrupted oral epithelial barrier function via myosin light chain kinase.

Our previous studies have found that the composition ratio of Prevotella melaninogenica (Pm) on buccal mucosa surface of oral lichen planus (OLP) patients increased significantly compared with control. Furthermore, Pm could invade the epithelium of OLP patients. This study aimed to further explore the impact of Pm on oral keratinocytes. The Pm-human oral keratinocyte (HOK) co-culture model was established to detect monolayer permeability, zona occludens-1 (ZO-1) expression, and intracellular survival of Pm. We performed RNA-seq followed by identification of differentially expressed genes (DEGs) and Gene Ontology (GO) analysis, with a particular focus on myosin light chain kinase (MLCK). An MLCK inhibitor ML-7 was utilized in Pm-HOK co-culture model to assess its effects on monolayer permeability and ZO-1 expression. HOK monolayer permeability was increased, and ZO-1 expression was decreased after co-culture (p < 0.05). Pm could survive in HOK cells. RNA-seq revealed MLCK was an upregulated common DEG. The expression of MLCK in the Pm-HOK co-culture model was upregulated. Inhibition of MLCK rescued the increased epithelial permeability, and ZO-1 expression was upregulated (p < 0.05). MLCK may be involved in disrupting epithelial barrier function by Pm.

Bifidobacterium bifidum inhibits the TNF-α induced increase in intestinal epithelial tight junction permeability via a TLR-2/TLR-6 and PPAR-γ dependent inhibition of NF-κB p50/p65

Background: Bifidobacterium bifidum (BB) is one of the most widely used probiotic bacterial species and BB has been shown to have a protective effect against intestinal inflammation by an enhancement of the intestinal epithelial tight junction (TJ) barrier. Previous studies from our laboratory showed that a specific BB strain, referred to as BB1, caused a marked enhancement of intestinal epithelial TJ barrier, while strain BB4 had no effect. TNF-α contributes to the development of intestinal inflammation in inflammatory conditions of the gut, in part by increasing intestinal permeability and antigenic penetration. TNF-α induced increase in intestinal epithelial TJ permeability is mediated by an NF-κB p50/p65 activation of myosin light chain kinase (MLCK) gene and MLCK-induced opening of the intestinal TJ barrier. Peroxisome proliferator-activated receptor gamma (PPAR-γ) is an important nuclear receptor known to have anti-inflammatory activity by interfering with NF-κB activation. However, the protective effect of BB1 against TNF-α induced increase in intestinal TJ permeability remain unclear. Aims: The major purpose of this study was to delineate the protective effect of BB1 against the TNF-α induced increase in intestinal TJ permeability and to elucidate the intracellular mechanism involved. Methods: Filter-grown Caco-2 monolayers ( in vitro) and recycling intestinal perfusion of live mice ( in vivo) were the model systems used to assess intestinal TJ permeability. Results: 1) TNF-α caused an increase Caco-2 TJ permeability, as assed by Caco-2 trans-epithelial resistance (TER) and trans-epithelial flux of paracellular marker, FITC-Dextran 10kD. 2) Addition of BB1 (1x108 CFU/ml) to the apical membrane compartment inhibited the TNF-α increase in Caco-2 epithelial TJ permeability, while strain BB4 had no effect. 3) BB1 but not BB4, caused an increase in Caco-2 TLR-2 expression on the apical membrane surface, and siRNA induced knockdown of TLR-2 prevent the BB1 inhibition of TNF-α induced increase in Caco-2 TJ permeability. 4) TNF-α caused a rapid phosphorylation and degradation of Caco-2 IKK-α, as well as and phosphorylation and nuclear translocation of NF-κB p50/p65, indicating NF-κB activation. BB1, but not BB4, inhibited the NF-κB activation. 5) TNF-α activation of NF-κB was associated with an increase in MLCK gene (increase in MLCK promoter activity and mRNA transcription) and kinase activity. BB1, but not BB4, inhibited the TNF-α induced increase in MLCK gene and kinase activity. 6) BB1 but not BB4, caused a rapid PPAR-γ activation (PPAR-γ phosphorylation and nuclear translocation), and siRNA induced knockdown of PPAR-γ inhibited the TNF-α induced activation of NF-κB and increase in Caco-2 TJ permeability. 7) Intraperitoneal injection of TNF-α (5 mg) also caused time-dependent increase in mouse small intestinal permeability (IP) to FITC-Dextran 10kD. TNF-α caused a degradation of IkB-α in intestinal tissue and nuclear translocation of NF-kB in mouse enterocyte. 8) BB1, but not BB4, selectively activated PPAR-γ in mouse enterocytes and inhibited the TNF-α-induced activation of NF-κB, resulting in a decrease in mouse IP. 9) BB1 inhibition of TNF-α activation of mouse enterocyte NF-κB and increase in IP was prevented in mice with intestinal epithelial cell-specific knockout of PPAR-γ Villin-cre and TLR-2 Villin-cre mice. Conclusion: BB1 protects against the TNF-α induced increase in intestinal TJ permeability in a strain-specific manner. BB1 protects against the TNF-α induced increase in intestinal TJ permeability via a novel intracellular mechanism involving BB1/TLR-2 pathway activation of PPAR-γ, and PPAR-γ mediated inhibition of NF-κB p50/p65 and MLCK gene in enterocyte. This research project was supported by the National Institute of Diabetes and Digestive and Kidney Diseases R01-DK-121073-01. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Lack of Pkd1 enhances intestinal integrity and motility prior to accelerated kidney cyst growth

Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disorder, primarily caused by PKD1 germline mutations. Polycystin-1, PKD1’s encoded protein, is essential to maintain kidney epithelial cell integrity, but its role in the gut has yet to be determined. Considering macrophage infiltration precedes accelerated cyst growth in Pkd1 knockout ( Pkd1KO) mice, and the gut is the body’s largest reservoir of inflammatory cells, we hypothesized that loss of Pkd1 would weaken the intestinal epithelial cell barrier and mount an inflammatory response, potentially preceding kidney damage. Plasma, urine, kidney, and intestines were collected from 11-12 week old male and female conditional Pkd1KO (CAGG-creER; tamoxifen-induced at 5-6 weeks) and flox (control) mice. Plasma was used to measure intestinal permeability as well as monocyte chemoattractant protein-1 (MCP-1) and lipocalin-2 (LCN-2) concentration. Urine was utilized to assess kidney injury, while tissues were used for RT-PCR and histology. Pkd1KO mice had significantly elevated kidney-to-body weight ratio, cyst ratio, and expression of tubular injury markers kidney injury molecule 1 (Kim1) and Lcn2 compared to flox mice. Despite this cellular injury, kidney inflammatory gene expression and urine osmolality did not differ between genotypes. No overt morphological changes were observed in the ileum or colon, nor were there differences in expression of proinflammatory chemokines or cytokines in either tissue. Plasma abundance of MCP-1 and LCN-2 was similar between genotypes. Epithelial permeability in the small and large intestines was significantly decreased in Pkd1KO versus flox mice. Moreover, Pkd1KO mice had a significantly reduced whole gut transit time compared to flox mice. This corresponded with increases in colonic expression of epithelial integrity marker occludin and water channels aquaporin 4 and aquaporin 8 in Pkd1KO mice. Together, these data insinuate that loss of Pkd1 in the intestines leads to a tighter intestinal barrier accompanied by increased water channel expression and gut motility. These findings appear unrelated to inflammatory involvement in early PKD. These studies were supported by NIH T32 DK007545 to RS, R01 DK132028 to TS, and R03 DK119717 to TS. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Role of CRF1 and CRF2 Receptors in Stress-induced Increase of Intestinal Epithelial Permeability in the Mouse Colon

Background and Aims: Stress is known to increase intestinal permeability, which may contribute to the development of irritable bowel syndrome (IBS) and other intestinal disorders. Corticotropin releasing factor (CRF) has been proven to mediate stress-induced changes in gut function, such as increased colonic motility, ion secretion, and intestinal permeability. Actions of CRF are mediated by two G-protein coupled receptors, CRF1 and CRF2; both are expressed in the intestine. However, the roles of the CRF1 and CRF2 receptors in stress-induced increase of intestinal epithelial permeability remain unknown. Methods: Adult male and female C57BL/6 mice were restrained (1h/d) for five consecutive days. The selective CRF1 and CRF2 receptor antagonists were injected intraperitoneally 15 min prior to the restraint stress. Control mice were kept in their home cages without restraint. After the last restraint stress/control session, the proximal colon was removed, and the mucosa/submucosa preparations were dissected and mounted onto the Ussing Chamber System for the measurement of intestinal permeability. Transepithelial resistance (TER) and FITC-inulin (4kD) flux across the colonic epithelium were used as indicators of paracellular permeability. Horseradish peroxidase (HRP, 40kD) flux was used to measure transcellular permeability. Results: Restraint stress did not alter TER or FITC-inulin flux in male mice. Pretreatment with the CRF1 receptor antagonist NBI 27914 or the CRF2 receptor antagonist antisauvagine-30 had no effect on TER or FITC-inulin flux in male mice. In female mice, restraint stress significantly reduced TER and increased FITC-inulin flux in the proximal colon, which were blocked by NBI 27914 but not by antisauvagine-30. Restraint stress significantly increased HRP flux in both male and female mice. In male mice, pretreatment with NBI 27914 significantly reduced stress-evoked increase of HRP flux, while antisauvagine-30 had no effect. In female mice, both NBI 27914 and antisauvagine-30 significantly reduced stress-evoked increase of HRP flux. Conclusions: Stress evokes different changes in epithelial permeability in the proximal colon of male and female mice. In male mice, stress does not influence paracellular permeability but increases transcellular permeability, which is mediated by the CRF1 receptor. In female mice, stress increases both transcellular and paracellular permeability. Stress-induced increase of paracellular permeability was mediated by the CRF1 receptor, but the increased transcellular permeability was mediated by both CRF1 and CRF2 receptors. University of Wisconsin-La Crosse Undergraduate Research Grant (CS and LB). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Neuregulin-1 reduces necroptosis and protects mice from death caused by respiratory viral infection

Abstract Respiratory syncytial virus (RSV) and influenza type A virus (IAV) cause significant morbidity and mortality even with available vaccines. Pre-existing atopy provides protection against death in mice infected with Sendai virus (SeV), a rodent virus related to RSV. We found neuregulin-1 (NRG1) expressed at high levels in atopic mice, and when given exogenously (500 ng/day i.n for five days) to non-atopic mice provides significant protection against mortality from lethal doses of both SeV (2x106 pfu) and IAV (3x103 pfu PR8-mouse adapted). Further, using well-differentiated human airway epithelial cells, NRG-1 was found to improve epithelial barrier function following RSV infection. RSV has been reported to cause necroptosis, which can induce inflammation and alter epithelial membrane permeability. We hypothesized the survival advantage of NRG1 involves inhibition of necroptosis. Treating cells from the murine airway epithelial cell line LET-1 with NRG1 before inoculation with SeV significantly inhibited viral induced necroptosis as measured by a real-time non-lytic apoptosis/necroptosis assay. Furthermore, NRG1 reduced expression of genes promoting necroptosis (RIPK3 and MLKL) in well-differentiated human bronchial epithelial cells 72h post RSV infection. Together, these data suggest NRG1 modulates viral induced cell death through inhibition of necroptosis of epithelial cells and may explain the reduced airway leak and increased survival seen in our model.

The leaky gut: a narrative review on the role of epithelial cell permeability in necrotizing enterocolitis

The leaky gut: a narrative review on the role of epithelial cell permeability in necrotizing enterocolitis

Blockade of PI3K/AKT signaling pathway by Astragaloside IV attenuates ulcerative colitis via improving the intestinal epithelial barrier

BackgroundThe specific pathogenesis of UC is still unclear, but it has been clear that defects in intestinal barrier function play an important role in it. There is a temporary lack of specific drugs for clinical treatment. Astragaloside IV (AS-IV) is one of the main active ingredients extracted from Astragalus root and is a common Chinese herbal medicine for the treatment of gastrointestinal diseases. This study aimed to determine whether AS-IV has therapeutic value for DSS or LPS-induced intestinal epithelial barrier dysfunction in vivo and in vitro and its potential molecular mechanisms.MethodsThe intestinal tissues from UC patients and colitis mice were collected, intestinal inflammation was observed by colonoscopy, and mucosal barrier function was measured by immunofluorescence staining. PI3K/AKT signaling pathway activator YS-49 and inhibitor LY-29 were administered to colitic mice to uncover the effect of this pathway on gut mucosal barrier modulation. Then, network pharmacology was used to screen Astragaloside IV (AS-IV), a core active component of the traditional Chinese medicine Astragalus membranaceus. The potential of AS-IV for intestinal barrier function repairment and UC treatment through blockade of the PI3K/AKT pathway was further confirmed by histopathological staining, FITC-dextran, transmission electron microscopy, ELISA, immunofluorescence, qRT-PCR, and western blotting. Finally, 16 S rRNA sequencing was performed to uncover whether AS-IV can ameliorate UC by regulating gut microbiota homeostasis.ResultsMucosal barrier function was significantly damaged in UC patients and murine colitis, and the activated PI3K/AKT signaling pathway was extensively involved. Both in vivo and vitro showed that the AS-IV-treated group significantly relieved inflammation and improved intestinal epithelial permeability by inhibiting the activation of the PI3K/AKT signaling pathway. In addition, microbiome data found that gut microbiota participates in AS-IV–mediated intestinal barrier recovery as well.ConclusionsOur study highlights that AS-IV exerts a protective effect on the integrality of the mucosal barrier in UC based on the PI3K/AKT pathway, and AS-IV may serve as a novel AKT inhibitor to provide a potential therapy for UC.Graphical abstract

Sclareol protected against intestinal barrier dysfunction ameliorating Crohn's disease-like colitis via Nrf2/NF-B/MLCK signalling

BackgroundInflammation-induced intestinal barrier dysfunction is not only a pathological feature of Crohn's disease (CD) but also an important therapeutic target. Sclareol (SCL) is a nontoxic natural plant compound with anti-inflammatory effect, but its role in CD has not been established. MethodsIn vivo studies of mice with TNBS-induced colitis were carried out to evaluate the effects of SCL on CD-like colitis and intestinal barrier function. In vitro, a TNF-α-induced colonic organoid model was established to test the direct effect of SCL on inflammation-induced intestinal barrier injure and inflammatory response. The Nrf2/NF-κB/MLCK signalling was analysed to explore the mechanism of SCL. ResultsIn vivo, SCL largely alleviated the colitis in TNBS mice, as evidenced by improvements in the weight loss, colitis symptoms, endoscopic score, macroscopic histological score, and histological inflammation score. Moreover, SCL significantly improved intestinal barrier dysfunction, manifested as reduced intestinal permeability and decreased intestinal bacterial translocation in TNBS mice. Importantly, SCL antagonised the intestinal mucosal inflammation while protecting tight junctions in TNBS mice. In vitro, SCL largely depressed pro-inflammatory cytokines levels and improved intestinal epithelial permeability in a TNF-α-induced colonic organoid model. In the context of CD, the protective effects of SCL against inflammation and intestinal barrier damage are at least partially results from the Nrf2 signalling activation and the NF-κB/MLCK signalling inhibition. ConclusionsSCL improved intestinal barrier dysfunction and alleviated CD-like colitis, possibly through modulation of Nrf2/NF-κB/MLCK signalling. In view of SCL's safety profile, there is hope that it will be useful in the clinic.

Modulation of intestinal epithelial permeability by chronic small intestinal helminth infections.

Increased permeability of the intestinal epithelial layer is linked to the pathogenesis and perpetuation of a wide range of intestinal and extra-intestinal diseases. Infecting humans with controlled doses of helminths, such as human hookworm (termed hookworm therapy), is proposed as a treatment for many of the same diseases. Helminths induce immunoregulatory changes in their host which could decrease epithelial permeability, which is highlighted as a potential mechanism through which helminths treat disease. Despite this, the influence of a chronic helminth infection on epithelial permeability remains unclear. This study uses the chronically infecting intestinal helminth Heligmosomoides polygyrus to reveal alterations in the expression of intestinal tight junction proteins and epithelial permeability during the infection course. In the acute infection phase (1 week postinfection), an increase in intestinal epithelial permeability is observed. Consistent with this finding, jejunal claudin-2 is upregulated and tricellulin is downregulated. By contrast, in the chronic infection phase (6 weeks postinfection), colonic claudin-1 is upregulated and epithelial permeability decreases. Importantly, this study also investigates changes in epithelial permeability in a small human cohort experimentally challenged with the human hookworm, Necator americanus. It demonstrates a trend toward small intestinal permeability increasing in the acute infection phase (8 weeks postinfection), and colonic and whole gut permeability decreasing in the chronic infection phase (24 weeks postinfection), suggesting a conserved epithelial response between humans and mice. In summary, our findings demonstrate dynamic changes in epithelial permeability during a chronic helminth infection and provide another plausible mechanism by which chronic helminth infections could be utilized to treat disease.

Gallic acid mitigates intestinal inflammation and loss of tight junction protein expression using a 2D-Caco-2 and RAW 264.7 co-culture model

A 2D-intestinal epithelial Caco-2/RAW 264.7 macrophage co-culture model was developed to demonstrate the relative efficacy of different phenolic acids to mitigate changes in Caco-2 epithelial cell redox state initiated both directly by autoxidation products, H2O2, and indirectly through cell communication events originating from cytokine stimulated macrophage. An inducer cocktail (lipopolysaccharide + interferon gamma) was used to activate RAW 264.7 cells in the 2D- Caco-2/RAW co-culture and intracellular changes in Caco-2 cell redox signaling occurred in response to positive changes (p < 0.05) in inflammatory biomarkers derived in macrophage that included IL-6, TNF-α, nitric oxide and peroxynitrite, respectively. Phenolic acids varied in relative capacity to reduce NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) in cocktail inflamed induced macrophage. This response in addition to the relative predisposition of gallic acid (GA) to undergo autoxidation to generate H2O2 activity (p < 0.05), culminated in downstream cell signaling in Caco-2 nuclear factor erythroid 2-related factor (Nrf2) activity (increase 26.9 %), altered monolayer integrity (increase 33.7 %), and release of interleukin 8 (IL-8) (decrease 80.5 %) (p < 0.05). It can be concluded that the co-culture model described herein was useful to assess the importance of communication between cytokine stimulated macrophage and intestinal cells. Moreover, the relative unique efficacy of GA, compared to other phenolic acids tested to protect against activated macrophage induced changes related to intestinal dysfunction were particularly relevant to epithelial redox signaling, intestinal permeability and regulation of tight junction proteins. This study concludes that phenolic acids are not equal in the capacity to protect against intestinal cell dysfunction despite some indication of biological activity.

A novel approach to the dual sugar test for the assessment of intestinal epithelium permeability in response to exertional heat stress and nutritional intervention

An acute increase in intestinal epithelium permeability is induced by prolonged exertion in the heat, resulting in the translocation of pathogenic bacteria and endotoxins from the lumen into the circulation, causing a systemic inflammatory response and debilitating symptoms(1). Acute exercise-induced gastrointestinal syndrome mimics chronic health conditions with which an impaired intestinal barrier function is associated, including coeliac disease, inflammatory bowel disease, diabetes, Alzheimer’s and liver diseases(2). Intestinal epithelium permeability is typically assessed using a dual sugar absorption test, by administering a drink containing non-metabolisable sugars (e.g. lactulose [L] and L-rhamnose [R]) that can enter the circulation by paracellular translocation when the epithelium is compromised, and are subsequently excreted and measured cumulatively in the urine(3). We aimed to demonstrate that our recently developed ion chromatography protocol(4) can be used to accurately quantify L/R ratio in the plasma of participants exercising in hot ambient conditions and to determine the impact of nutritional intervention on intestinal epithelium permeability. Further, we hypothesised that measuring L/R in plasma collected at intervals during the post-exercise recovery period would reveal more information about intestinal permeability compared to previously published cumulative urine L/R data(3). Endurance-trained participants completed a set of randomised crossover studies, consisting of 2 h running at 60% V˙O2max in temperate, warm and hot ambient conditions (n = 8) and/or in the heat while consuming water, carbohydrate or protein (n = 9). The dual sugar solution was ingested at 90 min of exercise and blood was sampled at 0, 1, 2 and 4 h post-exercise. Plasma sugars were quantified by high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and L/R ratios were compared by two-way repeated measures ANOVA with Tukey’s multiple comparisons. Plasma L/R increased immediately post-exercise in the heat (0.15 ± 0.11) compared with temperate (0.06 ± 0.04, p&lt;0.001) and warm (0.09 ± 0.08, p&lt;0.01) conditions, while consuming glucose before and during exercise alleviated this (0.02 ± 0.02, p&lt;0.001), and this novel information was otherwise missed when measuring urine L/R. Consuming glucose or whey protein hydrolysate during exercise attenuated intestinal permeability from exertional heat stress throughout recovery, with the mean plasma L/R over 4 h reduced from 0.11 ± 0.05 to 0.04 ± 0.03 (p&lt;0.001) and 0.06 ± 0.04 (p&lt;0.01) with glucose and protein, respectively. We recommend using the dual sugar test with quantification of plasma sugars at intervals by HPAEC-PAD to maximise intestinal permeability data collection in exercise gastroenterology research and beyond, as this gives additional acute response information compared to urinary measurements. Our approach can be employed to investigate and develop personalised nutrition strategies that prevent intestinal hyperpermeability during exertional heat stress. This has implications for athlete performance and safety, and can also build upon occupational health and safety practices and inform chronic disease management.

Assessment of the endothelial glycocalyx state in patients with axial spondyloarthritis associated with Crohn’s disease

BACKGROUND: Epithelial barrier dysfunction, including their permeability increment, is supposed to be a pathogenetic link in the development of various immunoinflammatory diseases, including Crohn’s disease and axial spondyloarthritits. Endothelial permeability regulation is provided by both intercellular contacts and endothelial glycocalyx. AIM: To study the microcirculatory state by measuring the endothelial glycocalyx thickness in patients with axial spondyloarthritits associated with Crohn’s disease. To identify an association between the endothelial glycocalyx thinning and the epithelial permeability severity in patients with Crohn’s disease, axial spondyloarthritits and their combination. MATERIALS AND METHODS: 12 patients with axial spondyloarthritits associated with Crohn’s disease (group A), 22 patients with idiopathic axial spondyloarthritits (group B), 24 patients with Crohn’s disease (group C) and 16 healthy individuals (group D) were examined. The endothelial glycocalyx thickness was assessed in the sublingual region using dark-field microscopy. Fecal calprotectin and fecal zonulin was studied. RESULTS: A statistically significant increase in the perfused boundary region high flow was found (p = 0.001), as well as the MicroVascular Health Score (p = 0.004) in group A compared with the control. In Group A, a positive correlation between perfused boundary region high flow and scores Ankylosing Spondylitis Disease Activity Score, Bath Ankylosing Spondylitis Disease Activity Index, Bath Ankylosing Spondylitis Metrology Index 10 (p 0.05) was found; perfused boundary region 5–9 mµ correlated positively with the fecal calprotectin (p = 0.007). Perfused boundary region high flow measurement provides 77.1% accuracy for correct classification of axial spondyloarthritits associated with Crohn’s disease and idiopathic axial spondyloarthritits (p = 0.010). In group C, a positive correlation between fecal zonulin and perfused boundary region 5–9 mµ, 10–19 mµ, 5–25 mµ (p 0.05) was also found. CONCLUSIONS: The findings suggest that the patients with axial spondyloarthritits associated with Crohn’s disease have endothelial glycocalyx impairment; the detection of these abnormalities may be useful in assessing the disease integral activity and in the differential diagnosis of axial spondyloarthritits associated with Crohn’s disease and idiopathic axial spondyloarthritits. Further studies of microcirculation will allow better understanding of spondyloarthritis and inflammatory bowel diseases pathogenesis and improve the management of patients with these diseases.