Impaired Epithelial Barrier Function Research Articles

A New Strategy for Dietary Nutrition to Improve Intestinal Homeostasis in Diarrheal Irritable Bowel Syndrome: A Perspective on Intestinal Flora and Intestinal Epithelial Interaction.

Although a reasonable diet is essential for promoting human health, precise nutritional regulation presents a challenge for different physiological conditions. Irritable Bowel Syndrome (IBS) is characterized by recurrent abdominal pain and abnormal bowel habits, and diarrheal IBS (IBS-D) is the most common, seriously affecting patients' quality of life. Therefore, the implementation of precise nutritional interventions for IBS-D has become an urgent challenge in the fields of nutrition and food science. IBS-D intestinal homeostatic imbalance involves intestinal flora disorganization and impaired intestinal epithelial barrier function. A familiar interaction is evident between intestinal flora and intestinal epithelial cells (IECs), which together maintain intestinal homeostasis and health. Dietary patterns, such as the Mediterranean diet, have been shown to regulate gut flora, which in turn improves the body's health by influencing the immune system, the hormonal system, and other metabolic pathways. This review summarized the relationship between intestinal flora, IECs, and IBS-D. It analyzed the mechanism behind IBS-D intestinal homeostatic imbalance by examining the interactions between intestinal flora and IECs, and proposed a precise dietary nutrient intervention strategy. This increases the understanding of the IBS-D-targeted regulation pathways and provides guidance for designing related nutritional intervention strategies.

Interactions between the gut microbiome, associated metabolites and the manifestation and progression of heart failure with preserved ejection fraction in ZSF1 rats

BackgroundHeart failure with preserved ejection fraction (HFpEF) is associated with systemic inflammation, obesity, metabolic syndrome, and gut microbiome changes. Increased trimethylamine-N-oxide (TMAO) levels are predictive for mortality in HFpEF. The TMAO precursor trimethylamine (TMA) is synthesized by the intestinal microbiome, crosses the intestinal barrier and is metabolized to TMAO by hepatic flavin-containing monooxygenases (FMO). The intricate interactions of microbiome alterations and TMAO in relation to HFpEF manifestation and progression are analyzed here.MethodsHealthy lean (L-ZSF1, n = 12) and obese ZSF1 rats with HFpEF (O-ZSF1, n = 12) were studied. HFpEF was confirmed by transthoracic echocardiography, invasive hemodynamic measurements, and detection of N-terminal pro-brain natriuretic peptide (NT-proBNP). TMAO, carnitine, symmetric dimethylarginine (SDMA), and amino acids were measured using mass-spectrometry. The intestinal epithelial barrier was analyzed by immunohistochemistry, in-vitro impedance measurements and determination of plasma lipopolysaccharide via ELISA. Hepatic FMO3 quantity was determined by Western blot. The fecal microbiome at the age of 8, 13 and 20 weeks was assessed using 16s rRNA amplicon sequencing.ResultsIncreased levels of TMAO (+ 54%), carnitine (+ 46%) and the cardiac stress marker NT-proBNP (+ 25%) as well as a pronounced amino acid imbalance were observed in obese rats with HFpEF. SDMA levels in O-ZSF1 were comparable to L-ZSF1, indicating stable kidney function. Anatomy and zonula occludens protein density in the intestinal epithelium remained unchanged, but both impedance measurements and increased levels of LPS indicated an impaired epithelial barrier function. FMO3 was decreased (− 20%) in the enlarged, but histologically normal livers of O-ZSF1. Alpha diversity, as indicated by the Shannon diversity index, was comparable at 8 weeks of age, but decreased by 13 weeks of age, when HFpEF manifests in O-ZSF1. Bray–Curtis dissimilarity (Beta-Diversity) was shown to be effective in differentiating L-ZSF1 from O-ZSF1 at 20 weeks of age. Members of the microbial families Lactobacillaceae, Ruminococcaceae, Erysipelotrichaceae and Lachnospiraceae were significantly differentially abundant in O-ZSF1 and L-ZSF1 rats.ConclusionsIn the ZSF1 HFpEF rat model, increased dietary intake is associated with alterations in gut microbiome composition and bacterial metabolites, an impaired intestinal barrier, and changes in pro-inflammatory and health-predictive metabolic profiles. HFpEF as well as its most common comorbidities obesity and metabolic syndrome and the alterations described here evolve in parallel and are likely to be interrelated and mutually reinforcing. Dietary adaption may have a positive impact on all entities.Graphical abstract

Integrating 16S rRNA Sequencing, Microflora Metabolism, and Network Pharmacology to Investigate the Mechanism of SBL in Alleviating HDM-Induced Allergic Rhinitis.

Allergic rhinitis (AR) is a series of allergic reactions to allergens in the nasal mucosa and is one of the most common allergic diseases that affect both children and adults. Shi-Bi-Lin (SBL) is the modified formula of Cang Er Zi San (CEZS), a traditional Chinese herbal formula used for treating AR. Our study aims to elucidate the anti-inflammatory effects and mechanisms of SBL in house dust mite-induced AR by regulating gut microflora metabolism. In vivo studies showed that nasal allergies and the infiltration of inflammatory cells in the nasal epithelium were significantly suppressed by SBL. Moreover, SBL restored the impaired nasal epithelial barrier function with an increased tight junction protein expression and reduced the endothelial nitric oxide synthase (eNOS). Interestingly, SBL significantly reconstituted the abundance and composition of gut microbiota in AR mice; it increased the relative abundance of potentially beneficial genera and decreased the relative abundance of harmful genera. SBL also restored immune-related metabolisms, which were significantly increased and correlated with suppressing inflammatory cytokines. Furthermore, a network analysis and molecular docking indicated IL-6 was a possible target drug candidate for the SBL treatment. SBL dramatically reduced the IL-6 level in the nasal lavage fluid (NALF), suppressing the IL-6 downstream Erk1/2 and AKT/PI3K signaling pathways. In conclusion, our study integrates 16S rRNA sequencing, microflora metabolism, and network pharmacology to explain the immune mechanism of SBL in alleviating HDM-induced allergic rhinitis.

Postoperative Ileus Murine Model.

Most patients experience postoperative ileus (POI) after surgery, which is associated with increased morbidity, mortality, and hospitalization time. POI is a consequence of mechanical damage during surgery, resulting in disruption of motility in the gastrointestinal tract. The mechanisms of POI are related to aberrant neuronal sensitivity, impaired epithelial barrier function, and increased local inflammation. However, the details remain enigmatic. Therefore, experimental murine models are crucial for elucidating the pathophysiology and mechanism of POI injury and for the development of novel therapies. Here, we introduce a murine model of POI generated via intestinal manipulation (IM) that is similar to clinical surgery; this is achieved by mechanical damage to the small intestine by massaging the abdomen 1-3 times with a cotton swab. IM delayed gastrointestinal transit 24 h after surgery, as assessed by FITC-dextran gavage and fluorescence detection of the segmental digestive tract. Moreover, tissue swelling of the submucosa and immune cell infiltration were investigated by hematoxylin and eosin staining and flow cytometry. Proper pressure of the IM and a hyperemic effect on the intestine are critical for the procedure. This murine model of POI can be utilized to study the mechanisms of intestinal damage and recovery after abdominal surgery.

Circadian rhythm disruption-mediated downregulation of Bmal1 exacerbates DSS-induced colitis by impairing intestinal barrier.

Circadian rhythm disruption (CRD) is thought to increase the risk of inflammatory bowel disease. The deletion of Bmal1, a core transcription factor, leads to a complete loss of the circadian rhythm and exacerbates the severity of dextran sodium sulfate (DSS)-induced colitis in mice. However, the underlying mechanisms by which CRD and Bmal1 mediate IBD are still unclear. We used a CRD mouse model, a mouse colitis model, and an in vitro model of colonic epithelial cell monolayers. We also knocked down and overexpressed Bmal1 in Caco-2 cells by transfecting lentivirus in vitro. The collected colon tissue and treated cells were assessed and analyzed using immunohistochemistry, immunofluorescence staining, quantitative reverse transcription-polymerase chain reaction, western blot, flow cytometry, transmission electron microscopy, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling staining. We found that CRD mice with downregulated Bmal1 expression were more sensitive to DSS-induced colitis and had more severely impaired intestinal barrier function than wild-type mice. Bmal1-/- mice exhibited more severe colitis, accompanied by decreased tight junction protein levels and increased apoptosis of intestinal epithelial cells compared with wild-type mice, which were alleviated by using the autophagy agonist rapamycin. Bmal1 overexpression attenuated Lipopolysaccharide-induced apoptosis of intestinal epithelial cells and impaired intestinal epithelial cells barrier function in vitro, while inhibition of autophagy reversed this protective effect. This study suggests that CRD leads to the downregulation of Bmal1 expression in the colon, which may exacerbate DSS-induced colitis in mice, and that Bmal1 may serve as a novel target for treating inflammatory bowel disease.

Apelin-13 improves pulmonary epithelial barrier function in a mouse model of LPS-induced acute lung injury by inhibiting Chk1-mediated DNA damage

Apelin-13, a type of active peptide, can alleviate lipopolysaccharide (LPS)-induced acute lung injury (ALI). However, the specific mechanism is unclear. Cell cycle checkpoint kinase 1 (Chk1) plays an important role in DNA damage. Here, we investigated the regulatory effect of Apelin on Chk1 in ALI. Chk1-knockout and −overexpression mice were used to explore the role of Chk1 in LPS-induced ALI mice treated with or without Apelin-13. In addition, A549 cells were also treated with LPS to establish a cell model. Chk1 knockdown inhibited the destruction of alveolar structure, the damage of lung epithelial barrier function, and DNA damage in the ALI mouse model. Conversely, Chk1 overexpression had the opposite effect. Furthermore, Apelin-13 reduced Chk1 expression and DNA damage to improve the impaired lung epithelial barrier function in the ALI model. However, the high expression of Chk1 attenuated the protective effect of Apelin-13 on ALI. Notably, Apelin-13 promoted Chk1 degradation through autophagy to regulate DNA damage in LPS-treated A549 cells. In summary, Apelin-13 regulates the expression of Chk1 by promoting autophagy, thereby inhibiting epithelial DNA damage and repairing epithelial barrier function.

Study on the mechanism of hirudin multi target delaying renal function decline in chronic kidney disease based on the “gut-kidney axis” theory

The disorder of the “gut-kidney axis” exacerbates renal function decline in chronic kidney disease (CKD), and current CKD therapy is insufficient to address this issue. Hirudin has a palliative effect on the decline of renal function. However, whether hirudin can delay CKD by regulating the “intestinal renal axis” disorder remains unclear. Unilateral ureteral ligation (UUO) induced CKD rat model, and the rats were treated with bifidobacterium and hirudin for 36 days. After 14 and 36 days of modeling, kidney and colon tissues were collected for pathology, western blot (WB) assay, and quantitative real-time PCR (qPCR) detection. Serum samples were collected for renal function testing. Fecal samples were used for 16S rRNA sequencing and research on fecal bacterial transplantation. Lipopolysaccharide combine with adenosine 5’-triphosphate (LPS + ATP)-induced intestinal epithelial cell injury was treated with a nod-like receptor pyrin domain-associated protein 3 (NLRP3) inhibitor and hirudin. Protein expression was detected using WB and qPCR. The kidneys and colons of the CKD rats exhibited varying degrees of lesions. Creatinine (CRE), blood urea nitrogen (BUN), N-acetyl-β-D-glucosidase (NAG) enzyme, and serum uremic toxins were elevated. The expression of claudin-1 and occludin was decreased, NLRP3 inflammatory-related proteins were increased, and the gut microbiota was disrupted. These pathological changes were more pronounced after 36 days of modeling. Meanwhile, high-dose hirudin treatment significantly improved these lesions and restored the intestinal flora to homeostasis in CKD rats. In vitro, hirudin demonstrated comparable effects to NLRP3 inhibitors by upregulating claudin-1 and occludin expression, and downregulating NLRP3 inflammatory-related proteins expression. The dysbiosis of the gut microbiota and impaired intestinal epithelial barrier function in CKD are associated with renal dysfunction in CKD. Hirudin delays the progression of CKD by regulating the disorder of the “gut-kidney axis” and inhibiting the activation of the NLRP3-ASC-caspase-1 pathway.

The rise of the opportunists: What are the drivers of the increase in infectious diseases caused by environmental and commensal bacteria?

AbstractThe aquaculture sector is currently experiencing a global disease crisis. Emerging bacterial diseases—often opportunistic or syndromic—have collapsed production in nations across the world. Losses in the shrimp industry associated with opportunistic Vibrio spp. exceed 40% of global capacity. This paper reviews potential drivers of syndromic diseases involving opportunistic bacterial pathogens affecting global aquaculture. We provide key examples from major industries where such conditions have prompted greater antibiotic use and have resulted in significant mortality. We challenge the conventional definitions of opportunistic pathogens and propose a fluid categorisation that acknowledges the continuum of host adaptation and the complexity of microbial ecology. We discuss the implications of environmental and dietary stressors such as climate change, coastal eutrophication and pollution, and the transition to plant‐based feeds, which have been linked to impaired epithelial barrier function, gut health disorders and increased disease susceptibility. We critique the ‘one‐pathogen one‐disease’ paradigm, suggesting that Rothman's causal pie model is more useful for understanding opportunistic infections as it emphasises the multicausal nature of disease. We provide examples of bacterial and viral interactions in aquatic disease and occurrence of bacterial diseases resulting from host damage from eukaryotic parasites or increasing frequency and severity of interventions to control such parasites. We recognise the need for corroborative evidence to validate the rise of opportunistic bacterial pathogens as a global trend, and we advocate for the application of nuanced disease causation models to reduce the incidence of opportunistic infections and improve the sustainability of the aquaculture industry.

Mulberry Leaf-Derived Morin Activates β-Catenin by Binding to Frizzled7 to Promote Intestinal Stem Cell Expansion upon Heat-Stable Enterotoxin b Injury.

Intestinal stem cells (ISCs) sustain epithelial renewal by dynamically altering behaviors of proliferation and differentiation in response to various nutrition and stress inputs. However, how ISCs integrate bioactive substance morin cues to protect against heat-stable enterotoxin b (STb) produced by Escherichia coli remains an uncertain question with implications for treating bacterial diarrhea. Our recent work showed that oral mulberry leaf-derived morin improved the growth performance in STb-challenged mice. Furthermore, morin supplementation reinstated the impaired small-intestinal epithelial structure and barrier function by stimulating ISC proliferation and differentiation as well as supporting intestinal organoid expansion ex vivo. Importantly, the Wnt/β-catenin pathway, an ISC fate commitment signal, was reactivated by morin to restore the jejunal crypt-villus architecture in response to STb stimulation. Mechanically, the extracellular morin-initiated β-catenin axis is dependent or partially dependent on the Wnt membrane receptor Frizzled7 (FZD7). Our data reveal an unexpected role of leaf-derived morin, which represents molecular signaling targeting the FZD7 platform instrumental for controlling ISC regeneration upon STb injury.

PDE12 disrupts mitochondrial oxidative phosphorylation and mediates mitochondrial dysfunction to induce oral mucosal epithelial barrier damage in oral submucous fibrosis

Oral submucous fibrosis (OSF) is a chronic oral mucosal disease. The pathological changes of OSF include epithelial damage and subepithelial matrix fibrosis. This study aimed to reveal the epithelial injury mechanism of OSF. A histopathological method was used to analyze oral mucosal tissue from OSF patients and OSF rats. The expression of PDE12 in the oral epithelium was analyzed by immunohistochemistry. The epithelial–mesenchymal transition (EMT) and tight junction proteins in arecoline-treated HOKs were explored by western blotting. Epithelial leakage was assessed by transepithelial electrical resistance and lucifer yellow permeability. The expression of PDE12 and the mitochondrial morphology, mitochondrial permeability transition pore opening, mitochondrial membrane potential, and mitochondrial reactive oxygen species (mtROS) were evaluated in arecoline-induced HOKs. Oxidative phosphorylation (OXPHOS) complexes and ATP content were also explored in HOKs. The results showed significant overexpression of PDE12 in oral mucosal tissue from OSF patients and rats. PDE12 was also overexpressed and aggregated in mitochondria in arecoline-induced HOKs, resulting in dysfunction of OXPHOS and impaired mitochondrial function. An EMT, disruption of tight junctions with epithelial leakage, and extracellular matrix remodeling were also observed. PDE12 overexpression induced by PDE12 plasmid transfection enhanced the mtROS level and interfered with occludin protein localization in HOKs. Interestingly, knockdown of PDE12 clearly ameliorated arecoline-induced mitochondrial dysfunction and epithelial barrier dysfunction in HOKs. Therefore, we concluded that overexpression of PDE12 impaired mitochondrial OXPHOS and mitochondrial function and subsequently impaired epithelial barrier function, ultimately leading to OSF. We suggest that PDE12 may be a new potential target against OSF.

GROUP 3 INNATE LYMPHOID CELLS RELY ON ER STRESS RESPONSE FOR CYTOKINE PRODUCTION IN IBD

Abstract BACKGROUNDS Group 3 innate lymphoid cells (ILC3s) recently emerged as important regulators and potential drug targets for IBD. However, the response of ILC3s to environmental stimuli during intestinal inflammation remains elusive. IRE1a-XBP1 serves as the regulatory hub of the endoplasmic reticulum stress response that plays a vital role in intestinal inflammation. METHODS We generated conditional knockout Ire1aflo/flox,Rorc-Cre (Ire1aΔRorc) and Rag-/-Ire1aΔRorc mice with Ire1a deleted in ILC3s. Intestinal infections were induced by Citrobacter rodentium and Clostridium difficile. ILC3s-derived cytokines including IL-22 is crucial for host defense in the acute phase of C. rodentium and C. difficile infection. Dextran sodium sulfate (DSS) was given in drinking water to induce acute colitis. We recruited patients with active Crohn’s disease (CD) and collected ileal biopsies during routine colonoscopy before starting ustekinumab, a non-selective anti-IL-23 antibody. Mucosal ILC3s were isolated for analysis of ILC3s by flow cytometry. RESULTS In murine intestinal ILC3s, the activity of IRE1a-XBP1 pathway follows a robust 24h circadian rhythm which parallels that of clock genes and cytokines including IL-22. XBP1s is activated in ILC3s ex vivo by vasoactive intestinal peptide, which was previous shown to orchestrate the circadian expression of IL-22 by ILC3s. IRE1a-XBP1 in intestinal ILC3s is further activated in response to IL-23 and colitis in mice, as well as in inflamed IBD tissues compared to normal tissues from healthy individuals. We further showed that IRE1a-XBP1 activation in stimulated ILC3s requires mitochondrial reactive oxygen species (mtROS) (Fig 1). Using the Ire1aΔRorc mice, we demonstrated that IRE1a-XBP1 is critical for cytokine expression by ILC3s. Ire1aΔRorc mice are highly susceptible to acute C. rodentium and C. difficile infection as well as acute DSS colitis (similar vulnerabilities were found in Rag-/-Ire1aΔRorc mice compared to Rag-/-Ire1aflox/flox littermates). Ire1aΔRorc mice exhibit reduced IL-22 and IL-17A in ILC3s and impaired epithelial barrier function with diminished expression of mucins and antimicrobial peptides. The susceptibilities of Ire1aΔRorc and Rag-/-Ire1aΔRorc mice to colitis were rescued by administration of recombinant IL-22 (Fig 2). Moreover, the frequency of XBP1s+ ILC3s in ileal mucosa from CD patients before initiation of ustekinumab positively correlates with response to therapy. CONCLUSIONS We demonstrate that a non-canonical mtROS-IRE1a-XBP1 pathway augments cytokine production by intestinal ILC3s. Loss of IRE1a-XBP1 in ILC3s impairs the optimal and rhythmic expression of protective cytokines and renders the mice more susceptible to intestinal infections and colitis. We also identify XBP1+ ILC3s as a potential biomarker for predicting responsiveness to anti-IL-23 therapies in IBD.

DOP89 Unfolded protein response controls group 3 innate lymphoid cells in IBD

Abstract Background Group 3 innate lymphoid cells (ILC3s) have recently emerged as important regulators and potential drug targets for IBD. However, the response of ILC3s to environmental stimuli during intestinal inflammation remains elusive. IRE1a-XBP1 serves as the regulatory hub of endoplasmic reticulum stress and the unfolded protein response (UPR) that plays a vital role in intestinal inflammation. Methods We generated conditional knockout Ire1aflo/flox,Rorc-Cre (Ire1aΔRorc) and Rag-/-Ire1aΔRorc mice with Ire1a deleted in ILC3s. Intestinal infections were induced by Citrobacter rodentium and Clostridium difficile. ILC3s-derived cytokines including IL-22 is crucial for host defense in the acute phase of C. rodentium and C. difficile infection. Dextran sodium sulfate (DSS) was given in drinking water to induce acute colitis. We recruited patients with active Crohn’s disease (CD) and collected ileal biopsies during routine colonoscopy before starting ustekinumab, a non-selective anti-IL-23 antibody. Mucosal ILC3s were isolated for analysis of ILC3s by flow cytometry. Results In murine intestinal ILC3s, the activity of IRE1a-XBP1 pathway follows a robust 24h circadian rhythm which parallels that of clock genes and cytokines including IL-22. XBP1s is activated in ILC3s ex vivo by vasoactive intestinal peptide, which was previous shown to orchestrate the circadian expression of IL-22 by ILC3s. IRE1a-XBP1 in intestinal ILC3s is further activated in response to IL-23 and colitis in mice, as well as in inflamed IBD tissues compared to normal tissues from healthy individuals. We further showed that IRE1a-XBP1 activation in stimulated ILC3s requires mitochondrial reactive oxygen species (mtROS) (Fig 1). Using the Ire1aΔRorc mice, we demonstrated that IRE1a-XBP1 is critical for cytokine expression by ILC3s. Ire1aΔRorc mice are highly susceptible to acute C. rodentium and C. difficile infection as well as acute DSS colitis (similar vulnerabilities were found in Rag-/-Ire1aΔRorc mice compared to Rag-/-Ire1aflox/flox littermates). Ire1aΔRorc mice exhibit reduced IL-22 and IL-17A in ILC3s and impaired epithelial barrier function with diminished expression of mucins and antimicrobial peptides. The susceptibilities of Ire1aΔRorc and Rag-/-Ire1aΔRorc mice to colitis were rescued by administration of recombinant IL-22 (Fig 2). Moreover, the frequency of XBP1s+ ILC3s in ileal mucosa from CD patients before initiation of ustekinumab positively correlates with response to therapy. Conclusion We demonstrate that a non-canonical mtROS-IRE1a-XBP1 pathway augments cytokine production by intestinal ILC3s. We also identify XBP1+ ILC3s as a potential biomarker for predicting responsiveness to anti-IL-23 therapies in IBD.

Death receptor 5 is required for intestinal stem cell activity during intestinal epithelial renewal at homoeostasis

Intestinal epithelial renewal, which depends on the proliferation and differentiation of intestinal stem cells (ISCs), is essential for epithelial homoeostasis. Understanding the mechanism controlling ISC activity is important. We found that death receptor 5 (DR5) gene deletion (DR5-/-) mice had impaired epithelial absorption and barrier function, resulting in delayed weight gain, which might be related to the general reduction of differentiated epithelial cells. In DR5-/- mice, the expression of ISC marker genes, the number of Olfm4+ ISCs, and the number of Ki67+ and BrdU+ cells in crypt were reduced. Furthermore, DR5 deletion inhibited the expression of lineage differentiation genes driving ISC differentiation into enterocytes, goblet cells, enteroendocrine cells, and Paneth cells. Therefore, DR5 gene loss may inhibit the intestinal epithelial renewal by dampening ISC activity. The ability of crypts from DR5-/- mice to form organoids decreased, and selective DR5 activation by Bioymifi promoted organoid growth and the expression of ISC and intestinal epithelial cell marker genes. Silencing of endogenous DR5 ligand TRAIL in organoids down-regulated the expression of ISC and intestinal epithelial cell marker genes. So, DR5 expressed in intestinal crypts was involved in the regulation of ISC activity. DR5 deletion in vivo or activation in organoids inhibited or enhanced the activity of Wnt, Notch, and BMP signalling through regulating the production of Paneth cell-derived ISC niche factors. DR5 gene deletion caused apoptosis and DNA damage in transit amplifying cells by inhibiting ERK1/2 activity in intestinal crypts. Inhibition of ERK1/2 with PD0325901 dampened the ISC activity and epithelial regeneration. In organoids, when Bioymifi’s effect in activating ERK1/2 activity was completely blocked by PD0325901, its role in stimulating ISC activity and promoting epithelial regeneration was also eliminated. In summary, DR5 in intestinal crypts is essential for ISC activity during epithelial renewal under homoeostasis.

D-Mannose promotes recovery from experimental colitis by inducing AMPK phosphorylation to stimulate epithelial repair.

Delayed mucosal healing and impaired intestinal epithelial barrier function have been implicated in the pathogenesis of ulcerative colitis (UC). Accordingly, restoration of epithelial barrier function as a means to reshape mucosal homeostasis represents an important strategy for use in the treatment of UC. In this study, we examined the role and mechanisms of D-mannose in the recovery of colitis as assessed in both animal and cell models. We found that D-mannose ameliorated inflammation, promoted mucosal healing in the colon and therefore was able to induce the recovery of UC. Furthermore, D-mannose increased the expression of tight junction (TJ) proteins and reduced the intestinal permeability during the recovery of colitis. Moreover, D-mannose inhibited M1 macrophage polarization and promoted M2 macrophage polarization via inducing AMPK phosphorylation while reducing mTOR phosphorylation in both models. In addition, increased TJ protein expression and decreased paracellular permeability were observed in NCM460 cells when incubated with the supernatants of D-mannose-treated RAW264.7 cells, suggesting that M1/M2 polarization induced by D-mannose modulates the expression of TJ proteins. Further study showed that D-mannose significantly upregulated the expression of TJ proteins in DSS-treated NCM460 cells by inducing AMPK phosphorylation, indicating a direct protective effect on epithelial cells. Finally, the protective effects of D-mannose were significantly abrogated by the presence of compound C, an AMPK inhibitor. Taken together, our data indicate that D-mannose can alleviate inflammation and foster epithelial restitution in UC recovery by inducing the TJ protein expression, which are achieved by inducing AMPK phosphorylation in the epithelium and/or macrophages.

LOSS OF EPITHELIAL DUOX2 SIGNALING IS ACCOMPANIED BY A REDUCTION IN AKKERMANSIACEAE AND PROMOTES THE DEVELOPMENT OF METABOLIC SYNDROME IN A MICROBIOME-DEPENDENT MANNER

Dysbiosis, an imbalance in microbial composition or function, is a key player in the development of conditions associated with chronic inflammation. The gut epithelium regulates host-microbiome dynamics through a variety of mechanisms, including the release of reactive oxygen species by NADPH oxidases. Dual oxidase 2 (DUOX2) is an antimicrobial NADPH oxidase expressed in the thyroid gland and gut epithelium and its activity is associated with dysbiosis. Loss of function mutations in DUOX2 are linked to the development of metabolic and inflammatory conditions such as congenital hypothyroidism and inflammatory bowel disease. Metabolic syndrome (MetS) is a cluster of conditions characterized by dysbiosis, inflammation, obesity, glucose intolerance, and hepatic steatosis. Dysbiosis in MetS is associated with a reduction in Bacteroidaceae and Akkermansiaceae. Despite this association, the mechanism by which dysbiosis leads to chronic inflammation and MetS is poorly defined. Here, we investigate the relationship between impaired epithelial barrier function and the development of MetS by focusing for the first time on intestinal specific DUOX2 deficiency as a mechanistic link.

Skin Physiology, Mucosal Functions, and Symptoms Are Modulated by Grass Pollen and Ozone Double Exposure in Allergic Patients

Introduction: Along with climate changes, we see an increase in allergic symptoms and the number of pollen-allergic patients in many countries. Increased allergic symptoms are associated with an elevated ozone exposure which may be linked by impaired epithelial barrier function. This study aimed to quantify the clinical effect of ozone and pollen double exposure (DE). We tested whether ozone impairs barrier-related skin physiology and mucosal functions under DE with pollen in grass pollen-allergic patients versus healthy controls. Methods: This case-control study included 8 grass pollen-allergic patients and 8 non-allergic healthy subjects exposed to grass pollen and ozone in the GA²LEN pollen chamber, comparing shorter and longer DE duration. Non-invasive skin physiological parameters were assessed, including stratum corneum hydration, skin redness, surface pH, and basal transepidermal water loss as a parameter for epidermal barrier function. The subjects’ general well-being, bronchial, nasal, and ocular symptoms were documented. Results: Skin physiology tests revealed that DE in allergic patients deteriorates the epidermal barrier function and increases the surface pH and skin redness. DE significantly induced nasal secretion in pollen-allergic versus healthy subjects, which was more pronounced with longer DE. The general well-being was significantly impaired under DE versus pollen or ozone alone, with a negative influence of DE duration. No relevant bronchial symptoms were recorded. Conclusion: Skin physiology and nasal mucosal symptoms are negatively affected by ozone and grass pollen DE in allergic patients. The negative effects showed, in some parameters, a dose (time)-response relationship. The pH can be regarded as a possible modulatory mechanism.

Dietary zearalenone alters gill epithelial morphology and subsequent risk of infection: Insights from a freshwater grass carp (Ctenopharyngodon idella) model

The mechanical action of zearalenone (ZEA) on different species has been the focus of attention. However, current knowledge of the mode of action and detailed mechanism of ZEA in aquatic species remains limited. According to the maximum detectable dose of commercial fish feed (511 μg kg−1), 1440 freshwater edible grass carp (Ctenopharyngodon idella) with an initial weight of 15.34 ± 0.06 g that were fed six graded levels of dietary ZEA (0, 535, 1041, 1548, 2002 and 2507 μg kg−1 diet). After 70 d, impaired gill epithelial barrier function was confirmed in ZEA-exposed grass carp in terms of the differential expression of tight junctions (TJs), increased oxidative stress levels, and excessive apoptosis promoted cell death. Subsequently, the health status of the fish was determined by immersion (3 h) of Flavobacterium columnare. ZEA-exposed fish displayed the highest incidence of gill rot at 3 d postinfection. Concordantly, at higher doses, ZEA could significantly decrease immune component [(e.g., acid phosphatase (ACP), lysozyme (LZ) and Complement 3 (C3)] activities and antimicrobial peptides (AMPs) transcription abundance and increase substantially inflammatory cytokines. Furthermore, we found that the nuclear factor-erythroid 2-related factor 2 (Nrf2), the nuclear factor κB (NF-κB), and the target of rapamycin (TOR) may be partially involved in associated regulation, thus indicating a potential regulatory network affected by ZEA. In summary, our findings suggest that ZEA disrupts the barrier function of fish gills and increases the risks of pathogen infection. The current recommended guidance values for ZEA in aquaculture (2000 μg kg−1) are insufficient to preclude toxic potential, and increased public concern about ZEA in feed and feedstuff is essential.

Hydrogel‐Based Artificial Mucosa Restores Local Immune and Microbial Homeostasis for Treating Ulcerative Colitis

AbstractUlcerative colitis (UC) is a common inflammatory bowel disease driven by the dyshomeostasis of intestinal immune system and microbiota, but its treatment options are limited due to impaired epithelial barrier integrity and function. Herein, a multifunctional biomimetic hydrogel is developed as artificial colonic mucosa, which can readily establish a bioadhesive barrier at the ulcer site and restore local immune and microbial homeostasis to exert potent anti‐inflammatory effect. The hydrogel framework is generated through the thrombin‐mediated activation and polymerization of fibrinogen that resembles natural blood coagulation mechanism. During this process, the antibacterial agent Ag+ is used for the crosslinking between the generated fibrin units and dopamine/thiol‐modified hyaluronic acid biopolymers via Ag+‐thiol coordination to convey enhanced bioadhesiveness, while the interconnected porous network is employed to encapsulate basic fibroblast growth factor (bFGF) and alanyl‐glutamine (ALG) for mucosa regeneration. The hydrogel stably adheres to the ulcer site to protect and repair the damaged epithelium and establishes an anti‐inflammatory microenvironment by inducing locoregional M1‐to‐M2 macrophage repolarization while eliminating harmful gut flora, which can act in a cooperative manner to substantially alleviate UC symptoms. The biomimetic hydrogel here presents a promising approach for effective UC therapy in the clinics.

Crustin Defense against Vibrio parahaemolyticus Infection by Regulating Intestinal Microbial Balance in Litopenaeus vannamei.

Crustins are a kind of antimicrobial peptide (AMP) that exist in crustaceans. Some crustins do not have direct antimicrobial activity but exhibit in vivo defense functions against Vibrio. However, the underlying molecular mechanism is not clear. Here, the regulatory mechanism was partially revealed along with the characterization of the immune function of a type I crustin, LvCrustin I-2, from Litopenaeus vannamei. LvCrustin I-2 was mainly detected in hemocytes, intestines and gills and was apparently up-regulated after Vibrio parahaemolyticus infection. Although the recombinant LvCrustin I-2 protein possessed neither antibacterial activity nor agglutinating activity, the knockdown of LvCrustin I-2 accelerated the in vivo proliferation of V. parahaemolyticus. Microbiome analysis showed that the balance of intestinal microbiota was impaired after LvCrustin I-2 knockdown. Further transcriptome analysis showed that the intestinal epithelial barrier and immune function were impaired in shrimp after LvCrustin I-2 knockdown. After removing the intestinal bacteria via antibiotic treatment, the phenomenon of impaired intestinal epithelial barrier and immune function disappeared in shrimp after LvCrustin I-2 knockdown. This indicated that the impairment of the shrimp intestine after LvCrustin I-2 knockdown was caused by the dysbiosis of the intestinal microbiota. The present data suggest that crustins could resist pathogen infection through regulating the intestinal microbiota balance, which provides new insights into the functional mechanisms of antimicrobial peptides during pathogen infection.

LOSS OF EPITHELIAL DUOX2 SIGNALING IS ACCOMPANIED BY A REDUCTION IN AKKERMANSIACEAE AND PROMOTES THE DEVELOPMENT OF METABOLIC SYNDROME IN A MICROBIOME-DEPENDENT MANNER

Abstract BACKGROUND Dysbiosis, an imbalance in microbial composition or function, is a key player in the development of conditions associated with chronic inflammation. The gut epithelium regulates host-microbiome dynamics through a variety of mechanisms, including the release of reactive oxygen species by NADPH oxidases. Dual oxidase 2 (DUOX2) is an antimicrobial NADPH oxidase expressed in the thyroid gland and gut epithelium and its activity is associated with dysbiosis. Loss of function mutations in DUOX2 are linked to the development of metabolic and inflammatory conditions such as congenital hypothyroidism and inflammatory bowel disease. Metabolic syndrome (MetS) is a cluster of conditions characterized by dysbiosis, inflammation, obesity, glucose intolerance, and hepatic steatosis. Dysbiosis in MetS is associated with a reduction in Bacteroidaceae and Akkermansiaceae. Despite this association, the mechanism by which dysbiosis leads to chronic inflammation and MetS is poorly defined. Here, we investigate the relationship between impaired epithelial barrier function and the development of MetS by focusing for the first time on intestinal specific DUOX2 deficiency as a mechanistic link. METHODS Mice carrying an intestinal epithelial-specific deletion of DUOX2 (DA IEC-KO), and wild-type (WT) littermates were fed a standard diet and euthanized at 24 weeks. DUOX2 activity was determined by Amplex Red. Metabolic alterations were determined by glucose tolerance tests and body, adipose tissue, and liver weight measurements and histological and gene expression assessments. Intestinal permeability was determined by FITC-dextran and microbial translocation assessments. The role of the microbiome was assessed by RNA-sequencing of stool and in antibiotic-treated mice. RESULTS DA IEC-KO mice produced less epithelial H2O2 and exhibited increased body, liver, and adipose tissue weights that were accompanied by glucose intolerance and increased plasma triglycerides, lipid deposition in the liver, and lipid droplet size in adipocytes. Expression of macrophage and thermogenesis markers, F4/80, CD68, and UCP1 were altered in the liver and adipose tissue of these mice. DA IEC-KO mice showed altered gut permeability and increased bacterial translocation to the liver and adipocytes. Microbial sequencing revealed that loss of intestinal epithelial DUOX2 activity led to significant differences in alpha and beta diversity, and a marked reduction in the family Akkermansiaceae. Antibiotic depletion of the microbiota abrogated all MetS phenotypes observed. CONCLUSION Our findings suggest that loss of intestinal epithelial DUOX2 activity leads to increased intestinal permeability and MetS in a microbiome dependent manner. These findings are significant in that they elucidate a novel role for epithelial DUOX2 at the host-microbial interface with distant systemic effects.