Effect of toll-like receptor 3 agonist poly I:C on intestinal mucosa and epithelial barrier function in mouse models of acute colitis

AIM To investigate potential effects of poly I:C on mucosal injury and epithelial barrier disruption in dextran sulfate sodium(DSS)-induced acute colitis.METHODS Thirty C57BL/6 mice were given either regular drinking water(control group) or 2%(w/v) DSS drinking water(model and poly I:C groups) ad libitum for 7 d. Poly I:C was administrated subcutaneously (20 μg/mouse) 2 h prior to DSS induction in mice of the poly I:C group. Severity of colitis was evaluated by disease activity index, body weight, colon length, histology and myeloperoxidase (MPO) activity, as well as the production of proinflammatory cytokines, including tumor necrosis factor-α(TNF-α), interleukin 17(IL-17) and interferon-γ (IFN-γ). Intestinal permeability was analyzed by the fluorescein isothiocyanate labeleddextran (FITC-D) method. Ultrastructural features of the colon tissue were observed under electron microscopy. Expressions of tight junction(TJ) proteins, including zo-1, occludin and claudin-1, were measured by immunohistochemistry/immunofluorescence, Western blot and real-time quantitative polymerase chain reaction(RT-qPCR).RESULTS DSS caused significant damage to the colon tissue in the model group. Administration of poly I:C dramatically protected against DSS-induced colitis, as demonstrated by less body weight loss, lower disease activity index score, longer colon length, colonic MPO activity, and improved macroscopic and histological scores. It also ameliorated DSS-induced ultrastructural changes of the colon epithelium, as observed under scanning electron microscopy, as well as FITC-D permeability. The m RNA and protein expressions of TJ protein, zo-1, occludin and claudin-1 were also found to be significantly enhanced in the poly I:C group, as determined by immunohistochemistry/immunofluorescence, Western blot and RT-q PCR. By contrast, poly I:C pretreatment markedly reversed the DSS-induced up-regulated expressions of the inflammatory cytokines TNF-α, IL-17 and IFN-γ.CONCLUSION Our study suggested that poly I:C may protect against DSS-induced colitis through maintaining integrity of the epithelial barrier and regulating innate immune responses, which may shed light on the therapeutic potential of poly I:C in human colitis.

Omeprazole decreases magnesium transport across Caco-2 monolayers

AIM:To elucidate the effect and underlying mechanisms of omeprazole action on Mg 2+ transport across the intestinal epithelium.METHODS:Caco-2 monolayers were cultured in various dose omeprazole-containing media for 14 or 21 d before being inserted into a modified Ussing chamber apparatus to investigate the bi-directional Mg 2+ transport and electrical parameters.Paracellular permeability of the monolayer was also observed by the dilution potential technique and a cation permeability study.An Arrhenius plot was performed to elucidate the activation energy of passive Mg 2+ transport across the Caco-2 monolayers.RESULTS:Both apical to basolateral and basolateral to apical passive Mg 2+ fluxes of omeprazole-treated epithelium were decreased in a dose-and time-dependent manner.Omeprazole also decreased the paracellular cation selectivity and changed the paracellular selective permeability profile of Caco-2 epithelium to Li +,Na +,K +,Rb +,and Cs + from seriesⅦto seriesⅥof the Eisenman sequence.The Arrhenius plot revealed the higher activation energy for passive Mg 2+ transport in omeprazoletreated epithelium than that of control epithelium,indicating that omeprazole affected the paracellular channel of Caco-2 epithelium in such a way that Mg 2+ movement was impeded.CONCLUSION:Omeprazole decreased paracellular cation permeability and increased the activation energy for passive Mg 2+ transport of Caco-2 monolayers that led to the suppression of passive Mg 2+ absorption.

Role of pregnane X-receptor in regulating bacterial translocation in chronic liver diseases

Bacterial translocation(BT) has been impeccably implicated as a driving factor in the pathogenesis of a spectrum of chronic liver diseases(CLD). Scientific evidence accumulated over the last four decades has implied that the disease pathologies in CLD and BT are connected as a loop in the gut-liver axis and exacerbate each other. Pregnane X receptor(PXR) is a ligandactivated transcription factor and nuclear receptor that is expressed ubiquitously along the gut-liver-axis. PXR has been intricately associated with the regulation of various mechanisms attributed in causing BT. The importance of PXR as the mechanistic linker molecule in the gutliver axis and its role in regulating bacterial interactions with the host in CLD has not been explored. Pub Med was used to perform an extensive literature search using the keywords PXR and bacterial translocation, PXR and chronic liver disease including cirrhosis. In an adequate expression state, PXR acts as a sensor for bile acid dysregulation and bacterial derived metabolites, and in response shapes the immune profile beneficial to the host. Activation of PXR could be therapeutic in CLD as it counter-regulates endotoxin mediated inflammation and maintains the integrity of intestinal epithelium. This review mainly focuses PXR function and its regulation in BT in the context of chronic liver diseases.

Effect of Clostridium perfringens enterotoxin on gastric cancer cells SGC7901 which highly expressed claudin-4 protein

AIM To investigate the effects of Clostridium perfringens enterotoxin(CPE) on gastric cancer cells which highly expressed claudin-4(CL4) protein.METHODS In this study, we detected expression of CL4 protein in different gastric cancer cell lines. Then, we investigated the effects of CPE on SGC7901 cells which highly expressed CL4 protein and the effects of CPE on sub-cutaneous tumor in nude mice models.RESULTS CL4 are highly expressed in SGC7901 cells. CPE expressedsignificant cytotoxicity in SGC7901 cells. Suppression of CL4 expression significantly decreased CPE-mediated cytotoxicity. CPE also inhibited tumor growth in subcutaneous tumor xenograft models.CONCLUSION CPE showed CL4 mediated cytotoxicity on gastric cancer cells SGC7901 and inhib-ited tumor growth in nude mice models.

Fluctuation of zonulin levels in blood vs stability of antibodies

AIM To evaluate the measurement of zonulin level and antibodies of zonulin and other tight junction proteins in the blood of controls and celiac disease patients.METHODS This study was conducted to assess the variability or stability of zonulin levels vs Ig A and Ig G antibodies against zonulin in blood samples from 18 controls at 0, 6, 24 and 30 h after blood draw. We also measured zonulin level as well as zonulin, occludin, vinculin, aquaporin 4 and glial fibrillary acidic protein antibodies in the sera of 30 patients with celiac disease and 30 controls using enzyme-linked immunosorbent assay methodology. RESULTS The serum zonulin level in 6 out of 18 subjects was low or < 2.8 ng/m L and was very close to the detection limit of the assay. The other 12 subjects had zonulin levels of > 2.8 ng/m L and showed significant fluctuation from sample to sample. Comparatively, zonulin antibody measured in all samples was highly stable and reproducible from sample to sample. Celiac disease patients showed zonulin levels with a mean of 8.5 ng/m L compared to 3.7 ng/m L in controls(P < 0.0001). Elevation of zonulin level at 2SD above the mean was demonstrated in 37% of celiac disease patients, while antibodies against zonulin, occludin and other tight junction proteins was detected in up to 86% of patients with celiac disease. CONCLUSION Due to its fluctuation, a single measurement of zonulin level is not recommended for assessment of intestinal barrier integrity. Measurement of Ig G and Ig A antibodies against zonulin, occludin, and other tight junction proteins is proposed for the evaluation of the loss of intestinal barrier integrity.

Gastro protective properties of the novel prostone SPI-8811 against acid-injured porcine mucosa

AIM:To evaluate the protective properties of novel prostone ClC-2 agonist SPI-8811 in porcine model of gastric acid injury. METHODS:Porcine gastric mucosa was mounted in Ussing chambers and injured by bathing mucosal tissues in an HCl Ringer's solution (pH = 1.5) with or without SP1-8811 (1 μmol/L), cystic fibrosis transmem-brane conductance regulator (CFTR) inhibitor (inhibitor 172, 10 μmol/L, apical) and ClC-2 inhibitor ZnCl 2 , 300 μmol/L, apical), on the apical surface of tissues. Transepithelial resistance and mucosal-to-serosal 3 H-mannitol fluxes were measured over a 90-min period. Tissues were analyzed by morph metric techniques, Immunofluorescence and by western blots. RESULTS:Compared with control tissues, acid exposure decreased transepithelial electrical resistance (TER) and increased 3 H-mannitol flux. Pretreatment of gastric mucosa with SPI-8811 was protective against acid-induced decreases in TER (TER, 50 Ω . cm 2 vs 100 Ω . cm 2 ) and abolished increases in flux ( 3 H-mannitol flux, 0.10 μmol/L . cm 2 vs 0.04 μmol/L . cm 2 ). Evidence of histological damage in the presence of acid was markedly at- tenuated by SPI-0811. Immunofluorescence and western analysis for occludin revealed enhanced localization to the region of the tight junction (TJ) after treatment with SPI-8811. Pretreatment with the ClC-2 inhibitor ZnCl 2 , but not the selective CFTR inhibitor 172, attenuated SPI-8811-mediated mucosal protection, suggesting a role for ClC-2. Prostone may serve both protective and reparative roles in injured tissues. CONCLUSION: ClC-2 agonist SPI-8811 stimulated enhancement of mucosal barrier function by protecting TJ protein occludin in porcine gastric mucosa and thus protected the gastric acid injury in porcine stomach.