Search

Introduction: Tight junctions (TJs) and their constituent proteins play pivotal roles in cellular physiology and anatomy by establishing functional boundaries within and between neighboring cells. While the involvement of TJ proteins, such as claudins, in cancer is extensively studied, studies highlighting their interaction with immune system are still meager. Studies indicate that alterations in cytokines and immune cell populations can affect TJ proteins, compromising TJ barrier function and exacerbating pro-inflammatory conditions, potentially leading to epithelial cell malignancy. Disrupted TJs in established tumors may foster a pro-tumor immune microenvironment, facilitating tumor progression, invasion, epithelial-to-mesenchymal transition and metastasis. Although previous literature contains many studies describing the involvement of TJs in pathogenesis of malignancies their role in modulating the immune microenvironment of tumors is just beginning to be unleashed., Areas Covered: This article for the first time attempts to discern the importance of interaction between TJs and immune microenvironment in malignancies. To achieve the above aim a thorough search of databases like PubMed and Google Scholar was conducted to identify the recent and relevant articles on the topic., Expert Opinion: Breaking the vicious cycle of dysbiosis/infections/chemical/carcinogen-induced inflammation-TJ remodeling-malignancy-TJ dysregulation-more inflammation can be used as a strategy to complement the effect of immunotherapies in various malignancies.

The structural and functional destruction of the blood-testis barrier (BTB) following uropathogenic E. coli (UPEC) infection may be a critical component of the pathologic progress of orchitis. Recent findings indicate that the mammalian target of the rapamycin (mTOR)-signaling pathway is implicated in the regulation of BTB assembly and restructuring. To explore the mechanisms underlying BTB damage induced by UPEC infection, we analyzed BTB integrity and the involvement of the mTOR-signaling pathway using in vivo and in vitro UPEC-infection models. We initially confirmed that soluble virulent factors secreted from UPEC trigger a stress response in Sertoli cells and disturb adjacent cell junctions via down-regulation of junctional proteins, including occludin, zonula occludens-1 (ZO-1), F-actin, connexin-43 (CX-43), β-catenin, and N-cadherin. The BTB was ultimately disrupted in UPEC-infected rat testes, and blood samples from UPEC-induced orchitis in these animals were positive for anti-sperm antibodies. Furthermore, we herein also demonstrated that mTOR complex 1 (mTORC1) over-activation and mTORC2 suppression contributed to the disturbance in the balance between BTB "opening" and "closing." More importantly, rapamycin (a specific mTORC1 inhibitor) significantly restored the expression of cell-junction proteins and exerted a protective effect on the BTB during UPEC infection. We further confirmed that short-term treatment with rapamycin did not aggravate spermatogenic degeneration in infected rats. Collectively, this study showed an association between abnormal activation of the mTOR-signaling pathway and BTB impairment during UPEC-induced orchitis, which may provide new insights into a potential treatment strategy for testicular infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lu, Liu, Tursi, Yan, Cao, Che, Yang and Dong.)

Background: Tumor protein p63 has been shown to be important for epithelial dysfunction, including epithelial barrier defects and mucosal inflammation, in the development of chronic rhinosinusitis with nasal polyps (CRSwNP). Basonuclin1 (BNC1), an epithelial-specific transcriptional factor, is a direct downstream target of p63 and thus might be involved in the pathogenesis of CRSwNP., Objective: We sought to investigate whether BNC1 was associated with p63-mediated epithelial barrier defects and nasal mucosal inflammation in CRSwNP., Methods: Nasal tissue biopsies were obtained from 91 patients to CRSwNP, 49 chronic rhinosinusitis without nasal polyps (CRSsNP) patients, and 28 control subjects. Immunohistochemistry and immunofluorescence staining were used to determine the distribution of BNC1 in tissues and localization in cells, respectively. Quantitative PCR was performed to detect the expression levels of BNC1, TP63, epithelial barrier proteins, and type-2 helper T-cell inflammation-related genes., Results: BNC1 mRNA expression was significantly elevated in the tissues in CRSwNP patients compared with CRSsNP (1.96-fold, p = 0.0003) and control groups (2.40-fold, p < 0.0001). BNC1 staining was strongly positive in the nasal epithelium and co-localized with p63-positive epithelial cells. The expression of BNC1 mRNA was strongly correlated with TP63 mRNA level both in tissue biopsies (r = 0.78, p < 0.0001) and epithelial scrapings (r = 0.97, p < 0.0001). BNC1 expression was also positively correlated with epithelial barrier protein genes (CDH1, CLDN1, CLDN4, TJP1, and TJP2) and epithelial genes involved in TH2 inflammation (IL33, CCL26, CLC, and ALOX15)., Conclusions: Overexpression of BNC1 may be associated with increased expression of TP63, and possibly contribute to the epithelial barrier defects and TH2 inflammation in CRSwNP., (© 2021 S. Karger AG, Basel.)

Protein arginine deiminase 4 (PAD4) serves a critical role in differentiation, development and apoptosis through gene regulation and has emerged as a potential therapeutic target for the treatment of various diseases. However, the roles of PAD4 in lipopolysaccharide (LPS)-induced acute lung injury (ALI) remain largely unknown. To investigate the roles of PAD4 during LPS-induced ALI, the present study detected the trend of PAD4 expression in the lung tissues of ALI mice. Subsequently, the efficiency of TDFA on PAD4 and citrullinated H3 histone were detected. And then, histology, the wet/dry weight ratio, survival rate, activated cells infiltration, oxidative stress levels, tight junction proteins and proinflammatory cytokine expression were detected. In addition, the level of transepithelial electrical resistance (TEER) was assessed. Finally, the level of nuclear P65, total phosphorylated P65 and P65 were measured in vivo and in vitro. The results showed that PAD4 expression was upregulated in the lung tissues of LPS-induced ALI. TDFA efficiently decreased the severity of the lung edema, attenuated the severity of pulmonary injury and improved the survival rate following lethal LPS administration. Besides, TDFA reduced activated cells infiltration and suppressed inflammation related parameters, including proinflammatory cytokines production (TNF-α, IL-6 and IL-1β) and oxidative stress (MDA, GSH and SOD). Furthermore, TDFA reversed the TEER downregulation tendency and tight junction proteins (ZO-1, Occludin, Claudin-4) levels that represent the integrity of alveolar epithelium. Eventually, TDFA exerts its protective roles through modulating nuclear localization of transcription factor NF-κB P65 in epithelial cells. Taken together, these results indicate that PAD4 inhibition may serve as a promising therapeutic approach for LPS-induced ALI., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interests regarding the publication of this paper., (Copyright © 2020. Published by Elsevier B.V.)

Sesamol, a liposoluble lignan extract, has already been proved to possess potent anti-inflammatory properties, and it could also regulate gut dysfunction. The purpose of the present research is to explore the protective effect of sesamol on colitis mice. In the current research, sesamol treatment (100 mg/kg bodyweight/day) for 6 weeks inhibited the dextran sulphate sodium (DSS)-induced bodyweight loss of mice. Transmission electron microscopy and hematoxylin and eosin staining results showed that the DSS-induced histopathological changes of mice were also recovered by sesamol supplementation. In addition, DSS-induced inflammatory responses were inhibited by sesamol supplementation via the NF-κB signaling pathway in mice colon. Moreover, sesamol treatment prevented gut barrier damages by enhancing the expression of tight junction proteins (occludin, claudin-1, and ZO-1) and recovering the loss of gut mucus layer. Furthermore, sesamol supplementation also increased the short-chain fatty acid (SCFAs) contents of acetate, propionate, and butyrate. Furthermore, sesamol supplementation changed the gut microbiome structure by enhancing the relative abundance of Coprococcuscus , Butyricicoccus , Odoribacter , and AF12 in colitis mice. In conclusion, sesamol could effectively ameliorate DSS-induced colitis by promoting gut microecology.

Long-term intake of a high-fat diet seriously affects the health of pregnant women and leads to increased levels of inflammation in the mammary gland. Therefore, to further explore the effect of a high-fat diet on mammary gland development and the tight junction (TJ) during pregnancy, we placed mice into two groups: a high-fat diet group and a control group. We detected the expression of proteins related to fat synthesis in the mammary gland by western blotting. The results showed that a high-fat diet could lead to an increase in fat synthesis in the mammary gland. Then, the inflammatory levels and acinar cell morphology in the mammary gland were detected by ELISA and H&E staining. We also measured the levels of MAPK and NF-κB signal pathway-related proteins by western blotting. The results showed that a high-fat diet activated the MAPK and NF-κB signaling pathways and promoted the expression of inflammatory factors. Finally, the development of the mammary gland and the integrity of the TJ were determined by immunohistochemistry, immunofluorescence and western blotting. The results showed that a high-fat diet inhibited the development of the mammary gland and the expression of tight junction proteins (TJs). Our study showed that a high-fat diet could promote the expression of inflammatory factors by activating the MAPK and NF-κB signaling pathways and could reshape the microenvironment through extramammary inflammation. Finally, a high-fat diet inhibited the development of the mammary gland during pregnancy and destroyed the integrity of the TJ.

Ginsenosides have a variety of pharmacological activities, including immunomodulatory, antitumor and anti-inflammatory activities. However, the effect of Rk3 on ulcerative colitis has rarely been reported. This study evaluated the effect of Rk3 on DSS-induced ulcerative colitis and preliminarily explored the anti-inflammatory mechanisms. Rk3 administration significantly attenuated the weight loss, increased DAI scores, colonic shortening, and increased MPO and iNOS activities caused by DSS in mice. Histological improvement was apparent, tight junctions in the colon were restored, and the levels of short-chain fatty acids (acetic acid, butyric acid and isovaleric acid) were increased. In addition, Rk3 reduced the expression of proinflammatory factors (TNF-α, IL-1β and IL-6), NLRP3, ASC, and Caspase-1, indicating blockade of the NLRP3 inflammasome pathway. These results show that Rk3 can improve DSS-induced ulcerative colitis by protecting intestinal barrier function and inhibiting NLRP3 inflammasome expression, indicating that Rk3 could be used as a potential drug for treating ulcerative colitis., Competing Interests: Declaration of Competing Interest The authors declare there are no conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The effects of skin wounds on the intestinal barrier function and the beneficial effects of the dietary administration of Shewanella putrefaciens (known as SpPdp11) in gilthead seabream (Sparus aurata L.) were studied. Two replicates of fish were fed a commercial diet (control, CON) or CON diet enriched with 10 9  cfu g -1 SpPdp11 (SP diet) for 30 days. After this time, half of the fish were sampled, while the others were injured below the lateral line (wounded fish, W) and fed the same diets for an extra week before sampling (CON + W and SP + W groups). The intestinal histology and gene expression of different genes relevant for the intestinal barrier function were studied. The results showed that injured fish had a disordered enterocyte nucleus disposition, a more intense infiltration of mixed leucocytes and a thicker lamina propria in the intestine compared to the control fish. However, the fish in the SP + W group did not present these pathological symptoms in the intestine. No significant variations in the number of goblet cells were detected among the different experimental groups. Pro-inflammatory cytokines (colony-stimulating factor receptor 1, CSF1R, myeloperoxidase, MPO and interleukin-1β, IL-1β), mucins (intestinal mucin, IMUC and mucin 2, MUC2), and immunoglobulin T heavy chain (IGHT) were up-regulated, while tight junction protein occludin was down-regulated in the intestine from fish of the CON + W group. Similarly, the dietary administration of SpPdp11 markedly depressed the gene expression of pro-inflammatory cytokines, MUC2 and IGHT, but increased the gene expression of anti-inflammatory cytokine transforming growth factor-β1 (TGF-β1) and the tight junction proteins tricellulin and occluding after wounding. In brief, the skin wounds provoked an intestinal inflammatory response that included changes in the mucus layer and tight junction disruptions. Besides this, preventive administration of SpPdp11 alleviated the intestinal dysfunctions caused by skin wounds in gilthead seabream., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

High ambient temperature has multiple potential effects on the organism such as hyperthermia, endotoxemia, and/or systemic inflammation. However, it is often difficult to discriminate between cause and consequence of phenotypic effects, such as the indirect influence of heat stress via reduced food intake. Lactating dairy cows are a particularly sensitive model to examine the effects of heat stress due to their intensive metabolic heat production and small surface:volume ratio. Results from this model show heat stress directly induced a so-far unknown infiltration of yet uncategorized cells into the mucosa and submucosa of the jejunum. Due to a pair-feeding design, we can exclude this effect being a consequence of the concurrent heat-induced reduction in feed intake. Isolation and characterization of the infiltrating cells using laser capture microdissection and RNA sequencing indicated a myeloic origin and macrophage-like phenotype. Furthermore, targeted transcriptome analyses provided evidence of activated immune- and phagocytosis-related pathways with LPS and cytokines as upstream regulators directly associated with heat stress. Finally, we obtained indication that heat stress may directly alter jejunal tight junction proteins suggesting an impaired intestinal barrier. The penetration of toxic and bacterial compounds during heat stress may have triggered a modulated immune repertoire and induced an antioxidative defense mechanism to maintain homeostasis between commensal bacteria and the jejunal immune system. Our bovine model indicates direct effects of heat stress on the jejunum of mammals already at moderately elevated ambient temperature. These results need to be considered when developing concepts to combat the negative consequences of heat stress., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Ulcerative colitis is a life-long, chronic, relapsing and remitting inflammatory disease of the large intestine with an unpredictable course characterized by debilitating gastrointestinal symptoms accompanied by healthcare and emotional burdens that reduce the quality of life and the ability to work, attend school, and be productive. Ulcerative colitis affects millions of people worldwide and is now considered a global disease. Although some form of primary immune abnormality is thought to underlie this illness, extensive laboratory research conducted since the mid-20th century has largely failed to definitively establish a primary antecedent immune abnormality in individuals with ulcerative colitis or their family members. An alternative approach employing a systems pathogenesis analysis has implicated a causal role for colonocyte-generated hydrogen peroxide in the pathogenesis of this illness. Significantly elevated levels of hydrogen peroxide in non-inflamed colonic mucosa have been demonstrated in individuals with ulcerative colitis, implying a build-up prior to the onset of inflammation and supporting a causal role for colonocyte hydrogen peroxide in the development of this disease. Hydrogen peroxide's unique properties of cell membrane permeability, long life, potent oxidizing potential, and the ability to attract white blood cells combine to promote oxidative disintegration of colonic epithelial tight junctional proteins while attracting white blood cells into the colonic epithelium, both of which lead to colonic inflammation and eventual ulcerative colitis.

Strong tight junctions and curtailed inflammatory responses under stressful conditions are key for optimal digestive health. Bacillus -based probiotics are increasingly being used to maintain broilers' health, but their mode of action is often not well-defined. In the present study we used Caco-2 cells as a model for intestinal epithelia and assessed the effect of three Bacillus -based probiotics on intestinal barrier function and intestinal inflammation. Experimental results showed that one of the three tested strains, Bs 29784, significantly reinforced intestinal barrier integrity under basal conditions through an up-regulation of the expression of tight junction's proteins, whereas the others had no or detrimental effects. When Caco-2 cells were pre-treated with Bacillus subtilis strains, the subsequent IL-8 release to various pro-inflammatory signals (IL-1β, deoxynivalenol, or flagellin) was blunted compared to cells that had not been pretreated, but to a different extent depending on the strain of Bacillus used. Bs 29784, was able to significantly decrease IL-8 production in all stressed conditions tested. Mechanistically, Bs 29784 appeared to limit nuclear translocation of NF-κB during IL-1β exposure by preventing IκB degradation. The effects of Bs 29784 were observed independently with supernatant and cells but in a lesser extent than with the combination, indicating that they can thus likely be attributed to both secreted metabolites and cell-associated compounds. Moreover, under inflammatory conditions, Bs 29784 significantly reduced the upregulation of iNOS protein levels further underlining its intestinal anti-inflammatory potential. Our data show that Bacillus -based probiotics may indeed improve digestive health by strengthening intestinal barrier and limiting inflammatory responses and that these properties are strain-dependent.

Sphingosine 1-phosphate (S1P) and S1P receptors (S1PRs) regulate migration of lymphocytes out of thymus to blood and lymph nodes (LNs) to efferent lymph, whereas their role in other tissue sites is not known. Here, we investigated the question of how these molecules regulate leukocyte migration from tissues through afferent lymphatics to draining LNs (dLNs). S1P, but not other chemokines, selectively enhanced human and murine CD4 T cell migration across lymphatic endothelial cells (LECs). T cell S1PR1 and S1PR4, and LEC S1PR2, were required for migration across LECs and into lymphatic vessels and dLNs. S1PR1 and S1PR4 differentially regulated T cell motility and vascular cell adhesion molecule-1 (VCAM-1) binding. S1PR2 regulated LEC layer structure, permeability, and expression of the junction molecules VE-cadherin, occludin, and zonulin-1 through the ERK pathway. S1PR2 facilitated T cell transcellular migration through VCAM-1 expression and recruitment of T cells to LEC migration sites. These results demonstrated distinct roles for S1PRs in comodulating T cell and LEC functions in migration and suggest previously unknown levels of regulation of leukocytes and endothelial cells during homeostasis and immunity., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

The expression pattern of tight junction proteins (TJPs) varies among organs and tumor types. In this study, we examined the immunoreactivity of claudin (CLDN)-1, -4, and -7, and JAM-A in salivary gland tumors (SGTs) by histological types and cell types to estimate their usefulness as differential diagnostic markers. Immunoreactivity of CLDN1 was higher in ductal epithelium cells of SGTs than in non-tumor tissues. Conversely, immunoreactivity of CLDN1 was significantly decreased in basal/myoepithelium cells of SGTs compared with that in non-tumor tissues. There was no significant difference between the immunoreactivity of CLDN1 in benign tumors and that in malignant tumors. Immunoreactivity of CLDN4, CLDN7, and JAM-A in ductal epithelium cells was higher in many SGTs than in non-tumor tissues. There was a difference depending on the histological type of SGT in immunoreactivity of CLDN4, CLDN7, and JAM-A in basaloid/myoepithelial cells. It was possible to classify SGTs by a hierarchical clustering using immunoreactivity of TJPs. The results suggest that an immunohistochemical marker panel including these TJPs may be useful for differential diagnosis of SGTs and that CLDN1 is associated with tumorigenesis of SGTs.

It is well-known that essential oil thymol exhibits antibacterial activity. The protective effects of thymol on pig intestine during inflammation is yet to be investigated. In this study, an in vitro lipopolysaccharide (LPS)-induced inflammation model using IPEC-J2 cells was established. Cells were pretreated with thymol for 1 h and then exposed to LPS for various assays. Interleukin 8 (IL-8) secretion, the mRNA abundance of cytokines, reactive oxygen species (ROS), nutrient transporters, and tight junction proteins was measured. The results showed that LPS stimulation increased IL-8 secretion, ROS production, and tumor necrosis factor alpha (TNF-α) mRNA abundance ( P < 0.05), but the mRNA abundance of sodium-dependent glucose transporter 1 (SGLT1), excitatory amino acid transporter 1 (EAAC1), and H + /peptide cotransporter 1 (PepT1) were decreased ( P < 0.05). Thymol blocked ROS production ( P < 0.05) and tended to decrease the production of LPS-induced IL-8 secretion ( P = 0.0766). The mRNA abundance of IL-8 and TNF-α was reduced by thymol pretreatment ( P < 0.05), but thymol did not improve the gene expression of nutrient transporters ( P > 0.05). The transepithelial electrical resistance (TEER) was reduced and cell permeability increased by LPS treatment ( P < 0.05), but these effects were attenuated by thymol ( P < 0.05). Moreover, thymol increased zonula occludens-1 (ZO-1) and actin staining in the cells. However, the mRNA abundance of ZO-1 and occludin-3 was not affected by either LPS or thymol treatments. These results indicated that thymol enhances barrier function and reduce ROS production and pro-inflammatory cytokine gene expression in the epithelial cells during inflammation. The regulation of barrier function by thymol and LPS may be at post-transcriptional or post-translational levels.

The present work is undertaken to characterize a Granny Smith apple procyanidin extract (AE) and investigate the beneficial effect of the AE in the intestine in vitro. Each AE was characterized via LC-ESI-MS. Caco-2 cells were used to study the preventive actions of the AE against the downregulation of tight junction protein expression, oxidative stress and inflammation induced by lipopolysaccharides (LPS). Phenolic compounds present in the AE, including chlorogenic acid, catechin, epicatechin, proanthocyanidin dimers, and proanthocyanidin trimers, were characterized. The expression of the tight junction protein, including occludin and zona occludens (ZO)-1, increased significantly in LPS + AE treated Caco-2 cells, compared to LPS induced Caco-2 cells. Proanthocyanidin dimers had the most potent effect on increasing tight junction protein expression. The addition of LPS to Caco-2 cells induced oxidative stress and inflammation. However, incubation with proanthocyanidin dimers prevented LPS-mediated oxidative stress, including the increase of SOD, HO-1, CAT, and GSH-Px mRNA expression, and counteracted LPS-mediated inflammation as evidenced by the down-regulation of inflammatory markers (NF-κβ, IL-6, and TNF-α mRNA expression). Our findings provide evidence that AE could upregulate tight junction protein expression, probably acting via the reduction of oxidative stress and inflammation.

Blood-spinal cord barrier (BSCB) disruption is a major process for the secondary injury of spinal cord injury (SCI) and is considered to be a therapeutic target for SCI. Previously, we demonstrated that metformin could improve functional recovery after SCI; however, the effect of metformin on BSCB is still unknown. In this study, we found that metformin could prevent the loss of tight junction (TJ) proteins at day 3 after SCI in vivo, but in vitro there was no significant difference of these proteins between control and metformin treatment in endothelial cells. This indicated that metformin-induced BSCB protection might not be mediated by up-regulating TJ proteins directly, but by inhibiting TJ proteins degradation. Thus, we investigated the role of metformin on MMP-9 and neutrophils infiltration. Neutrophils infiltration is the major source of the enhanced MMP-9 in SCI. Our results showed that metformin decreased MMP-9 production and blocked neutrophils infiltration at day 1 after injury, which might be related to ICAM-1 down-regulation. Also, our in vitro study showed that metformin inhibited TNF-α-induced MMP-9 up-regulation in neutrophils, which might be mediated via an AMPK-dependent pathway. Together, it illustrated that metformin prevented the breakdown of BSCB by inhibiting neutrophils infiltration and MMP-9 production, but not by up-regulating TJ proteins expression. Our study may help to better understand the working mechanism of metformin on SCI., (© 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

The present study explored the possible preventive effects of dietary glutamate (Glu) on LPS-induced oxidative damage, mRNA expression changes of tight junction (TJ) and defensin proteins, inflammatory and apoptosis response signaling molecules in fish intestine. Young Jian carp were fed five diets supplemental graded levels of Glu (0, 4, 8, 16 and 32 g kg -1 diet) for 63 days. The results indicated that Glu supplementation depressed LPS induced the production of reactive oxygen species (ROS) and severe oxidative damage (lipid peroxidation and protein oxidation) in fish intestine, which was partially due to the increased glutathione (GSH) content and antioxidant enzyme activities including superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione-S-transferase (GST), and glutathione reductase (GR) (P < 0.05). Further investigations indicated that Glu supplementation caused elevation of those antioxidant enzyme activities are related to the up-regulation of corresponding antioxidant enzymes and the related signaling factor Nrf2 mRNA levels (P < 0.05). Meanwhile, Glu pre-treatment significantly suppressed LPS-induced COX-2 and inflammatory cytokines mRNA expression and down-regulated NF-κB p65 and MAPK p38 transcription. Furthermore, pre-treatment with Glu prevented LPS induced apoptosis-related gene expression (caspase 3 and 9, P < 0.05). Lastly, Glu supplementation also attenuated LPS induced intestinal barrier function-related gene TJ proteins (ZO-1, occludin1, claudin2, 3, and 7), β-defensin1 and 3 mRNA expressions decreasing (P < 0.05). Taken together, the present results showed Glu could attenuate LPS induced the oxidative damage by Nrf2 signal pathway and depress LPS induced inflammation response (cytokines, COX-2, NF-κB p65, and MAPK p38), apoptosis (caspase3 and 9), and barrier function (ZO-1, occludin1, claudin2, 3 and 7, and β-defensin 1 and 3)-related gene expression changes of fish intestine., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Background: Postnatal development of the mammalian mucosal immune system is crucial for responding to the rapid colonization by commensal bacteria and possible exposure to pathogens. This study analyzed expression patterns for mRNAs and their relationship with microRNAs (miRNAs) in the bovine small intestine during the critical neonatal period (0 to 42 days). This analysis revealed molecular mechanisms regulating the postnatal development of the intestinal mucosal immune system., Results: Small intestine samples (jejunum and ileum) were collected from newborn male, Holstein calves immediately post-partum (n = 3) and at 7 (n = 5), 21 (n = 5), and 42 (n = 5) days of age and the transcriptomes were profiled using RNA-Seq. When analyzing all time points collectively, greater expression of genes encoding the complement functional pathway, as well as lower expression of genes encoding Toll-like receptors and NOD-like receptors were observed in the jejunum when compared to the ileum. In addition, significant changes in the expression of immune-related genes were detected within the first week post-partum in both jejunum and ileum. For example, increased expression of genes encoding tight junction proteins (claudin 1, claudin 4 and occludin), an antimicrobial peptide (Regenerating Islet-Derived 3-γ), NOD-like receptors (NACHT, LRR and PYD domain-containing protein 3), regulatory T cell marker (forkhead box P3), and both anti-inflammatory (interleukin 10) and pro-inflammatory (interleukin 8) cytokines was observed throughout the small intestine of 7-day-old calves when compared to newborn calves. Moreover, the expression of mucosal immune-related genes were either positively or negatively correlated with total bacterial population depending on both intestinal region and age. The integrated analysis of miRNAs and mRNAs supported the conclusion that miRNAs may regulate temporal changes in the expression of genes encoding tight junction proteins (miR-335), cytokines (miR-335) and bacterial recognition (miR-100) during the first week of small intestine development., Conclusion: The rapid development of transcriptional differences between jejunum and ileum reveal that these two intestinal regions make distinct contributions to the intestinal mucosal immune system during the early neonatal period. In addition, transcriptome analysis indicates that the first week after birth is a very dynamic developmental period for the intestinal mucosal immune system and these changes may be regulated by both miRNAs and microbial colonization. Findings from this study indicate that a detailed analysis of both the abundance and diversity of the colonizing microbiome may be necessary to understand factors regulating the rapid development of the mucosal immune system during the first week of life.

Intestinal mucosal immune components and mRNA levels of inflammatory cytokines, tight junction proteins, antioxidant enzymes and related signalling molecules in young grass carp (Ctenopharyngodon idellus) under dietary manganese (Mn) deficiency or excess were investigated. Fish were fed the diets containing graded levels of Mn [3.65-27.86 mg Mn kg(-1) diet] for 8 weeks. The results demonstrated that Mn deficiency significantly decreased the lysozyme and acid phosphatase (ACP) activities, up-regulated tumour necrosis factor α (TNF-α), interleukin 8 and the signalling factor nuclear factor-κB p65, and down-regulated interleukin 10 (IL-10), transforming growth factor β1, inhibitor of signalling factors κB-α and target of rapamycin mRNA levels in the proximal intestine (PI), mid intestine (MI) and distal intestine (DI). However, Mn deficiency did not change the C3 content in the PI, whereas it decreased the C3 contents in the MI and DI. Additionally, Mn depletion also resulted in significantly low mRNA levels for tight junction proteins (claudin-b, claudin-c, claudin-15, occludin and zonula occludens-1), antioxidant enzymes (MnSOD, GPx and CAT) and NF-E2-related factor-2 in the intestines of fish. Excessive Mn exhibited toxic effects similar to Mn deficiency, where optimal Mn contents reversed those indicators. In conclusion, Mn deficiency or excess causes the depression of intestinal immunity, induction of inflammation and dysfunction of the intestinal physical barrier relating to NF-κB, TOR and Nrf2 signalling in grass carp. Furthermore, quadratic regression analysis at 95% maximum response of lysozyme and acid phosphatase activities in the distal intestine of young grass carp revealed the optimum dietary Mn levels to be 8.90 and 8.99 mg kg(-1) diet, respectively., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Papain is commonly used in food, pharmaceutical, textile, and cosmetic industries and is known to induce occupational allergic asthma. We have previously shown that the papain-like cysteine protease Dermatophagoides pteronyssinus 1 from house dust mite exhibits percutaneous sensitization potential. We aimed here to investigate the potential of papain itself in epicutaneous sensitization. The effects of papain on tight junction (TJ) proteins were tested in vitro in human primary keratinocytes. Using C57BL/6 wild-type and Toll-like receptor 4 (TLR4)-deficient mice, we analyzed the sensitization potential of papain, its effects on the skin barrier, and immune cell recruitment. Our results show that papain affects the skin barrier by increasing transepidermal water loss, degrading TJ proteins and inducing vasodilation. When topically applied, papain exhibited a high epicutaneous inflammatory potential by recruiting neutrophils, mast cells, and CD3-positive cells and by induction of a TH2-biased antibody response. However, its high potency for specific sensitization via the skin was TLR4 independent and, in spite of its capacity to degrade epidermal TJ proteins, does not rely on its enzymatic function. From our data, we conclude that papain has all features to act as a strong allergen via the skin.

Interleukin-34 (IL-34) is a newly discovered cytokine as an additional ligand for colony stimulating factor-1 receptor (CSF1R), and its functions are expected to overlap with colony stimulating factor-1/macrophage-colony stimulating factor. We have previously shown that the IL-34 is primarily produced by neurons in the central nervous system (CNS) and induces proliferation and neuroprotective properties of microglia which express CSF1R. However, the functions of IL-34 in the CNS are still elucidative. Here we show that CNS capillary endothelial cells also express CSF1R. IL-34 protected blood-brain barrier integrity by restored expression levels of tight junction proteins, which were downregulated by pro-inflammatory cytokines. The novel function of IL-34 on the blood-brain barrier may give us a clue for new therapeutic strategies in neuroinflammatory and neurodegenerative diseases such as multiple sclerosis and Alzheimer's disease.

The choroid plexus (CP) is considered to be a point of leukocyte entry into the CNS during normal immune surveillance and in neuroinflammatory diseases. The structural and functional alterations within the CP that support this migration are not understood. We used quantitative, high-resolution, 3-dimensional (3-D) fluorescence imaging to analyze CP alterations associated with inflammatory responses in C57/Bl6 mice after the induction of experimental autoimmune encephalomyelitis by immunization with myelin oligodendrocyte glycoprotein (MOG) and complete Freund adjuvant/pertussis toxin (MOG-CFA/PTX) or adjuvants alone (CFA-PTX). The MOG-CFA/PTX and CFA/PTX produced similar effects, although those caused by the former were consistently more marked. Both treatments resulted in the accumulation of serum immunoglobulin G and leukocytes in the CP stroma, consistent with elevated stromal capillary permeability. They also provoked distortions and diminished immunostaining patterns of the tight junction adaptor protein ZO-1 in the choroidal epithelium but no obvious change in the patterns of the tight junction associated protein claudin-2. Only MOG-CFA/PTX triggered visible extravasation of immunoglobulin G and leukocytes across the choroidal epithelium. Our results suggest that CFA/PTX primes the CP for neuroinflammation by inducing structural changes that are exacerbated when there is an immune response to MOG and reinforce the CP as a gateway for leukocytes to enter the CNS by accessing the CSF and leptomeninges.

Rationale: HIV-1-induced interstitial pneumonitis (IP) is a serious complication of HIV-1 infection, characterized by inflammation and cellular infiltration in lungs, often leading to respiratory failure and death. The barrier function of the pulmonary endothelium is caused in part by tight junction (TJ) proteins, such as claudin-5. Peroxisome proliferator-activated receptor (PPAR)-γ is expressed in lung tissues and regulates inflammation. We hypothesize that HIV-1 induces vascular lung injury, and HIV-1-mediated damage of the pulmonary endothelium and IP is associated with dysregulation of PPAR-γ., Objectives: Investigate the effects of HIV-1 infection on the pulmonary microvasculature and the modulatory effects of the PPAR-γ ligands., Methods: Using human lung tissues, we demonstrated down-regulation of claudin-5 (marker of pulmonary barrier integrity), down-regulation of PPAR-γ transcription, and expression in lung tissues of HIV-1-infected humans with IP., Measurements and Main Results: Human lung microvascular endothelial cells expressed the TJ proteins claudin-5, ZO-1, and ZO-2; HIV-1 decreased TJ proteins expression and induced nuclear factor-κB promoter activity, which was reversed by PPAR-γ agonist. Using two murine HIV/AIDS models, we demonstrated decreased claudin-5 expression and increased macrophage infiltration in the lungs of HIV-1-infected animals. Activation of PPAR-γ prevented HIV-1-induced claudin-5 down-regulation and significantly reduced viremia and pulmonary macrophage infiltration., Conclusions: HIV-induced IP is associated with injury to the lung vascular endothelium, with decreased TJ and PPAR-γ expression, and increased pulmonary macrophage infiltration. PPAR-γ ligands abrogated these effects. Thus, regulation of PPAR-γ can be a therapeutic approach against HIV-1-induced vascular damage and IP in infected humans. Removal of Expression of Concern: Issues leading to the previous expression of concern for this article have been resolved after further revisions and editorial review. No further concerns exist.

Background and Objective: Previously we demonstrated uniformly strong expression of CD24 in the epithelial attachment to the tooth and in the migrating epithelium of the periodontitis lesion. Titers of serum antibodies autoreactive with CD24 peptide correlated with reduced severity of periodontal disease. Ligation of CD24 expressed by oral epithelial cells induced formation of tight junctions that limited paracellular diffusion. In this study, we aimed to reveal that the lack of uniform expression of tight junction components in the pocket epithelium of periodontitis lesions is likely to contribute to increased paracellular permeability to bacterial products. This is proposed as a potential driver of the immunopathology of periodontitis., Material and Methods: An epithelial culture model with close correspondence for expression patterns for tight junction components in periodontal epithelia was used. Immunohistochemical staining and confocal laser scanning microscopy were used to analyse patterns of expression of gingival epithelial tight junction components., Results: The minimally inflamed gingival attachment was characterized by uniformly strong staining at cell contacts for the tight junction components zona occludens-1, zona occludens-2, occludin, junction adhesion molecule-A, claudin-4 and claudin-15. In contrast, the pocket epithelium of the periodontal lesion showed scattered, uneven staining for these components. This pattern correlated closely with that of unstimulated oral epithelial cells in culture. Following ligation of CD24 expressed by these cells, the pattern of tight junction component expression of the minimally inflamed gingival attachment developed rapidly., Conclusion: There was evidence for non-uniform and focal expression only of tight junction components in the pocket epithelium. In the cell-culture model, ligation of CD24 induced a tight junction expression profile equivalent to that observed for the minimally inflamed gingival attachment. Ligation of CD24 expressed by gingival epithelial cells by lectin-like receptors of commensal oral streptococci could mediate the phenotype of health, whereas pathogenic organisms associated with periodontal disease might not signal effectively through CD24., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Interferon-γ (IFN-γ) plays an important role in intestinal barrier dysfunction. However, the mechanisms are not fully understood. As hypoxia-inducible factor-1 (HIF-1) is a critical determinant response to hypoxia and inflammation, which has been shown to be deleterious to intestinal barrier function, we hypothesized that IFN-γ induces loss of barrier function through the regulation of HIF-1α activation and function. In this study, we detected the expressions of HIF-1α and tight junction proteins in IFN-γ-treated T84 intestinal epithelial cell line. IFN-γ led to an increase of HIF-1α expression in time- and dose-dependent manners but did not change the expression of HIF-1β. The IFN-γ-induced increase in HIF-1α was associated with an activation of NF-κB. Treatment with the NF-κB inhibitor, pyrolidinedithiocarbamate (PDTC), significantly suppressed the activation of NF-κB and the expression of HIF-1α. In addition, IFN-γ also increased intestinal epithelial permeability and depletion of tight junction proteins; inhibition of NF-κB or HIF-1α prevented the increase in intestinal permeability and alteration in tight junction protein expressions. Interestingly, we demonstrated that a significant portion of IFN-γ activation NF-kB and modulation tight junction expression is mediated through HIF-1α. Taken together, this study suggested that IFN-γ induced the loss of epithelial barrier function and disruption of tight junction proteins, by upregulation of HIF-1α expression through NF-κB pathway.

The tumorigenicity of human pluripotent stem cells is a major safety concern for their application in regenerative medicine. Here we identify the tight-junction protein Claudin-6 as a cell-surface-specific marker of human pluripotent stem cells that can be used to selectively remove Claudin-6-positive cells from mixed cultures. We show that Claudin-6 is absent in adult tissues but highly expressed in undifferentiated cells, where it is dispensable for human pluripotent stem cell survival and self-renewal. We use three different strategies to remove Claudin-6-positive cells from mixed cell populations: an antibody against Claudin-6; a cytotoxin-conjugated antibody that selectively targets undifferentiated cells; and Clostridium perfringens enterotoxin, a toxin that binds several Claudins, including Claudin-6, and efficiently kills undifferentiated cells, thus eliminating the tumorigenic potential of human pluripotent stem cell-containing cultures. This work provides a proof of concept for the use of Claudin-6 to eliminate residual undifferentiated human pluripotent stem cells from culture, highlighting a strategy that may increase the safety of human pluripotent stem cell-based cell therapies.