Tight junction physiology of pleural mesothelium

Pleura consists of visceral and parietal cell layers, producing a fluid, which is necessary for lubrication of the pleural space. Function of both mesothelial cell layers is necessary for the regulation of a constant pleural fluid volume and composition to facilitate lung movement during breathing. Recent studies have demonstrated that pleural mesothelial cells show a distinct expression pattern of tight junction proteins which are known to ubiquitously determine paracellular permeability. Most tight junction proteins provide a sealing function to epithelia, but some have been shown to have a paracellular channel function or ambiguous properties. Here we provide an in- depth review of the current knowledge concerning specific functional contribution of these proteins determining transport and barrier function of pleural mesothelium

Cannabidiol attenuates inflammatory impairment of intestinal cells expanding biomaterial-based therapeutic approaches

Cannabis-based biomaterials have the potential to deliver anti-inflammatory therapeutics specifically to desired cells, tissues, and organs, enhancing drug delivery and the effectiveness of anti-inflammatory treatment while minimizing toxicity. As a major component of Cannabis, Cannabidiol (CBD) has gained major attention in recent years because of its potential therapeutic properties, e.g., for restoring a disturbed barrier resulting from inflammatory conditions. The aim of this study was to test the hypothesis that CBD has beneficial effects under normal and inflammatory conditions in the established non-transformed intestinal epithelial cell model IPEC-J2. CBD induced a significant increase in transepithelial electrical resistance (TER) values and a decrease in the paracellular permeability of [³H]-D-Mannitol, indicating a strengthening effect on the barrier. Under inflammatory conditions induced by tumor necrosis factor alpha (TNFα), CBD stabilized the TER and mitigated the increase in paracellular permeability. Additionally, CBD prevented the barrier-disrupting effects of TNFα on the distribution and localization of sealing TJ proteins. CBD also affected the expression of TNF receptors. These findings demonstrate the potential of CBD as a component of Cannabis-based biomaterials used in the development of novel therapeutic approaches against inflammatory pathogenesis

Cellular Distribution Pattern of tjp1 (ZO-1) in Xenopus laevis Oocytes Heterologously Expressing Claudins

Epithelial barriers constitute a fundamental requirement in every organism, as they allow the separation of different environments and set boundaries against noxious and other adverse effectors. In many inflammatory and degenerative diseases, epithelial barrier function is impaired because of a disturbance of the paracellular seal. Recently, the Xenopus laevis oocyte has been established as a heterologous expression model for the analysis of transmembrane tight junction protein interactions and is currently considered to be a suitable screening model for barrier effectors. A prerequisite for this application is a physiological anchoring of claudins to the cytoskeleton via the major scaffolding protein tjp1 (tight junction protein 1, ZO-1). We have analyzed the oocyte model with regard to the interaction of heterologously expressed claudins and tjp1. Our experiments have revealed endogenous tjp1 expression in protein and mRNA analyses of unfertilized Xenopus laevis oocytes expressing human claudin 1 (CLDN1) to claudin 5 (CLDN5). The amphibian cell model can therefore be used for the analysis of claudin interactions

Concerted action of berberine in the porcine intestinal epithelial model IPEC-J2: Effects on tight junctions and apoptosis

The plant alkaloid berberine has been shown to have many beneficial effects on human health. This has led to its use as a treatment for various cancer types, obesity, and diabetes. Moreover, a described barrier-strengthening effect in human cancer cell lines indicates that it might be useful for the treatment of inflammatory bowel disease. Detailed information regarding its effects on intestinal epithelium remains limited. In our current study, we describe the impact of berberine on a non-transformed porcine small intestinal epithelial cell model, IPEC-J2. Incubation of IPEC-J2 monolayers with berberine revealed dose- and time-dependent effects on barrier properties. A viability assay confirmed the specific effect of berberine on the apoptotic pathway, paralleled by the internalization of the sealing tight-junction (TJ) proteins claudin-1, claudin-3, and occludin within 6 h. Hence, the barrier function of the cells was reduced, as shown by the reduced transepithelial electrical resistance and the increased [3H]-D-Mannitol flux. A decrease of claudin-1, claudin-3, and occludin expression was also observed after 24 h, whereas ZO-1 expression was not significantly changed. These data indicate an early effect on both cell viability and barrier integrity, followed by a general effect on TJ architecture. The intracellular co-localization of claudin-1 and occludin or claudin-3 and occludin points to an initial induction of apoptosis accompanied by the internalization of sealing TJ proteins. Although barrier strengthening has been reported in cancerogenic epithelial models, our results show a barrier-weakening action, which represents a new aspect of the effect of berberine on epithelia. These results agree with the known toxic potential of plant alkaloids in general and show that berberine is also capable of exerting adverse effects in the intestinal epithelium

Molecular Characterization of Barrier Properties in Follicle-Associated Epithelium of Porcine Peyer's Patches Reveals Major Sealing Function of Claudin-4

The pig represents a preferred model for the analysis of intestinal immunology. However, the barrier of the follicle-associated epithelium (FAE) covering porcine Peyer's patches (PP) has not yet been characterized in detail. This study aimed to perform this characterization in order to pave the way toward an understanding of the functional contribution of epithelial barrier properties in gut immunology. Porcine tissue specimens were taken from the distal small intestine in order to obtain electrophysiological data of PP FAE and neighboring villous epithelium (VE), employing the Ussing chamber technique. Transepithelial resistance (TER) and paracellular fluorescein flux were measured, and tissues were morphometrically compared. In selfsame tissues, expression and localization of major tight junction (TJ) proteins (claudin-1, -2, -3, -4, -5, and -8) were analyzed. PP FAE specimens showed a higher TER and a lower apparent permeability for sodium fluorescein than VE. Immunoblotting revealed an expression of all claudins within both epithelia, with markedly stronger expression of the sealing TJ protein claudin-4 in PP FAE compared with the neighboring VE. Immunohistochemistry confirmed the expression and localization of all claudins in both PP FAE and VE, with stronger claudin-4 abundance in PP FAE. The results are in accordance with the physiological function of the FAE, which strongly regulates and limits antigen uptake determining a mandatory transcellular route for antigen presentation, highlighting the importance of this structure for the first steps of the intestinal immune response. Thus, this study provides detailed insights into the specific barrier properties of the porcine FAE covering intestinal PP, at the interface of intestinal immunology and barriology

Effects of 1,2-Dimethylhydrazine on Barrier Properties of Rat Large Intestine and IPEC-J2 Cells

Colon cancer is accompanied by a decrease of epithelial barrier properties, which are determined by tight junction (TJ) proteins between adjacent epithelial cells. The aim of the current study was to analyze the expression of TJ proteins in a rat model of 1,2-dimethylhydrazine (DMH)-induced colorectal cancer, as well as the barrier properties and TJ protein expression of IPEC-J2 cell monolayers after incubation with DMH. Transepithelial electrical resistance and paracellular permeability for sodium fluorescein of IPEC-J2 were examined by an epithelial volt/ohm meter and spectrophotometry. The expression and localization of TJ proteins were analyzed by immunoblotting and immunohistochemistry. In the colonic tumors of rats with DMH-induced carcinogenesis, the expression of claudin-3 and -4 was significantly increased compared to controls. The transepithelial electrical resistance of IPEC-J2 cells increased, while paracellular permeability for sodium fluorescein decreased, accompanied by an increased expression of claudin-4. The increase of claudin-4 in rat colon after chronic DMH exposure was consistent with the acute effect of DMH on IPEC-J2 cells, which may indicate an essential role of this protein in colorectal cancer development

Effects of glucagon‐like peptides 1 and 2 and epidermal growth factor on the epithelial barrier of the rumen of adult sheep

Epidermal growth factor (EGF) and glucagon‐like peptides (GLP) modulate the tight junctions (TJ) of the intestinal epithelial barrier (EB) of monogastric animals. This work tried to elucidate whether GLP‐1, GLP‐2 and EGF can affect the EB of the rumen. Ovine ruminal epithelia were incubated in Ussing chambers for 7 hr with 25 or 250 nM of either GLP‐1 or GLP‐2 on the serosal side, with 2.5 nM of EGF on the serosal side or with 0.25 or 2.5 nM EGF on the mucosal side. No treatment affected tissue conductance. Short‐circuit current (Isc) was affected by time and treatment and their interactions. Only 250 nM of either GLP‐1 or GLP‐2 decreased Isc in certain periods compared with 25 nM GLP‐1 or 0.25 nM mucosally applied EGF; however, not when compared to control epithelia. Fluorescein flux rates (Jfluor) of ruminal epithelia were affected by treatment, time and time × treatment interaction. The time × treatment interaction was based on an increase in Jfluor between the first and last hour in epithelia incubated with 25 nM GLP‐1 or GLP‐2 and in epithelia incubated with EGF. After 7 hr incubation, claudin‐7 mRNA expression was downregulated in all treatments. Claudin‐1 mRNA was upregulated after incubation with 2.5 nM EGF on the serosal side, claudin‐4 mRNA was downregulated by 2.5 nM EGF on the mucosal side, and occludin mRNA was increased after incubation with 250 nM GLP‐2. The protein abundance of all tested TJ proteins was not influenced by treatment. We conclude that GLP‐1, GLP‐2, and EGF have no obvious acute effects on the EB of ruminal epithelia under simulated physiological conditions ex vivo. However, by decreasing the mRNA expression of claudin‐7 and partly affecting other TJ proteins, they may modulate EB in the longer term or under certain conditions

Blood-brain barrier protein claudin-5 expressed in paired Xenopus laevis oocytes mediates cell-cell interaction

Claudin-5 determines the sealing properties of blood-brain barrier tight junctions and its function is impaired in neurodegenerative and neuroinflammatory disorders. Focusing on the contribution of claudin-5 to the trans-interaction within the tight junction seal, we used Xenopus laevis oocytes as an expression system. Cells were clustered and challenged in a novel approach for the analysis of claudin interaction. We evaluated the strengthening effect of claudin-5 to cell-cell-connection in comparison to claudin-3. Application of a hydrostatic pressure impulse on clustered control oocyte pairs revealed a reduction of contact areas. In contrast, combinations with both oocytes expressing claudins maintained an enhanced connection between the cells (cldn5–cldn5, cldn3–cldn3). Strength of interaction was increased by both claudin-3 and claudin-5. This novel approach allowed an analysis of single claudins contributing to tight junction integrity, characterizing homophilic and hetrophilic trans-interaction of claudins. To test a new screening approach for barrier effectors, exemplarily, this 2-cell model of oocytes was used to analyze the effect of the absorption enhancer sodium caprate on the oocyte pairs

Selective Role of TNFα and IL10 in Regulation of Barrier Properties of the Colon in DMH-Induced Tumor and Healthy Rats

Recently it has been reported that the tumor adjacent colon tissues of 1,2-dymethylhydrazine induced (DMH)-rats revealed a high paracellular permeability. We hypothesized that the changes might be induced by cytokines. Colorectal cancer is accompanied by an increase in tumor necrosis factor alpha (TNFα) and interleukin 10 (IL10) that exert opposite regulatory effects on barrier properties of the colon, which is characterized by morphological and functional segmental heterogeneity. The aim of this study was to analyze the level of TNFα and IL10 in the colon segments of DMH-rats and to investigate their effects on barrier properties of the proximal and distal parts of the colon in healthy rats. Enzyme immunoassay analysis showed decreased TNFα in tumors in the distal part of the colon and increased IL10 in proximal tumors and in non-tumor tissues. Four-hour intraluminal exposure of the colon of healthy rats with cytokines showed reduced colon barrier function dependent on the cytokine: TNFα decreased it mainly in the distal part of the colon, whereas IL10 decreased it only in the proximal part. Western blot analysis revealed a more pronounced influence of IL10 on tight junction (TJ) proteins expression by down-regulation of the TJ proteins claudin-1, -2 and -4, and up-regulation of occludin only in the proximal part of the colon. These data may indicate a selective role of the cytokines in regulation of the barrier properties of the colon and a prominent role of IL10 in carcinogenesis in its proximal part