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Tight Junction Proteins (tight + junction_protein)

Distribution by Scientific Domains

Medical Sciences	53%
Life Sciences	40%

Selected Abstracts

Disruption of tight junction structure in salivary glands from Sjögren's syndrome patients is linked to proinflammatory cytokine exposure

ARTHRITIS & RHEUMATISM, Issue 5 2010
Patricia Ewert
Objective Disorganization of acinar cell apical microvilli and the presence of stromal collagen in the acinar lumen suggest that the labial salivary gland (LSG) barrier function is impaired in patients with Sjögren's syndrome. Tight junctions define cell polarity and regulate the paracellular flow of ions and water, crucial functions of acinar cells. This study was undertaken to evaluate the expression and localization of tight junction proteins in LSGs from patients with SS and to determine in vitro the effects of tumor necrosis factor , (TNF,) and interferon-, (IFN,) on tight junction integrity of isolated acini from control subjects. Methods Twenty-two patients and 15 controls were studied. The messenger RNA and protein levels of tight junction components (claudin-1, claudin-3, claudin-4, occludin, and ZO-1) were determined by semiquantitative reverse transcriptase,polymerase chain reaction and Western blotting. Tight junction protein localization was determined by immunohistochemistry. Tight junction ultrastructure was examined by transmission electron microscopy. Isolated acini from control subjects were treated with TNF, and IFN,. Results Significant differences in tight junction protein levels were detected in patients with SS. ZO-1 and occludin were strongly down-regulated, while claudin-1 and claudin-4 were overexpressed. Tight junction proteins localized exclusively to apical domains in acini and ducts of LSGs from controls. In SS patients, the ZO-1 and occludin the apical domain presence of decreased, while claudin-3 and claudin-4 was redistributed to the basolateral plasma membrane. Exposure of isolated control acini to TNF, and IFN, reproduced these alterations in vitro. Ultrastructural analysis associated tight junction disorganization with the presence of endocytic vesicles containing electron-dense material that may represent tight junction components. Conclusion Our findings indicate that local cytokine production in LSGs from SS patients may contribute to the secretory gland dysfunction observed in SS patients by altering tight junction integrity of epithelial cells, thereby decreasing the quality and quantity of saliva. [source]

Cell culture,produced hepatitis C virus does not infect peripheral blood mononuclear cells,

HEPATOLOGY, Issue 6 2008
Svetlana Marukian
Hepatitis C virus (HCV) replicates primarily in the liver, but HCV RNA has been observed in association with other tissues and cells including B and T lymphocytes, monocytes, and dendritic cells. We have taken advantage of a recently described, robust system that fully recapitulates HCV entry, replication and virus production in vitro to re-examine the issue of HCV infection of blood cell subsets. The HCV replicase inhibitor 2,C-methyl adenosine was used to distinguish HCV RNA replication from RNA persistence. Whereas cell culture,grown HCV replicated in Huh-7.5 hepatoma cells, no HCV replication was detected in B or T lymphocytes, monocytes, macrophages, or dendritic cells from healthy donors. No blood cell subset tested expressed significant levels of Claudin-1, a tight junction protein needed for HCV infection of Huh-7.5 cells. A B cell line expressing high levels of Claudin-1, CD81, and scavenger receptor BI remained resistant to HCV pseudoparticle infection. We bypassed the block in HCV entry by transfecting HCV RNA into blood cell subsets. Transfected RNA was not detectably translated and induced high levels of interferon-,. Supernatants from HCV RNA,transfected macrophages inhibited HCV replication in Huh-7.5 cells. Conclusion: We conclude that multiple blocks prevent blood cells from supporting HCV infection. (HEPATOLOGY 2008;48:1843-1850.) [source]

c-Jun N-terminal kinase is largely involved in the regulation of tricellular tight junctions via tricellulin in human pancreatic duct epithelial cells

JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2010
Takashi Kojima
Tricellulin (TRIC) is a tight junction protein at tricellular contacts where three epithelial cells meet, and it is required for the maintenance of the epithelial barrier. To investigate whether TRIC is regulated via a c-Jun N-terminal kinase (JNK) pathway, human pancreatic HPAC cells, highly expressed at tricellular contacts, were exposed to various stimuli such as the JNK activators anisomycin and 12- O -tetradecanoylphorbol 13-acetate (TPA), and the proinflammatory cytokines IL-1,, TNF,, and IL-1,. TRIC expression and the barrier function were moderated by treatment with the JNK activator anisomycin, and suppressed not only by inhibitors of JNK and PKC but also by siRNAs of TRIC. TRIC expression was induced by treatment with the PKC activator TPA and proinflammatory cytokines IL-1,, TNF,, and IL-1,, whereas the changes were inhibited by a JNK inhibitor. Furthermore, in normal human pancreatic duct epithelial cells using hTERT-transfected primary cultured cells, the responses of TRIC expression to the various stimuli were similar to those in HPAC cells. TRIC expression in tricellular tight junctions is strongly regulated together with the barrier function via the JNK transduction pathway. These findings suggest that JNK may be involved in the regulation of tricellular tight junctions including TRIC expression and the barrier function during normal remodeling of epithelial cells, and prevent disruption of the epithelial barrier in inflammation and other disorders in pancreatic duct epithelial cells. J. Cell. Physiol. 225: 720,733, 2010. © 2010 Wiley-Liss, Inc. [source]

A microfluidic bioreactor with integrated transepithelial electrical resistance (TEER) measurement electrodes for evaluation of renal epithelial cells

BIOTECHNOLOGY & BIOENGINEERING, Issue 4 2010
Nicholas Ferrell
Abstract We have developed a bilayer microfluidic system with integrated transepithelial electrical resistance (TEER) measurement electrodes to evaluate kidney epithelial cells under physiologically relevant fluid flow conditions. The bioreactor consists of apical and basolateral fluidic chambers connected via a transparent microporous membrane. The top chamber contains microfluidic channels to perfuse the apical surface of the cells. The bottom chamber acts as a reservoir for transport across the cell layer and provides support for the membrane. TEER electrodes were integrated into the device to monitor cell growth and evaluate cell,cell tight junction integrity. Immunofluorescence staining was performed within the microchannels for ZO-1 tight junction protein and acetylated ,-tubulin (primary cilia) using human renal epithelial cells (HREC) and MDCK cells. HREC were stained for cytoskeletal F-actin and exhibited disassembly of cytosolic F-actin stress fibers when exposed to shear stress. TEER was monitored over time under normal culture conditions and after disruption of the tight junctions using low Ca2+ medium. The transport rate of a fluorescently labeled tracer molecule (FITC-inulin) was measured before and after Ca2+ switch and a decrease in TEER corresponded with a large increase in paracellular inulin transport. This bioreactor design provides an instrumented platform with physiologically meaningful flow conditions to study various epithelial cell transport processes. Biotechnol. Bioeng. 2010;107:707,716. © 2010 Wiley Periodicals, Inc. [source]

Novel hepatic progenitor cell surface markers in the adult rat liver,

HEPATOLOGY, Issue 1 2007
Mladen I. Yovchev
Hepatic progenitor/oval cells appear in injured livers when hepatocyte proliferation is impaired. These cells can differentiate into hepatocytes and cholangiocytes and could be useful for cell and gene therapy applications. In this work, we studied progenitor/oval cell surface markers in the liver of rats subjected to 2-acetylaminofluorene treatment followed by partial hepatectomy (2-AAF/PH) by using rat genome 230 2.0 Array chips and subsequent RT-PCR, immunofluorescent (IF), immunohistochemical (IHC) and in situ hybridization (ISH) analyses. We also studied expression of the identified novel cell surface markers in fetal rat liver progenitor cells and FAO-1 hepatoma cells. Novel cell surface markers in adult progenitor cells included tight junction proteins, integrins, cadherins, cell adhesion molecules, receptors, membrane channels and other transmembrane proteins. From the panel of 21 cell surface markers, 9 were overexpressed in fetal progenitor cells, 6 in FAO-1 cells and 6 are unique for the adult progenitors (CD133, claudin-7, cadherin 22, mucin-1, ros-1, Gabrp). The specificity of progenitor/oval cell surface markers was confirmed by ISH and double IF analyses. Moreover, study of progenitor cells purified with Ep-CAM antibodies from D-galactosamine injured rat liver, a noncarcinogenic model of progenitor cell activation, verified that progenitor cells expressed these markers. Conclusion: We identified novel cell surface markers specific for hepatic progenitor/oval cells, which offers powerful tool for their identification, isolation and studies of their physiology and pathophysiology. Our studies also reveal the mesenchymal/epithelial phenotype of these cells and the existence of species diversity in the hepatic progenitor cell identity. (HEPATOLOGY 2007;45:139,149.) [source]

Evaluation of human nasal RPMI 2650 cells grown at an air,liquid interface as a model for nasal drug transport studies

JOURNAL OF PHARMACEUTICAL SCIENCES, Issue 3 2008
Shuhua Bai
Abstract This study tests the hypothesis that human nasal RPMI 2650 cells grown at an air,liquid interface is a feasible model for drug transport studies via the nasal route. RPMI 2650 cells were cultured in Eagle's minimal essential medium (MEM) at both air,liquid and liquid,liquid interfaces. For each culture regimen, monolayer integrity was tested by measuring the transepithelial resistance (TEER) as well as the transport of paracellular and transcellular markers across the monolayer. The expression of tight junction proteins,differentiation markers,in cells of the different monolayers was studied by western blot analysis and confocal microscopy. The highest TEER values (192,±,3 ,,·,cm2) were observed for RPMI 2650 cells seeded onto collagen-coated permeable polytetrafluoroethylene inserts and grown at an air,liquid interface for 10 days; a seeding density of 4,×,105/cm2 generated and maintained a cell monolayer with suitable barrier properties at days 9,12. Microscopic examination showed that RPMI 2650 cells grown on filter inserts formed a fully confluent monolayer. The apparent permeability coefficients of the paracellular marker, [14C] mannitol, and the transcellular marker, [3H] propranolol, were 5.07,±,0.01,×,10,6 cm/s and 16.1,±,0.1,×,10,6 cm/s, respectively. Western blot analysis indicated the presence of four tight junction proteins: ZO-1, occludin, claudin-1 and E-cadherin; and the quantities of ZO-1, occludin, and E-cadherin were significantly higher in cells grown at an air,liquid interface than in cells grown at a liquid,liquid interface. Confocal microscopic studies showed ZO-1, F-actin, occludin and claudin-1 proteins at cell-cell contacts and revealed significant differences in the distributions and densities of ZO-1 protein in cells grown at the two types of interface. The data indicate that RPMI 2650 cells grown at an air,liquid interface form polarized monolayers with the cells interconnected by tight junction proteins. This human nasal cell line model could provide a useful tool for in vitro screening of nasal drug candidates. © 2007 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 97:1165,1178, 2008 [source]

Transforming growth factor-, induces epithelial to mesenchymal transition by down-regulation of claudin-1 expression and the fence function in adult rat hepatocytes

LIVER INTERNATIONAL, Issue 4 2008
Takashi Kojima
Abstract Background/Aims: Transforming growth factor-, (TGF-,) initiates and maintains epithelial,mesenchymal transition (EMT), which causes disassembly of tight junctions and loss of epithelial cell polarity. In mature hepatocytes during EMT induced by TGF-,, changes in the expression of tight junction proteins and the fence function indicated that epithelial cell polarity remains unclear. Methods: In the present study, using primary cultures of adult rat hepatocytes at day 10 after plating, in which epithelial cell polarity is well maintained by tight junctions, we examined the effects of 0.01,20 ng/ml TGF-, on the expression of the integral tight junction proteins, claudin-1, -2 and occludin, as well as the fence function. Results: In adult rat hepatocytes, TGF-, induced EMT, which was indicated as upregulation of Smad-interacting protein-1 (SIP1) and Snail and down-regulation of E-cadherin. Down-regulation of claudin-1 and upregulation of occludin were observed beginning from a low dose of TGF-,, whereas upregulation of claudin-2 was observed at a high dose of TGF-,. Furthermore, treatment with TGF-, caused disruption of the fence function, which was closely associated with the expression of claudin-1 via p38 mitogen-activated protein kinase (MAPK), phosphoinositide-3 kinase and protein kinase C but not MAPK signalling pathways. Conclusion: These results suggest that in mature hepatocytes in vitro, TGF-, induces EMT by down-regulation of claudin-1 and the fence function via distinct signalling pathways. [source]

Differential distribution of tight junction proteins suggests a role for tanycytes in blood-hypothalamus barrier regulation in the adult mouse brain

THE JOURNAL OF COMPARATIVE NEUROLOGY, Issue 7 2010
Amandine Mullier
The median eminence is one of the seven so-called circumventricular organs. It is located in the basal hypothalamus, ventral to the third ventricle and adjacent to the arcuate nucleus. This structure characteristically contains a rich capillary plexus and features a fenestrated endothelium, making it a direct target of blood-borne molecules. The median eminence also contains highly specialized ependymal cells called tanycytes, which line the floor of the third ventricle. It has been hypothesized that one of the functions of these cells is to create a barrier that prevents substances in the portal capillary spaces from entering the brain. In this paper, we report on our use of immunohistochemistry to study the expression of tight junction proteins in the cells that compose the median eminence in adult mice. Our results indicate that tanycytes of the median eminence express occludin, ZO-1, and claudin 1 and 5, but not claudin 3. Remarkably, these molecules are organized as a continuous belt around the cell bodies of the tanycytes that line the ventral part of the third ventricle. In contrast, the tanycytes at the periphery of the arcuate nucleus do not express claudin 1 and instead exhibit a disorganized expression pattern of occludin, ZO-1, and claudin 5. Consistent with these observations, permeability studies using peripheral or central injections of Evans blue dye show that only the tanycytes of the median eminence are joined at their apices by functional tight junctions, whereas tanycytes located at the level of the arcuate nucleus form a permeable layer. In conclusion, this study reveals a unique expression pattern of tight junction proteins in hypothalamic tanycytes, which yields new insights into their barrier properties. J. Comp. Neurol. 518:943,962, 2010. © 2009 Wiley-Liss, Inc. [source]

Differential distribution of tight junction proteins suggests a role for tanycytes in blood-hypothalamus barrier regulation in the adult mouse brain

THE JOURNAL OF COMPARATIVE NEUROLOGY, Issue 7 2010
Amandine Mullier
Abstract The median eminence is one of the seven so-called circumventricular organs. It is located in the basal hypothalamus, ventral to the third ventricle and adjacent to the arcuate nucleus. This structure characteristically contains a rich capillary plexus and features a fenestrated endothelium, making it a direct target of blood-borne molecules. The median eminence also contains highly specialized ependymal cells called tanycytes, which line the floor of the third ventricle. It has been hypothesized that one of the functions of these cells is to create a barrier that prevents substances in the portal capillary spaces from entering the brain. In this paper, we utilize immunohistochemistry to study the expression of tight junction proteins in the cells that compose the median eminence in adult mice. Our results indicate that tanycytes of the median eminence express occludin, ZO-1, and claudin 1 and 5, but not claudin 3. Remarkably, these molecules are organized as a continuous belt around the cell bodies of the tanycytes that line the ventral part of the third ventricle. In contrast, the tanycytes at the periphery of the arcuate nucleus do not express claudin 1 and instead exhibit a disorganized expression pattern of occludin, ZO-1, and claudin 5. Consistent with these observations, permeability studies using peripheral or central injections of Evans blue dye show that only the tanycytes of the median eminence are joined at their apices by functional tight junctions, whereas tanycytes located at the level of the arcuate nucleus form a permeable layer. In conclusion, this study reveals a unique expression pattern of tight junction proteins in hypothalamic tanycytes, which yields new insights into their barrier properties. J. Comp. Neurol. 518:943,962, 2010. © 2009 Wiley-Liss, Inc. [source]

Disruption of tight junction structure in salivary glands from Sjögren's syndrome patients is linked to proinflammatory cytokine exposure

The action of pro-inflammatory cytokines on retinal endothelial cell barrier permeability: protective effect of corticosteroids

ACTA OPHTHALMOLOGICA, Issue 2008
AF AMBROSIO
Purpose The pro-inflammatory cytokines interleukin-1, (IL-1,) and tumor necrosis factor-alpha (TNF-,) were found to be increased in the vitreous of diabetic patients and in diabetic rat retinas, and increased cytokine levels were correlated with elevated retinal vascular permeability. In this work, we investigated the mechanisms underlying IL-1,- and TNF-,-induced retinal endothelial cell permeability and evaluated the ability of a glucocorticoid, dexamethasone (DEX), to prevent changes in permeability. Methods Primary cultures of bovine retinal endothelial cells (BRECs) were grown on transwell filters and exposed to IL-1, and TNF-,. BRECs permeability to 70 kDa RITC-dextran was measured. The content and localization of tight junction proteins was assessed by Western blotting and immunocytochemistry. Results IL-1, and TNF-, increased retinal endothelial cell permeability in a concentration- and time-dependent manner, but TNF-, was more effective (increased permeability at a lower dose and shorter time-point). The increase in permeability was not due to changes in cell viability. IL-1, and TNF-, altered ZO-1 and claudin-5 content. TNF-, also decreased ZO-1 staining at the cell border. Pre-treatment with DEX prevented TNF-,-induced cell permeability, and the protective effect of DEX was partially abolished by the glucocorticoid receptor antagonist RU486. Conclusion These data demonstrate that TNF-, and IL-1, potently induce endothelial cell permeability through alterations in tight junctions. Also, the study supports the potential therapeutic use of glucocorticoids to reduce retinal vascular permeability. Support: FCT (Portugal), NIH, JDRF and Allergan [source]

Abrogation of IFN-, mediated epithelial barrier disruption by serine protease inhibition

CLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 2 2005
L. E. M. Willemsen
Summary The intestinal barrier function is often impaired in a variety of diseases including chronic inflammatory bowel disease. Increased intestinal permeability during episodes of active disease correlates with destruction or rearrangement of the tight junction protein complex. IFN-, has been widely studied for its effect on barrier function and tight junction structures but its mode of action remains unclear. Since the claudin family of tight junction proteins is proposed to be involved in barrier maintenance we studied the effect of IFN-, on claudin expression in relation to epithelial barrier function. Cycloheximide and protease inhibitors were used to study mechanisms of IFN-, mediated barrier disruption. Intestinal epithelial cells were exposed to IFN-, and permeability was evaluated by horse radish peroxidase (HRP) and 4 kD FITC-dextran fluxes. Occludin and claudin-1, -2, -3, and -4 tight junction protein expression was determined by Western blotting. Occludin and claudin-2 protein expression was dramatically reduced after IFN-, exposure, which correlated with increased permeability for HRP and FITC-dextran. Interestingly, cleavage of claudin-2 was observed after incubation with IFN-,. Serine protease inhibitor AEBSF completely abrogated IFN-, mediated barrier disruption which was associated with preservation of claudin-2 expression. Moreover, IFN-, induced loss of barrier integrity was found to affect claudin-2 and occludin expression through different mechanisms. Since inhibition of serine protease activity abrogates IFN-, mediated barrier disruption this may be an important target for therapeutic intervention. [source]