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tight junction [keywords]
- The Beneficial Effect of Melatonin in Brain Endothelial Cells against Oxygen-Glucose Deprivation Followed by Reperfusion-Induced Injury. [Journal Article]
- Oxid Med Cell Longev 2014.:639531.
Melatonin has a cellular protective effect in cerebrovascular and neurodegenerative diseases. Protection of brain endothelial cells against hypoxia and oxidative stress is important for treatment of central nervous system (CNS) diseases, since brain endothelial cells constitute the blood brain barrier (BBB). In the present study, we investigated the protective effect of melatonin against oxygen-glucose deprivation, followed by reperfusion- (OGD/R-) induced injury, in bEnd.3 cells. The effect of melatonin was examined by western blot analysis, cell viability assays, measurement of intracellular reactive oxygen species (ROS), and immunocytochemistry (ICC). Our results showed that treatment with melatonin prevents cell death and degradation of tight junction protein in the setting of OGD/R-induced injury. In response to OGD/R injury of bEnd.3 cells, melatonin activates Akt, which promotes cell survival, and attenuates phosphorylation of JNK, which triggers apoptosis. Thus, melatonin protects bEnd.3 cells against OGD/R-induced injury.
- Signalling at tight junctions during epithelial differentiation and microbial pathogenesis. [Journal Article]
- J Cell Sci 2014 Aug 15; 127(Pt 16):3401-13.
Tight junctions are a component of the epithelial junctional complex, and they form the paracellular diffusion barrier that enables epithelial cells to create cellular sheets that separate compartments with different compositions. The assembly and function of tight junctions are intimately linked to the actomyosin cytoskeleton and, hence, are under the control of signalling mechanisms that regulate cytoskeletal dynamics. Tight junctions not only receive signals that guide their assembly and function, but transmit information to the cell interior to regulate cell proliferation, migration and survival. As a crucial component of the epithelial barrier, they are often targeted by pathogenic viruses and bacteria, aiding infection and the development of disease. In this Commentary, we review recent progress in the understanding of the molecular signalling mechanisms that drive junction assembly and function, and the signalling processes by which tight junctions regulate cell behaviour and survival. We also discuss the way in which junctional components are exploited by pathogenic viruses and bacteria, and how this might affect junctional signalling mechanisms.
- Helicobacter pylori targets cancer-associated apical-junctional constituents in gastroids and gastric epithelial cells. [JOURNAL ARTICLE]
- Gut 2014 Aug 14.
Helicobacter pylori strains that express the oncoprotein CagA augment risk for gastric cancer. However, the precise mechanisms through which cag(+) strains heighten cancer risk have not been fully delineated and model systems that recapitulate the gastric niche are critical for understanding pathogenesis. Gastroids are three-dimensional organ-like structures that provide unique opportunities to study host-H. pylori interactions in a preclinical model. We used gastroids to inform and direct in vitro studies to define mechanisms through which H. pylori modulates expression of the cancer-associated tight junction protein claudin-7.Gastroids were infected by luminal microinjection, and MKN28 gastric epithelial cells were cocultured with H. pylori wild-type cag(+) strains or isogenic mutants. β-catenin, claudin-7 and snail localisation was determined by immunocytochemistry. Proliferation was assessed using 5-ethynyl-2'-deoxyuridine, and levels of claudin-7 and snail were determined by western blot and flow cytometry.Gastroids developed into a self-organising differentiation axis and H. pylori induced mislocalisation of claudin-7 and increased proliferation in a CagA- and β-catenin-dependent manner. In MKN28 cells, H pylori-induced suppression of claudin-7 was regulated by β-catenin and snail. Similarly, snail expression was increased and claudin-7 levels were decreased among H. pylori-infected individuals.H. pylori increase proliferation in a strain-specific manner in a novel gastroid system. H. pylori also alter expression and localisation of claudin-7 in gastroids and human epithelial cells, which is mediated by β-catenin and snail activation. These data provide new insights into molecular interactions with carcinogenic potential that occur between H. pylori and epithelial cells within the gastric niche.
- Cadherin cytoplasmic domains inhibit the cell surface localization of endogenous e-cadherin, blocking desmosome and tight junction formation and inducing cell dissociation. [Journal Article]
- PLoS One 2014; 9(8):e105313.
The downregulation of E-cadherin function has fundamental consequences with respect to cancer progression, and occurs as part of the epithelial-mesenchymal transition (EMT). In this study, we show that the expression of the Discosoma sp. red fluorescent protein (DsRed)-tagged cadherin cytoplasmic domain in cells inhibited the cell surface localization of endogenous E-cadherin, leading to morphological changes, the inhibition of junctional assembly and cell dissociation. These changes were associated with increased cell migration, but were not accompanied by the down-regulation of epithelial markers and up-regulation of mesenchymal markers. Thus, these changes cannot be classified as EMT. The cadherin cytoplasmic domain interacted with β-catenin or plakoglobin, reducing the levels of β-catenin or plakoglobin associated with E-cadherin, and raising the possibility that β-catenin and plakoglobin sequestration by these constructs induced E-cadherin intracellular localization. Accordingly, a cytoplasmic domain construct bearing mutations that weakened the interactions with β-catenin or plakoglobin did not impair junction formation and adhesion, indicating that the interaction with β-catenin or plakoglobin was essential to the potential of the constructs. E-cadherin-α-catenin chimeras that did not require β-catenin or plakoglobin for their cell surface transport restored cell-cell adhesion and junction formation.
- Vascular disruption and blood-brain barrier dysfunction in intracerebral hemorrhage. [Journal Article, Review]
- Fluids Barriers CNS 2014.:18.
This article reviews current knowledge of the mechanisms underlying the initial hemorrhage and secondary blood-brain barrier (BBB) dysfunction in primary spontaneous intracerebral hemorrhage (ICH) in adults. Multiple etiologies are associated with ICH, for example, hypertension, Alzheimer's disease, vascular malformations and coagulopathies (genetic or drug-induced). After the initial bleed, there can be continued bleeding over the first 24 hours, so-called hematoma expansion, which is associated with adverse outcomes. A number of clinical trials are focused on trying to limit such expansion. Significant progress has been made on the causes of BBB dysfunction after ICH at the molecular and cell signaling level. Blood components (e.g. thrombin, hemoglobin, iron) and the inflammatory response to those components play a large role in ICH-induced BBB dysfunction. There are current clinical trials of minimally invasive hematoma removal and iron chelation which may limit such dysfunction. Understanding the mechanisms underlying the initial hemorrhage and secondary BBB dysfunction in ICH is vital for developing methods to prevent and treat this devastating form of stroke.
- Claudin-4 expression in gastric cancer cells enhances the invasion and is associated with the increased level of matrix metalloproteinase-2 and -9 expression. [JOURNAL ARTICLE]
- Oncol Lett 2014 Sep; 8(3):1367-1371.
Claudin-4 is a member of a large family of transmembrane proteins known as claudins, which are essential for the formation and maintenance of tight junctions. Our previous studies have revealed that claudin-4 proteins are overexpressed in metastatic gastric cancer. To clarify the roles of claudin-4 in gastric cancer metastasis, human gastric adenocarcinoma (AGS) cells constitutively expressing wild-type claudin-4 were generated. Expression of claudin-4 in AGS cells was found to increase cell invasion and migration, as measured by Boyden invasion chamber assays. Moreover, the claudin-4-expressing AGS cells were found to have increased matrix metalloproteinase (MMP)-2 and -9 expression, indicating that claudin-mediated increased invasion may be mediated through the activation of the MMP protein. Overall, the results suggest that claudin-4 overexpression may promote gastric cancer metastasis through the increased invasion of gastric cancer cells.
- Overexpression of netrin-1 increases the expression of tight junction-associated proteins, claudin-5, occludin, and ZO-1, following traumatic brain injury in rats. [JOURNAL ARTICLE]
- Exp Ther Med 2014 Sep; 8(3):881-886.
The function of the blood-brain barrier (BBB) depends on the integrity of tight junction (TJ)-associated proteins. Netrin-1 is known to promote angiogenesis and may also regulate the BBB. To understand the association between netrin-1 and the TJ-associated proteins, the expression levels of proteins involved in maintaining the integrity of the BBB, including netrin-1, claudin-5, occludin and zonula occluden (ZO)-1, were investigated in the present study using quantitative polymerase chain reaction, western blot analysis and immunofluorescence. The aim of the present study was to determine the changes in BBB permeability and whether pZsGreen1-N1 mediated overexpression of netrin-1 increased the expression of the TJ-associated proteins following traumatic brain injury (TBI). The results demonstrated that the levels of mRNA transcription and protein expression of the TJ-associated proteins, claudin-5, occludin and ZO-1, were significantly reduced following TBI. Furthermore, the changes in the expression of these three TJ proteins were consistent with the changes in the BBB permeability, indicating that weakening intercellular junctions leads to BBB opening. The present study also demonstrated that netrin-1 significantly increased the downregulation of claudin-5, occludin and ZO-1 expression levels induced by TBI, which provided a basis for further investigation on the role of netrin-1 in the integrity of TJs and proper functioning of the BBB.
- In-silico analysis of claudin-5 reveals novel putative sites for post-translational modifications: Insights into potential molecular determinants of blood-brain barrier breach during HIV-1 infiltration. [JOURNAL ARTICLE]
- Infect Genet Evol 2014 Aug 10.
The blood-brain barrier (BBB) poses a huge challenge and is a serious issue in deciphering the pathophysiology of central nervous system disorders. Endothelial tight junctions play an essential role in maintaining the integrity of the BBB. Post-translational modifications (PTMs) in endothelial tight junction proteins are known to cause deleterious functional impairment and possible disruptions in BBB integrity. PTMs in tight junction proteins play an important role in human immunodeficiency virus type 1 (HIV-1) entry through the BBB. Human claudin-5 is one of the highly expressed brain endothelial tight junction protein and various PTMs in claudin-5 are expected to aid HIV-1 in crossing the BBB. A precise characterization of PTMs in claudin-5 is important for understanding its role in HIV-1 brain infiltration. In this study, we have examined post-translational crosstalk between phosphorylation, O-glycosylation, palmitoylation and methylation sites in claudin-5, which could alter claudin-5's ability to maintain BBB integrity. To the best of our knowledge, this is the first report on claudin-5 protein that suggests a novel interplay between potential PTM sites. PTMs of predicted residues in claudin-5, suggested in this study, can serve as compelling targets for potential therapeutic agents against HIV-1 induced neuropathogenesis. Further site-specific experimental studies in this aspect are highly recommended.
- CAR expression in human embryos and hESC illustrates its role in pluripotency and tight junctions. [JOURNAL ARTICLE]
- Reproduction 2014 Aug 12.
Coxsackie virus and adenovirus receptor (CAR) is present during embryogenesis and is involved in tissue regeneration, cancer and intercellular adhesion. We investigated the expression of CAR in human preimplantation embryos and embryonic stem cells (hESC) to identify its role in early embryogenesis and differentiation. CAR protein was ubiquitously present during preimplantation development. It was nuclear in uncommitted cells from the cleavage stage up to the precursor epiblast and corresponded with the presence of soluble CAR3/7 splice variant. CAR was displayed on the membrane, participating in tight junction formations at compaction and blastocyst stages in both outer and inner cells, and CAR corresponded with the full-length CAR-containing transmembrane domain. In trophectodermal cells of hatched blastocysts, CAR was reduced in the membrane and concentrated in the nucleus, which correlated with the switch in RNA expression to the CAR4/7 and CAR2/7 splice variants. Cells of the outer layer in hESC colonies contained CAR on the membrane and all the cells of the colony had CAR in the nucleus, corresponding with the transmembrane CAR and CAR4/7. Upon hESC differentiation into cells representing the three germ layers and trophoblast lineage, CAR expression was downregulated. We concluded that CAR is differentially expressed during human preimplantation development. We described various CAR expressions: (1) soluble CAR marking undifferentiated blastomeres; (2) transmembrane CAR related to epithelial-like cell types, such as the trophectoderm and the outer layer of hESC colonies; and (3) soluble CAR present in trophectoderm nuclei after hatching. The functions of these distinct forms remain to be elucidated.
- Toll-like receptor 3 activation is required for normal skin barrier repair following UV damage. [JOURNAL ARTICLE]
- J Invest Dermatol 2014 Aug 12.
Ultraviolet (UV) damage to the skin leads to the release of noncoding RNA (ncRNA) from necrotic keratinocytes that activates toll-like receptor 3 (TLR3). This release of ncRNA triggers inflammation in the skin following UV damage. Recently, TLR3 activation was also shown to aid wound repair and increase expression of genes associated with permeability barrier repair. Here, we sought to test if skin barrier repair after UVB damage is dependent on the activation of TLR3. We observed that multiple ncRNAs induced expression of skin barrier repair genes, that the TLR3 ligand Poly (I:C) also induced expression and function of tight junctions, and that the ncRNA U1 acts in a TLR3-dependent manner to induce expression of skin barrier repair genes. These observations were shown to have functional relevance as Tlr3(-/-) mice displayed a delay in skin barrier repair following UVB damage. Combined, these data further validate the conclusion that recognition of endogenous RNA by TLR3 is an important step in the program of skin barrier repair.Journal of Investigative Dermatology accepted article preview online, 12 August 2014; doi:10.1038/jid.2014.354.