scholarly journals The Role of E-Cadherin in Maintaining the Barrier Function of Corneal Epithelium after Treatment with Cultured Autologous Oral Mucosa Epithelial Cell Sheet Grafts for Limbal Stem Deficiency

2016 ◽  
Vol 2016 ◽  
pp. 1-13 ◽  
Author(s):  
Fawzia Bardag-Gorce ◽  
Richard H. Hoft ◽  
Andrew Wood ◽  
Joan Oliva ◽  
Hope Niihara ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Cell Membrane ◽  
Oral Mucosa ◽  
Barrier Function ◽  
Cell Sheet ◽  
Cellular Adhesion ◽  
Beta Catenin ◽  
Junction Protein ◽  
E Cadherin

The role of E-cadherin in epithelial barrier function of cultured autologous oral mucosa epithelial cell sheet (CAOMECS) grafts was examined. CAOMECS were cultured on a temperature-responsive surface and grafted onto rabbit corneas with Limbal Stem Cell Deficiency (LSCD). E-cadherin levels were significantly higher in CAOMECS compared to normal and LSCD epithelium. Beta-catenin colocalized with E-cadherin in CAOMECS cell membranes while phosphorylated beta-catenin was significantly increased. ZO-1, occludin, and Cnx43 were also strongly expressed in CAOMECS. E-cadherin and beta-catenin localization at the cell membrane was reduced in LSCD corneas, while CAOMECS-grafted corneas showed a restoration of E-cadherin and beta-catenin expression. LSCD corneas did not show continuous staining for ZO-1 or for Cnx43, while CAOMECS-grafted corneas showed a positive expression of ZO-1 and Cnx43. Cascade Blue® hydrazide did not pass through CAOMECS. Because E-cadherin interactions are calcium-dependent, EGTA was used to chelate calcium and disrupt cell adhesion. EGTA-treated CAOMECS completely detached from cell culture surface, and E-cadherin levels were significantly decreased. In conclusion, E cadherin high expression contributed to CAOMECS tight and gap junction protein recruitment at the cell membrane, thus promoting cellular adhesion and a functional barrier to protect the ocular surface.

A-kinase anchoring proteins contribute to loss of E-cadherin and bronchial epithelial barrier by cigarette smoke

2014 ◽  
Vol 306 (6) ◽  
pp. C585-C597 ◽  
Author(s):  
Anouk Oldenburger ◽  
Wilfred J. Poppinga ◽  
Fleur Kos ◽  
Harold G. de Bruin ◽  
Wolter F. Rijks ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Protein Expression ◽  
Cigarette Smoke ◽  
Barrier Function ◽  
Epithelial Barrier ◽  
Chronic Obstructive ◽  
Bronchial Epithelial ◽  
Anchoring Proteins ◽  
E Cadherin

Airway epithelium, which forms the first barrier towards environmental insults, is disturbed by cigarette smoking, a major risk factor for developing chronic obstructive pulmonary disease (COPD). A-kinase anchoring proteins (AKAP) maintain endothelial barrier function and coordinate subcellular localization of protein kinase A (PKA). However, the role of AKAPs in epithelial barrier function is unknown. We studied the role of AKAPs in regulating human bronchial epithelial (Hogg JC, Timens W. Annu Rev Pathol 4: 435–459, 2009; HBE) barrier. Cigarette smoke extract (CSE) reduced barrier function in 16HBE cells and the expression of the adhesion molecule E-cadherin specifically at the cell membrane. In addition, CSE reduced the protein expression of the AKAP family member AKAP9 at the cell membrane. The expression of AKAP5 and AKAP12 was unaffected by CSE. AKAP9 interacted and colocalized with E-cadherin at the cell membrane, suggesting that the reduction of both proteins may be related. Interestingly, disruption of AKAP-PKA interactions by st-Ht31 prevented the CSE-induced reduction of E-cadherin and AKAP9 protein expression and subsequent loss of barrier function. Silencing of AKAP9 reduced the functional epithelial barrier and prevented the ability of st-Ht31 to restore membrane localization of E-cadherin. Our data suggest the possibility of a specific role for AKAP9 in the maintenance of the epithelial barrier. E-cadherin, but not AKAP9, protein expression was reduced in lung tissue from COPD patients compared with controls. However, AKAP9 mRNA expression was decreased in primary bronchial epithelial cells from current smokers compared with non/ex-smokers. In conclusion, our results indicate that AKAP proteins, most likely AKAP9, maintain the bronchial epithelial barrier by regulating the E-cadherin expression at the cell membrane.

l-Glutamine ameliorates acetaldehyde-induced increase in paracellular permeability in Caco-2 cell monolayer

2004 ◽  
Vol 287 (3) ◽  
pp. G510-G517 ◽  
Author(s):  
A. Seth ◽  
S. Basuroy ◽  
P. Sheth ◽  
R. K. Rao
Keyword(s):  
Barrier Function ◽  
Egf Receptor ◽  
Kinase Inhibitor ◽  
Cell Monolayer ◽  
Intercellular Junctions ◽  
Paracellular Permeability ◽  
Dependent Manner ◽  
Dose Dependent ◽  
E Cadherin

Role of l-glutamine in the protection of intestinal epithelium from acetaldehyde-induced disruption of barrier function was evaluated in Caco-2 cell monolayer. l-Glutamine reduced the acetaldehyde-induced decrease in transepithelilal electrical resistance and increase in permeability to inulin and lipopolysaccharide in a time- and dose-dependent manner; d-glutamine, l-aspargine, l-arginine, l-lysine, or l-alanine produced no significant protection. The glutaminase inhibitor 6-diazo-5-oxo-l-norleucine failed to affect the l-glutamine-mediated protection of barrier function. l-Glutamine reduced the acetaldehyde-induced redistribution of occludin, zonula occludens-1 (ZO-1), E-cadherin, and β-catenin from the intercellular junctions. Acetaldehyde dissociates occludin, ZO-1, E-cadherin, and β-catenin from the actin cytoskeleton, and this effect was reduced by l-glutamine. l-Glutamine induced a rapid increase in the tyrosine phosphorylation of EGF receptor, and the protective effect of l-glutamine was prevented by AG1478, the EGF-receptor tyrosine kinase inhibitor. These results indicate that l-glutamine prevents acetaldehyde-induced disruption of the tight junction and increase in the paracellular permeability in Caco-2 cell monolayer by an EGF receptor-dependent mechanism.

Carrier-free Cultured Autologous Oral Mucosa Epithelial Cell Sheet (CAOMECS) for Corneal Epithelium Reconstruction: A Histological Study

2015 ◽  
Vol 13 (2) ◽  
pp. 150-163 ◽  
Author(s):  
Fawzia Bardag-Gorce ◽  
Joan Oliva ◽  
Andrew Wood ◽  
Richard Hoft ◽  
Derek Pan ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Oral Mucosa ◽  
Corneal Epithelium ◽  
Histological Study ◽  
Cell Sheet ◽  
Carrier Free

Assembly of the cadherin-catenin complex in vitro with recombinant proteins

1994 ◽  
Vol 107 (12) ◽  
pp. 3655-3663 ◽  
Author(s):  
H. Aberle ◽  
S. Butz ◽  
J. Stappert ◽  
H. Weissig ◽  
R. Kemler ◽  
...  
Keyword(s):  
Cytoplasmic Domain ◽  
Beta Catenin ◽  
Molecular Architecture ◽  
Adhesive Properties ◽  
Protein Protein Interaction ◽  
Signalling Molecules ◽  
Classical Cadherins ◽  
E Cadherin

The cytoplasmic domain of classical cadherins is tightly associated with three proteins termed alpha-, beta- and gamma-catenin. These accessory proteins are of central importance for the adhesive properties of this class of cell adhesion molecules. In order to examine the molecular architecture of the cadherin-catenin complex in more detail we have expressed the catenins and the cytoplasmic domain of E-cadherin as fusion proteins in Escherichia coli, and analyzed the interaction of purified recombinant cadherin and catenins in combinatorial protein-protein interaction experiments. The cytoplasmic domain of E-cadherin cannot directly associate with alpha-catenin but interacts with high affinity with beta-catenin, whereas the binding of gamma-catenin (plakoglobin) to E-cadherin is less efficient. alpha- and beta-catenin assemble into a 1:1 heterodimeric complex. The analysis of various truncated beta-catenins revealed that an alpha-catenin binding site in beta-catenin is localized between amino acid positions 120 and 151. The central role of beta-catenin for the assembly of the heterotrimeric E-cadherin/alpha-catenin/beta-catenin complex in mixing experiments with all components was demonstrated. The reconstitution in vitro of the cadherin-catenin complex should allow the study of the interaction with signalling molecules and with the actin-based cytoskeleton.

scholarly journals Hedgehog signaling regulates E-cadherin expression for the maintenance of the actin cytoskeleton and tight junctions

2010 ◽  
Vol 299 (6) ◽  
pp. G1252-G1265 ◽  
Author(s):  
Chang Xiao ◽  
Sally A. Ogle ◽  
Michael A. Schumacher ◽  
Neal Schilling ◽  
Robert A. Tokhunts ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Epithelial Cell ◽  
Tight Junctions ◽  
Hedgehog Signaling ◽  
Protein Complexes ◽  
Cell Lineage ◽  
Adherens Junction ◽  
Epithelial Cell Differentiation ◽  
Junction Protein ◽  
E Cadherin

In the stomach, strictly regulated cell adherens junctions are crucial in determining epithelial cell differentiation. Sonic Hedgehog (Shh) regulates epithelial cell differentiation in the adult stomach. We sought to identify whether Shh plays a role in regulating adherens junction protein E-cadherin as a mechanism for epithelial cell differentiation. Mouse nontumorigenic gastric epithelial (IMGE-5) cells treated with Hedgehog signaling inhibitor cyclopamine and anti-Shh 5E1 antibody or transduced with short hairpin RNA against Skinny Hedgehog (IMGE-5Ski) were cultured. A mouse model expressing a parietal cell-specific deletion of Shh (HKCre/ShhKO) was used to identify further changes in adherens and tight junctions. Inhibition of Hedgehog signaling in IMGE-5 cells caused loss of E-cadherin expression accompanied by disruption of F-actin cortical expression and relocalization of zonula occludens-1 (ZO-1). Loss of E-cadherin was also associated with increased proliferation in IMGE-5Ski cells and increased expression of the mucous neck cell lineage marker MUC6. Compared with membrane-expressed E-cadherin and ZO-1 protein in controls, dissociation of E-cadherin/β-catenin and ZO-1/occludin protein complexes was observed in HKCre/ShhKO mice. In conclusion, we demonstrate that Hedgehog signaling regulates E-cadherin expression that is required for the maintenance of F-actin cortical expression and stability of tight junction protein ZO-1.

scholarly journals A Synthetic Peptide Corresponding to the Extracellular Domain of Occludin Perturbs the Tight Junction Permeability Barrier

1997 ◽  
Vol 136 (2) ◽  
pp. 399-409 ◽  
Author(s):  
Vivian Wong ◽  
Barry M. Gumbiner
Keyword(s):  
Epithelial Cell ◽  
Tight Junction ◽  
Adherens Junction ◽  
Extracellular Domain ◽  
Integral Membrane Protein ◽  
Epithelial Cell Line ◽  
Permeability Barrier ◽  
Junction Protein ◽  
Tight Junction Permeability

Occludin, the putative tight junction integral membrane protein, is an attractive candidate for a protein that forms the actual sealing element of the tight junction. To study the role of occludin in the formation of the tight junction seal, synthetic peptides (OCC1 and OCC2) corresponding to the two putative extracellular domains of occludin were assayed for their ability to alter tight junctions in Xenopus kidney epithelial cell line A6. Transepithelial electrical resistance and paracellular tracer flux measurements indicated that the second extracellular domain peptide (OCC2) reversibly disrupted the transepithelial permeability barrier at concentrations of < 5 μM. Despite the increased paracellular permeability, there were no changes in gross epithelial cell morphology as determined by scanning EM. The OCC2 peptide decreased the amount of occludin present at the tight junction, as assessed by indirect immunofluorescence, as well as decreased total cellular content of occludin, as assessed by Western blot analysis. Pulse-labeling and metabolic chase analysis suggested that this decrease in occludin level could be attributed to an increase in turnover of cellular occludin rather than a decrease in occludin synthesis. The effect on occludin was specific because other tight junction components, ZO-1, ZO-2, cingulin, and the adherens junction protein E-cadherin, were unaltered by OCC2 treatment. Therefore, the peptide corresponding to the second extracellular domain of occludin perturbs the tight junction permeability barrier in a very specific manner. The correlation between a decrease in occludin levels and the perturbation of the tight junction permeability barrier provides evidence for a role of occludin in the formation of the tight junction seal.

scholarly journals The serine protease-mediated increase in intestinal epithelial barrier function is dependent on occludin and requires an intact tight junction

2016 ◽  
Vol 311 (3) ◽  
pp. G466-G479 ◽  
Author(s):  
Natalie J. Ronaghan ◽  
Judie Shang ◽  
Vadim Iablokov ◽  
Raza Zaheer ◽  
Pina Colarusso ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Tight Junction ◽  
Serine Protease ◽  
Barrier Function ◽  
Serine Proteases ◽  
Cell Fractionation ◽  
Intestinal Epithelial ◽  
Tight Junction Proteins ◽  
Junction Protein ◽  
Junction Proteins

Barrier dysfunction is a characteristic of the inflammatory bowel diseases (IBD), Crohn's disease and ulcerative colitis. Understanding how the tight junction is modified to maintain barrier function may provide avenues for treatment of IBD. We have previously shown that the apical addition of serine proteases to intestinal epithelial cell lines causes a rapid and sustained increase in transepithelial electrical resistance (TER), but the mechanisms are unknown. We hypothesized that serine proteases increase barrier function through trafficking and insertion of tight junction proteins into the membrane, and this could enhance recovery of a disrupted monolayer after calcium switch or cytokine treatment. In the canine epithelial cell line, SCBN, we showed that matriptase, an endogenous serine protease, could potently increase TER. Using detergent solubility-based cell fractionation, we found that neither trypsin nor matriptase treatment changed levels of tight junction proteins at the membrane. In a fast calcium switch assay, serine proteases did not enhance the rate of recovery of the junction. In addition, serine proteases could not reverse barrier disruption induced by IFNγ and TNFα. We knocked down occludin in our cells using siRNA and found this prevented the serine protease-induced increase in TER. Using fluorescence recovery after photobleaching (FRAP), we found serine proteases induce a greater mobile fraction of occludin in the membrane. These data suggest that a functional tight junction is needed for serine proteases to have an effect on TER, and that occludin is a crucial tight junction protein in this mechanism.

scholarly journals CD151 regulates epithelial cell–cell adhesion through PKC- and Cdc42-dependent actin cytoskeletal reorganization

2003 ◽  
Vol 163 (1) ◽  
pp. 165-176 ◽  
Author(s):  
Masaki Shigeta ◽  
Noriko Sanzen ◽  
Masayuki Ozawa ◽  
Jianguo Gu ◽  
Hitoshi Hasegawa ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Epithelial Cell ◽  
Cytoskeletal Reorganization ◽  
Cortical Actin ◽  
A431 Cells ◽  
Kinase C ◽  
Integrin Α3β1 ◽  
E Cadherin ◽  
Cell Cell

CD151, a member of the tetraspanin family proteins, tightly associates with integrin α3β1 and localizes at basolateral surfaces of epithelial cells. We found that overexpression of CD151 in A431 cells accelerated intercellular adhesion, whereas treatment of cells with anti-CD151 mAb perturbed the integrity of cortical actin filaments and cell polarity. E-Cadherin puncta formation, indicative of filopodia-based adhesion zipper formation, as well as E-cadherin anchorage to detergent-insoluble cytoskeletal matrix, was enhanced in CD151-overexpressing cells. Levels of GTP-bound Cdc42 and Rac were also elevated in CD151-overexpressing cells, suggesting the role of CD151 in E-cadherin–mediated cell–cell adhesion as a modulator of actin cytoskeletal reorganization. Consistent with this possibility, engagement of CD151 by the substrate-adsorbed anti-CD151 mAb induced prominent Cdc42-dependent filopodial extension, which along with E-cadherin puncta formation, was strongly inhibited by calphostin C, a protein kinase C (PKC) inhibitor. Together, these results indicate that CD151 is involved in epithelial cell–cell adhesion as a modulator of PKC- and Cdc42-dependent actin cytoskeletal reorganization.

Interactions of Modern Aesthetic Materials for Prosthetic Restorations with Oral Mucosa A pilot study

2018 ◽  
Vol 69 (2) ◽  
pp. 386-390
Author(s):  
Bogdan Calenic ◽  
Mihaela Pantea ◽  
Ana Maria Cristina Tancu ◽  
Paula Perlea ◽  
Maria Greabu ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Cell Line ◽  
Polymethyl Methacrylate ◽  
Oral Mucosa ◽  
Precious Metal ◽  
Metal Alloy ◽  
Health Science ◽  
Epithelial Cell Line ◽  
Gingival Epithelial Cell ◽  
E Cadherin

The specific objective of the present study is to assess the interactions between cells from a human gingival epithelial cell line and various aesthetic materials used in modern prosthetic dentistry. For this study six types of dental materials were selected: Cr-Co non-precious metal alloy, ceramics applied on Cr-Co non-precious metal alloy, zirconia, ceramics applied on zirconia, polymethyl methacrylate and pressed ceramics/lithium disilicate. Cells from a human gingival epithelial cell line, Ca9-22 (Health Science Research Resources Bank), were cultured on the chosen surfaces for 3, 5 and 7 days. Cellular proliferation, cell attachment (using Multiplex Arrays Technology) and cytotoxicity (MTT- Assay) were evaluated at distinct predetermined intervals. Measurements performed at each distinct predetermined interval showed no significant difference for cell proliferation and cytotoxicity between the selected surfaces, however the highest levels were registered for the polymethyl methacrylate surface. Different attachment patterns were observed for epithelial cells attached on substrates, such as significantly different levels of adherence of E-Cadherin and N-Cadherin molecules; E-Cadherin adhesion levels indicate that pressed ceramics may be the dental material which, compared to the selected materials, influences the least the homeostasis of oral mucosa.

Sign in / Sign up

Export Citation Format

Share Document