scholarly journals Polarized entry and release of Junín virus, a New World arenavirus

2005 ◽  
Vol 86 (5) ◽  
pp. 1475-1479 ◽  
Author(s):  
Sandra M. Cordo ◽  
Maximiliano Cesio y Acuña ◽  
Nélida A. Candurra
Keyword(s):  
Epithelial Cells ◽  
Membrane Surface ◽  
The Body ◽  
Apical Plasma Membrane ◽  
Apical Surface ◽  
High Level Expression ◽  
Virus Family ◽  
Junin Virus ◽  
High Level ◽  
Junín Virus

Junín virus (JUNV), the causative agent of Argentine haemorrhagic fever, is a human pathogen that naturally enters the body through the epithelial cells of the respiratory and digestive tracts. The interaction of JUNV with two types of polarized epithelial cultures, Vero C1008 and A549, was investigated. Radioactive virus-binding assays showed that JUNV infects polarized lines preferentially through the apical surface. High-level expression of viral nucleoprotein was detected in polarized cell lines infected through the apical domain. Virus production from apical media was about 100-fold higher than that found into the basolateral medium. Confocal-immunofluorescence analysis revealed high-level expression of glycoprotein at the apical-membrane surface. Disruption of the microtubule network by colchicine impaired JUNV vectorial release. This is the first study to analyse the interaction between a member of the virus family Arenaviridae and polarized epithelial cells, showing preferential entry and release from the apical plasma membrane.

CFTR channel insertion to the apical surface in rat duodenal villus epithelial cells is upregulated by VIP in vivo

1999 ◽  
Vol 112 (6) ◽  
pp. 887-894
Author(s):  
N.A. Ameen ◽  
B. Martensson ◽  
L. Bourguinon ◽  
C. Marino ◽  
J. Isenberg ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Apical Membrane ◽  
Surface Expression ◽  
Apical Plasma Membrane ◽  
Intracellular Camp ◽  
Apical Surface ◽  
C Terminus ◽  
Confocal Immunofluorescence ◽  
Small Intestinal Villus

cAMP activated insertion of the cystic fibrosis transmembrane conductance regulator (CFTR) channels from endosomes to the apical plasma membrane has been hypothesized to regulate surface expression and CFTR function although the physiologic relevance of this remains unclear. We previously identified a subpopulation of small intestinal villus epithelial cells or CFTR high expressor (CHE) cells possessing very high levels of apical membrane CFTR in association with a prominent subapical vesicular pool of CFTR. We have examined the subcellular redistribution of CFTR in duodenal CHE cells in vivo in response to the cAMP activated secretagogue vasoactive intestinal peptide (VIP). Using anti-CFTR antibodies against the C terminus of rodent CFTR and indirect immunofluorescence, we show by quantitative confocal microscopy that CFTR rapidly redistributes from the cytoplasm to the apical surface upon cAMP stimulation by VIP and returns to the cytoplasm upon removal of VIP stimulation of intracellular cAMP levels. Using ultrastructural and confocal immunofluorescence examination in the presence or absence of cycloheximide, we also show that redistribution was not dependent on new protein synthesis, changes in endocytosis, or rearrangement of the apical cytoskeleton. These observations suggest that physiologic cAMP activated apical membrane insertion and recycling of CFTR channels in normal CFTR expressing epithelia contributes to the in vivo regulation of CFTR mediated anion transport.

Differential targeting of facilitative glucose transporters in polarized epithelial cells

1996 ◽  
Vol 271 (2) ◽  
pp. C547-C554 ◽  
Author(s):  
W. S. Pascoe ◽  
K. Inukai ◽  
Y. Oka ◽  
J. W. Slot ◽  
D. E. James
Keyword(s):  
Epithelial Cells ◽  
Glucose Uptake ◽  
Glucose Transporter ◽  
Mdck Cells ◽  
Intracellular Localization ◽  
Apical Plasma Membrane ◽  
Cellular Polarity ◽  
Glut 1 ◽  
Polarized Epithelial Cells ◽  
High Level

We have examined the intracellular localization of five facilitative glucose transporter proteins, one endogenous (GLUT-1) and four exogenous (GLUT-2, -3, -4, and -5), in polarized epithelial cells. GLUT-2, -3, -4, and -5 were stably transfected into Madin-Darby canine kidney (MDCK) cells, and peptide-specific antibodies were used to establish their distribution by immunofluorescence and immunoelectron-microscopic techniques. GLUT-1 and -2 were predominantly targeted to the basolateral domain of the cell, whereas GLUT-3 and -5 were targeted to the apical plasma membrane. The insulin-regulatable glucose transporter GLUT-4 was found in intracellular tubulovesicular structures beneath the surface of the cell. Vectorial 2-deoxy-D-glucose uptake measurements revealed that approximately 95% of glucose entry into wild-type MDCK cells occurs via the basolateral membranes. In GLUT-3-transfected cells, however, apical glucose uptake increased to approximately 55%; this was not observed in cells expressing the other GLUT isoforms. The discrete and differential intracellular localizations of the various GLUTs, in addition to the high level of sequence homology and predicted secondary structure similarity, render the GLUT family ideal for the study of intrinsic targeting motifs involved in the establishment and maintenance of cellular polarity.

Optimization of Plasmid Vectors for High-Level Expression in Lung Epithelial Cells

1997 ◽  
Vol 8 (5) ◽  
pp. 575-584 ◽  
Author(s):  
Nelson S. Yew ◽  
Donna M. Wysokenski ◽  
Kathryn X. Wang ◽  
Robin J. Ziegler ◽  
John Marshall ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Lung Epithelial Cells ◽  
High Level Expression ◽  
Plasmid Vectors ◽  
Lung Epithelial ◽  
High Level

Role of EP4 receptor and prostaglandin transporter in prostaglandin E2-induced alteration in colonic epithelial barrier integrity

2010 ◽  
Vol 299 (5) ◽  
pp. G1097-G1105 ◽  
Author(s):  
Manigandan Lejeune ◽  
Pearl Leung ◽  
Paul L. Beck ◽  
Kris Chadee
Keyword(s):  
Epithelial Cells ◽  
Apical Plasma Membrane ◽  
Epithelial Barrier ◽  
Prostaglandin E ◽  
Apical Surface ◽  
Barrier Integrity ◽  
Ep4 Receptor ◽  
Tnf Α ◽  
Mucosal Folds ◽  
Vectorial Transport

Prostaglandin E2 (PGE2) is a proinflammatory lipid mediator produced in excess in inflammatory bowel disease (IBD). PGE2 couples to and signals via four different E-prostanoid (EP) receptors, namely EP1, EP2, EP3, and EP4. In this study, we determined a role for PGE2 and EP4 receptors in altering colonic epithelial barrier integrity. In healthy colonic mucosa, EP4 receptors were localized on apical plasma membrane of epithelial cells at the tip of mucosal folds, whereas, in patients with IBD and in rats with dextran sodium sulfate (DSS)-induced colitis, they were diffusely overexpressed throughout the mucosa. Similarly, expression of EP4 receptor was polarized in T84 colonic epithelial monolayer and mimics the normal epithelium. Apical exposure of T84 monolayer with high levels of PGE2 decreased barrier integrity, which was abrogated by an EP4 receptor antagonist. To reveal the mechanism of vectorial transport of basally produced PGE2 toward apical EP4 receptors, we identified prostaglandin transporters (PGT) in human colonic epithelia. PGT were least expressed on epithelial cells at the colonic mucosal folds of control subjects but overexpressed in epithelial cells of patients with IBD or animals with DSS-induced colitis. T84 monolayer also expressed PGT, which increased twofold following stimulation with TNF-α. Importantly, in T84 monolayer stimulated with TNF-α, there was a corresponding increase in the uptake and vectorial transport of 3H-PGE2 to the apical surface. Knockdown or pharmacological inhibition of PGT significantly decreased vectorial transport of 3H-PGE2. These studies unravel a mechanism whereby EP4 receptor and PGT play a role in PGE2-induced alteration of epithelial barrier integrity in colitis.

scholarly journals Polarized epithelial cells secrete matriptase as a consequence of zymogen activation and HAI-1-mediated inhibition

2009 ◽  
Vol 297 (2) ◽  
pp. C459-C470 ◽  
Author(s):  
Jehng-Kang Wang ◽  
Ming-Shyue Lee ◽  
I-Chu Tseng ◽  
Feng-Pai Chou ◽  
Ya-Wen Chen ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Seminal Fluid ◽  
Secretory Granules ◽  
Functional Divergence ◽  
The Body ◽  
In Vivo Studies ◽  
Apical Plasma Membrane ◽  
Zymogen Activation

Matriptase, a transmembrane serine protease, is broadly expressed by, and crucial for the integrity of, the epithelium. Matriptase is synthesized as a zymogen and undergoes autoactivation to become an active protease that is immediately inhibited by, and forms complexes with, hepatocyte growth factor activator inhibitor (HAI-1). To investigate where matriptase is activated and how it is secreted in vivo, we determined the expression and activation status of matriptase in seminal fluid and urine and the distribution and subcellular localization of the protease in the prostate and kidney. The in vivo studies revealed that while the latent matriptase is localized at the basolateral surface of the ductal epithelial cells of both organs, only matriptase-HAI-1 complexes and not latent matriptase are detected in the body fluids, suggesting that activation, inhibition, and transcytosis of matriptase would have to occur for the secretion of matriptase. These complicated processes involved in the in vivo secretion were also observed in polarized Caco-2 intestinal epithelial cells. The cells target latent matriptase to the basolateral plasma membrane where activation, inhibition, and secretion of matriptase appear to take place. However, a proportion of matriptase-HAI-1 complexes, but not the latent matriptase, appears to undergo transcytosis to the apical plasma membrane for secretion. When epithelial cells lose their polarity, they secrete both latent and activated matriptase. Although most epithelial cells retain very low levels of matriptase-HAI-1 complex by rapidly secreting the complex, gastric chief cells may activate matriptase and store matriptase-HAI-1 complexes in the pepsinogen-secretory granules, suggesting an intracellular activation and regulated secretion in these cells. Taken together, while zymogen activation and closely coupled HAI-1-mediated inhibition are common features for matriptase regulation, the cellular location of matriptase activation and inhibition, and the secretory route for matriptase-HAI-1 complex may vary along with the functional divergence of different epithelial cells.

Adhesion of calcium oxalate monohydrate crystals to anionic sites on the surface of renal epithelial cells

1996 ◽  
Vol 270 (1) ◽  
pp. F192-F199 ◽  
Author(s):  
J. C. Lieske ◽  
R. Leonard ◽  
H. Swift ◽  
F. G. Toback
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Calcium Oxalate ◽  
Kidney Stones ◽  
Membrane Surface ◽  
Calcium Oxalate Monohydrate ◽  
Apical Surface ◽  
Renal Epithelial Cells ◽  
Renal Tubular Cells

Adhesion of microcrystals to the apical surface of renal tubular cells could be a critical step in the formation of kidney stones. The role of membrane surface charge as a determinant of the interaction between renal epithelial cells (BSC-1 line) and the most common crystal in kidney stones, calcium oxalate monohydrate (COM), was studied in a tissue culture model system. Adhesion of COM crystals to cells was blocked by cationized ferritin. Other cations that bind to cells including cetylpyridinium chloride and polylysine, as well as cationic dyes such as Alcian blue, also inhibited adhesion of COM crystals, but not all polycations shared this effect. Specific lectins including Triticum vulgaris (wheat germ agglutinin) blocked crystal binding to the cells. Furthermore, treatment of cells with neuraminidase inhibited binding of crystals. Therefore, anionic cell surface sialic acid residues appear to function as COM crystal receptors that can be blocked by specific cations or lectins. In vivo, alterations in the structure, function, quantity, or availability of these anionic cell surface molecules could lead to crystal retention and formation of renal calculi.

312. THE DISTRIBUTION OF PROMININ-1 IN THE RAT UTERUS DURING EARLY PREGNANCY

2010 ◽  
Vol 22 (9) ◽  
pp. 112
Author(s):  
S. N. Dowland ◽  
L. A. Lindsay ◽  
C. R. Murphy
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Early Pregnancy ◽  
Cell Types ◽  
Apical Plasma Membrane ◽  
Apical Surface ◽  
Uterine Epithelial Cells ◽  
Uterine Epithelial Cell ◽  
Terminal Web ◽  
Blastocyst Implantation

Prominin-1 is a recently discovered pentaspan membrane protein present in characteristic cholesterol-based vesicles and associated with microvilli. These vesicles are used to deliver prominin-1 to the apical plasma membrane in a number of cell types. Previous work on uterine epithelial cells has demonstrated a loss of microvilli and the presence of large, cholesterol-based vesicles at the time of implantation. Thus this study aims to determine a role for prominin-1 in rat uterine epithelial cells during early pregnancy. Immunofluorescence microscopy reveals punctate and diffuse prominin-1 staining below the apical plasma membrane on day 1 of pregnancy. At the time of blastocyst implantation (day 6) however, prominin-1 appears concentrated at the apical surface of the cell. Western blotting of isolated uterine epithelial cell lysate revealed a change in prominin-1 glycosylation during early pregnancy. Prominin-1 was determined to be glycosylated on day 1 of pregnancy, but these carbohydrate side chains were lost by the time of attachment. Results seen in the present study indicate that prominin-containing vesicles may be prevented from reaching the apical plasma membrane by the terminal web on day 1 of pregnancy. On day 6, the loss of the terminal web may allow the vesicles to approach and incorporate into the apical plasma membrane, as seen with other uterine vesicles. The deglycosylation of prominin-1 at this time is suggested to allow the protein to bind its ligand and activate downstream signalling pathways that permit implantation. This study constitutes the first reported observation of prominin in endometrial lumenal epithelial cells. These preliminary results, in consideration with previous reports of prominin expression in trophoblast cells, suggest an important role for this protein in early pregnancy.

Cathepsin K in thyroid epithelial cells: sequence, localization and possible function in extracellular proteolysis of thyroglobulin

2000 ◽  
Vol 113 (24) ◽  
pp. 4487-4498 ◽  
Author(s):  
C. Tepel ◽  
D. Bromme ◽  
V. Herzog ◽  
K. Brix
Keyword(s):  
Epithelial Cells ◽  
Limited Proteolysis ◽  
Cathepsin K ◽  
Cysteine Proteases ◽  
Apical Plasma Membrane ◽  
Apical Surface ◽  
Enzyme Cytochemistry ◽  
Neutral Ph ◽  
Extracellular Proteolysis

Extracellular proteolysis of thyroglobulin at the apical surface of thyroid epithelial cells results in liberation of thyroxine, and is mediated by lysosomal cysteine proteases such as cathepsins B and L. Here, we report on the expression of the cysteine protease cathepsin K in thyroid epithelial cells. The cDNA for porcine thyroid cathepsin K showed homologies ranging from 71% to 94% to the cDNA of cathepsin K from various species and cell types. The deduced amino acid sequence of porcine thyroid cathepsin K predicted a 37 kDa preproenzyme, with the active site residues Cys-140, His-277 and Asn-297, and one potential N-glycosylation site. The localization of cathepsin K was not restricted to lysosomes. Rather, secreted cathepsin K was predominantly found within the follicular lumen and in association with the apical plasma membrane of thyroid epithelial cells. Enzyme cytochemistry showed that cell-surface associated cathepsin K was proteolytically active at neutral pH. In vitro, recombinant cathepsin K liberated thyroxine from thyroglobulin by limited proteolysis at neutral pH. We postulate that its localization enables cathepsin K to contribute to the extracellular proteolysis of thyroglobulin, i.e. thyroid hormone liberation, at the apical surface of thyroid epithelial cells in situ.

scholarly journals Epithelial Microvilli Establish an Electrostatic Barrier to Microbial Adhesion

2014 ◽  
Vol 82 (7) ◽  
pp. 2860-2871 ◽  
Author(s):  
Kaila M. Bennett ◽  
Sharon L. Walker ◽  
David D. Lo
Keyword(s):  
Epithelial Cells ◽  
Surface Charge ◽  
Electrostatic Interactions ◽  
Membrane Surface ◽  
Cell Uptake ◽  
Mucosal Barrier ◽  
M Cells ◽  
Apical Surface ◽  
Microbial Adhesion ◽  
Electrostatic Charges

ABSTRACTMicrovilli are membrane extensions on the apical surface of polarized epithelia, such as intestinal enterocytes and tubule and duct epithelia. One notable exception in mucosal epithelia is M cells, which are specialized for capturing luminal microbial particles; M cells display a unique apical membrane lacking microvilli. Based on studies of M cell uptake under different ionic conditions, we hypothesized that microvilli may augment the mucosal barrier by providing an increased surface charge density from the increased membrane surface and associated glycoproteins. Thus, electrostatic charges may repel microbes from epithelial cells bearing microvilli, while M cells are more susceptible to microbial adhesion. To test the role of microvilli in bacterial adhesion and uptake, we developed polarized intestinal epithelial cells with reduced microvilli (“microvillus-minus,” or MVM) but retaining normal tight junctions. When tested for interactions with microbial particles in suspension, MVM cells showed greatly enhanced adhesion and uptake of particles compared to microvillus-positive cells. This preference showed a linear relationship to bacterial surface charge, suggesting that microvilli resist binding of microbes by using electrostatic repulsion. Moreover, this predicts that pathogen modification of electrostatic forces may contribute directly to virulence. Accordingly, the effacement effector protein Tir from enterohemorrhagicEscherichia coliO157:H7 expressed in epithelial cells induced a loss of microvilli with consequent enhanced microbial binding. These results provide a new context for microvillus function in the host-pathogen relationship, based on electrostatic interactions.

scholarly journals Bacterial Exposure Induces and Activates Matrilysin in Mucosal Epithelial Cells

2000 ◽  
Vol 148 (6) ◽  
pp. 1305-1315 ◽  
Author(s):  
Yolanda S. López-Boado ◽  
Carole L. Wilson ◽  
Lora V. Hooper ◽  
Jeffrey I. Gordon ◽  
Scott J. Hultgren ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Acute Infection ◽  
Human Colon ◽  
Cell Types ◽  
The Body ◽  
Apical Surface ◽  
Tracheal Epithelial Cells ◽  
Lung Carcinoma Cells ◽  
Tracheal Epithelial ◽  
And Function

Matrilysin, a matrix metalloproteinase, is expressed and secreted lumenally by intact mucosal and glandular epithelia throughout the body, suggesting that its regulation and function are shared among tissues. Because matrilysin is produced in Paneth cells of the murine small intestine, where it participates in innate host defense by activation of prodefensins, we speculated that its expression would be influenced by bacterial exposure. Indeed, acute infection (10–90 min) of human colon, bladder, and lung carcinoma cells, primary human tracheal epithelial cells, and human tracheal explants with type 1–piliated Escherichia coli mediated a marked (25–50-fold) and sustained (>24 h) induction of matrilysin production. In addition, bacterial infection resulted in activation of the zymogen form of the enzyme, which was selectively released at the apical surface. Induction of matrilysin was mediated by a soluble, non-LPS bacterial factor and correlated with the release of defensin-like bacteriocidal activity. Bacteria did not induce matrilysin in other cell types, and expression of other metalloproteinases by epithelial cells was not affected by bacteria. Matrilysin was not detected in germ-free mice, but the enzyme was induced after colonization with Bacteroides thetaiotaomicron. These findings indicate that bacterial exposure is a potent and physiologically relevant signal regulating matrilysin expression in epithelial cells.

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