Human colonic epithelial cells detect and respond to C5a via apically expressed C5aR through the ERK pathway

2012 ◽  
Vol 302 (12) ◽  
pp. C1731-C1740 ◽  
Author(s):  
Qi Cao ◽  
Shayla M. McIsaac ◽  
Andrew W. Stadnyk
Keyword(s):  
Epithelial Cells ◽  
Cell Lines ◽  
Complement Activation ◽  
Defense Mechanism ◽  
Apical Cell ◽  
Cell Types ◽  
Erk Pathway ◽  
C5a Receptor ◽  
Apical Cell Membrane ◽  
Proinflammatory Molecule

Intestinal epithelial cells (IECs) exhibit numerous adaptations to maintain barrier function as well as play sentinel roles by expressing receptors for microbial products and antimicrobial peptides. The complement system is another important innate sensing and defense mechanism of the host against bacteria and increasing evidence shows that complement plays a role in colitis. The split component C5a is a potent proinflammatory molecule, and the C5a receptor (C5aR) CD88 has been reported on multiple cell types. Here, we examined the question of whether human colonic cell lines can detect activated complement via C5aR and what signaling pathway is critical in the subsequent responses. T84, HT29, and Caco2 cell lines all possessed mRNA and protein for C5aR and the decoy receptor C5L2. Polarized cells expressed the proteins on the apical cell membrane. C5a binding to the C5aR on human IECs activates the ERK pathway, which proved critical for a subsequent upregulation of IL-8 mRNA, increased permeability of monolayers, and enhanced proliferation of the cells. The fact that human IECs are capable of detecting complement activation in the lumen via this anaphylatoxin receptor highlights the potential for IECs to detect pathogens indirectly through complement activation and be primed to amplify the host response through heightened inflammatory mediator expression to further recruit immune cells.

scholarly journals Differential Infection of Polarized Epithelial Cell Lines by Sialic Acid-Dependent and Sialic Acid-Independent Rotavirus Strains

2001 ◽  
Vol 75 (23) ◽  
pp. 11834-11850 ◽  
Author(s):  
Max Ciarlet ◽  
Sue E. Crawford ◽  
Mary K. Estes
Keyword(s):  
Sialic Acid ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Apical Cell ◽  
Paracellular Permeability ◽  
Apical Surface ◽  
Apical Cell Membrane ◽  
Basolateral Side ◽  
Epithelial Cell Lines

ABSTRACT Infection of epithelial cells by some animal rotaviruses, but not human or most animal rotaviruses, requires the presence ofN-acetylneuraminic (sialic) acid (SA) on the cell surface for efficient infectivity. To further understand how rotaviruses enter susceptible cells, six different polarized epithelial cell lines, grown on permeable filter membrane supports containing 0.4-μm pores, were infected apically or basolaterally with SA-independent or SA-dependent rotaviruses. SA-independent rotaviruses applied apically or basolaterally were capable of efficiently infecting both sides of the epithelium of all six polarized cell lines tested, while SA-dependent rotaviruses only infected efficiently through the apical surface of five of the polarized cell lines tested. Regardless of the route of virus entry, SA-dependent and SA-independent rotaviruses were released almost exclusively from the apical domain of the plasma membrane of polarized cells before monolayer disruption or cell lysis. The transepithelial electrical resistance (TER) of cells decreased at the same time, irrespective of whether infection with SA-independent rotaviruses occurred apically or basolaterally. The TER of cells infected apically with SA-dependent rotaviruses decreased earlier than that of cells infected basolaterally. Rotavirus infection decreased TER before the appearance of cytopathic effect and cell death and resulted in an increase in the paracellular permeability to [3H]inulin as a function of loss of TER. The presence of SA residues on either the apical or basolateral side was determined using a Texas Red-conjugated lectin, wheat germ agglutinin (WGA), which binds SA residues. WGA bound exclusively to SA residues on the apical surface of the cells, confirming the requirement for SA residues on the apical cell membrane for efficient infectivity of SA-dependent rotaviruses. These results indicate that the rotavirus SA-independent cellular receptor is present on both sides of the epithelium, but SA-dependent and SA-independent rotavirus strains infect polarized epithelial cells by different mechanisms, which may be relevant for pathogenesis and selection of vaccine strains. Finally, rotavirus-induced alterations of the epithelial barrier and paracellular permeability suggest that common mechanisms of pathogenesis may exist between viral and bacterial pathogens of the intestinal tract.

Bovine pancreatic duct cells express cAMP- and Ca(2+)-activated apical membrane Cl- conductances

1997 ◽  
Vol 273 (1) ◽  
pp. G204-G216 ◽  
Author(s):  
L. al-Nakkash ◽  
C. U. Cotton
Keyword(s):  
Epithelial Cells ◽  
Cell Membrane ◽  
Pancreatic Duct ◽  
Apical Membrane ◽  
Apical Cell ◽  
Primary Cultures ◽  
Apical Cell Membrane ◽  
Anion Secretion ◽  
Duct Epithelium ◽  
Cl Permeability

Secretion of salt and water by the epithelial cells that line pancreatic ducts depends on activation of apical membrane Cl- conductance. In the present study, we characterized two types of Cl- conductances present in the apical cell membrane of bovine pancreatic duct epithelial cells. Primary cultures of bovine main pancreatic duct epithelium and an immortalized cell line (BPD1) derived from primary cultures were used. Elevation of intracellular adenosine 3',5'-cyclic monophosphate (cAMP) or Ca2+ in intact monolayers of duct epithelium induced sustained anion secretion. Agonist-induced changes in plasma membrane Cl- permeability were accessed by 36 Cl- efflux, whole cell current recording, and measurements of transepithelial Cl- current across permeabilized epithelial monolayers. Elevation of intracellular cAMP elicited a sustained increase in Cl- permeability, whereas elevation of intracellular Ca2+ induced only a transient increase in Cl- permeability. Ca(2+)- but not cAMP-induced increases in Cl- permeability were abolished by preincubation of cells with the Ca2+ buffer 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, tetra(acetoxymethyl) ester (BAPTA-AM). N-phenylanthranilic acid (DPC; 1 mM) and glibenclamide (100 microM), but not 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS; 500 microM), inhibited the cAMP-induced increase in Cl- permeability. In contrast, DPC and DIDS, but not glibenclamide, inhibited the Ca(2+)-induced increase in Cl- permeability. We conclude from these experiments that bovine pancreatic duct epithelial cells express at least two types of Cl- channels, cAMP and Ca2+ activated, in the apical cell membrane. Because the Ca(2+)-activated increase in Cl- permeability is transient, the extent to which this pathway contributes to sustained anion secretion by the ductal epithelium remains to be determined.

Electroneutral H+ secretion in distal tubule of Amphiuma

1987 ◽  
Vol 252 (4) ◽  
pp. F691-F699 ◽  
Author(s):  
B. Stanton ◽  
A. Omerovic ◽  
B. Koeppen ◽  
G. Giebisch
Keyword(s):  
Basolateral Membrane ◽  
Apical Cell ◽  
Distal Tubule ◽  
Cell Types ◽  
Disulfonic Acid ◽  
Type I ◽  
Cellular Mechanisms ◽  
Apical Cell Membrane ◽  
Membrane Type

This study examines the cellular mechanisms of acid secretion by the in vitro perfused late distal tubule of Amphiuma kidney. Acidification of tubule fluid occurred against an electrochemical gradient of 16 mV; thus H+ secretion was active. Amiloride (1 mM) or a reduction of sodium in the perfusion fluid (from 83.7 to 7.7 mM) partially reduced acidification. Amiloride, in the presence of low sodium, completely inhibited acidification. Furthermore, acetazolamide and ouabain in the bath solution (0.1 mM) also inhibited acidification. Conductive properties of the epithelium and of individual cell membranes were determined by means of cable analysis of the tubule and intracellular voltage recordings. The transepithelial voltage and resistance averaged -0.4 +/- 0.4 mV, lumen negative, and 7,147 +/- 845 omega X cm, respectively. Two functionally different cell types were identified by intracellular microelectrodes. Type I cells had a basolateral membrane voltage (Vbl) of -67.7 mV. As determined by ion substitution experiments, the basolateral membrane was conductive to K+ and Cl-. This cell also had a 4-acetamido-4'-isothiocyanostilbene-2-2'-disulfonic acid (SITS)-sensitive Na+-dependent HCO3- exit pathway in the basolateral membrane. Type II cells had a Vbl of -76.1 mV (P less than 0.05 vs. type I) and the basolateral membrane was conductive to K+ and Cl- but not to HCO3-. HCO3- movement across the basolateral membrane in this cell may occur by electroneutral Cl- -HCO3- exchange. The apical cell membrane of both cell types did not contain measurable ionic conductances, as evidenced by a high value of apical membrane fractional resistance (0.98 +/- 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Neutrophil interaction with influenza-infected epithelial cells

Blood ◽  
1988 ◽  
Vol 72 (1) ◽  
pp. 142-149 ◽  
Author(s):  
DR Ratcliffe ◽  
SL Nolin ◽  
EB Cramer
Keyword(s):  
Influenza Virus ◽  
Epithelial Cells ◽  
Cell Membrane ◽  
Mdck Cells ◽  
Apical Cell ◽  
Apical Cell Membrane ◽  
Protein Insertion ◽  
Neutrophil Adherence ◽  
Vitro Model

Abstract An in vitro model system was used to study the early neutrophil response to influenza-infected epithelia. In the absence of serum, neutrophil adherence to influenza-infected confluent monolayers of Madin-Darby canine kidney epithelial cells (MDCK) was approximately 590 times greater than neutrophil binding to control cultures. The leukocytes bound specifically to virus-infected cells. Neutrophil adherence to influenza-infected MDCK cells was monitored during the course of one replication cycle, and binding began at a time (4.5 hours) that coincided with viral protein insertion in the apical cell membrane. Ultrastructural examination at 4.5 hours showed that greater than 90% of the neutrophils adhered to the epithelial cell membrane in the absence of budding virus and, at 6.5 hours, 100% of the neutrophils adhered to the epithelium with emerging virions. The number of neutrophils bound to influenza-infected MDCK cells was not affected by the presence or absence of calcium or magnesium but did depend on the amount of viral inoculum and on the temperature of the culture. In direct contrast to hemadsorption of RBCs, neutrophil binding to influenza-infected MDCK cells was 100% greater at 37 degrees C than at 4 degrees C. The neutrophil surface molecules that bound influenza virus appeared to become functionally polarized because the adherence of neutrophils to budding influenza virus or to a virus-coated surface inhibited the neutrophils from binding additional influenza virus to their nonadherent surface.

scholarly journals Antibodies to the major insoluble milk fat globule membrane-associated protein: specific location in apical regions of lactating epithelial cells.

1981 ◽  
Vol 89 (3) ◽  
pp. 485-494 ◽  
Author(s):  
W W Franke ◽  
H W Heid ◽  
C Grund ◽  
S Winter ◽  
C Freudenstein ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Lipid Production ◽  
Apical Cell ◽  
Cell Types ◽  
Specific Marker ◽  
Milk Lipid ◽  
Apical Surface ◽  
Milk Fat Globule ◽  
A Cell

Milk lipid globules of various species are surrounded by a membrane structure that is separated from the triglyceride core of the globule by a densely staining fuzzy coat layer of 10- to 50-nm thickness. This internal coat structure remains attached to the membrane during isolation and extraction with low- and high-salt buffers, is insoluble in nondenaturing detergents, and is enriched in an acidic glycoprotein (butyrophilin) with an apparent Mr of 67,000. Guinea pig antibodies against this protein, which show cross-reaction with the corresponding protein in some (goat) but not other (human, rat) species, have been used for localization of butyrophilin on frozen sections of various tissues from cow by immunofluorescence and electron microscopy. Significant reaction is found only in milk-secreting epithelial cells and not in other cell types of mammary gland and various epithelial tissues. In milk-secreting cells, the staining is restricted to the apical cell surface, including budding milk lipid globules, and to the periphery of the milk lipid globules contained in the alveolar lumina. These findings indicate that butyrophilin, which is constitutively secreted by surface budding in coordination with milk lipid production, is located at the apical surface and is not detected at basolateral surfaces, in endoplasmic reticulum, and in Golgi apparatus. This protein structure represents an example of a cell type-specific cytoskeletal component in a cell apex. It is suggested that this antigen provides a specific marker for the apical surface of milk-secreting cells and that butyrophilin is involved in the vectorial discharge of milk lipid globules.

scholarly journals Retargeting the Coxsackievirus and Adenovirus Receptor to the Apical Surface of Polarized Epithelial Cells Reveals the Glycocalyx as a Barrier to Adenovirus-Mediated Gene Transfer

2000 ◽  
Vol 74 (13) ◽  
pp. 6050-6057 ◽  
Author(s):  
Raymond J. Pickles ◽  
Jill A. Fahrner ◽  
JenniElizabeth M. Petrella ◽  
Richard C. Boucher ◽  
Jeffrey M. Bergelson
Keyword(s):  
Gene Transfer ◽  
Epithelial Cells ◽  
Basolateral Membrane ◽  
Apical Cell ◽  
Airway Epithelial Cells ◽  
Apical Surface ◽  
Apical Cell Membrane ◽  
Human Airway ◽  
Coxsackievirus And Adenovirus Receptor ◽  
Adenovirus Receptor

ABSTRACT Lumenal delivery of adenovirus vectors (AdV) results in inefficient gene transfer to human airway epithelium. The human coxsackievirus and adenovirus receptor (hCAR) was detected by immunofluorescence selectively at the basolateral surfaces of freshly excised human airway epithelial cells, suggesting that the absence of apical hCAR constitutes a barrier to adenovirus-mediated gene delivery in vivo. In transfected polarized Madin-Darby canine kidney cells, wild-type hCAR was expressed selectively at the basolateral membrane, whereas hCAR lacking the transmembrane and/or cytoplasmic domains was expressed on both the basolateral and apical membranes. Cells expressing apical hCAR still were not efficiently transduced by AdV applied to the apical surface. However, after the cells were treated with agents that remove components of the apical surface glycocalyx, AdV transduction occurred. These results indicate that adenovirus can infect via receptors located at the apical cell membrane but that the glycocalyx impedes interaction of AdV with apical receptors.

Deficient epithelial-fibroblast heterocellular gap junction communication can be overcome by co-culture with an intermediate cell type but not by E-cadherin transgene expression

1998 ◽  
Vol 111 (23) ◽  
pp. 3529-3539 ◽  
Author(s):  
T.L. Woodward ◽  
M.A. Sia ◽  
O.W. Blaschuk ◽  
J.D. Turner ◽  
D.W. Laird
Keyword(s):  
Epithelial Cells ◽  
Gap Junction ◽  
Cell Lines ◽  
Intercellular Communication ◽  
Cell Types ◽  
Intermediate Cell ◽  
Cell Type ◽  
Gap Junction Intercellular Communication ◽  
Intermediate Cells ◽  
E Cadherin

Epithelial, fibroblast and intermediate cell lines were employed to examine the mechanism(s) essential for heterocellular gap junction intercellular communication in vitro. These cell lines were characterized extensively for cell type based on morphology, intermediate cytoskeletal proteins, cell adhesion molecules and their associated proteins, tight junction proteins as well as functional differentiation. All cell types expressed connexin43 and were dye-coupled in homocellular culture. Epithelial and intermediate cells or fibroblasts and intermediate cells readily assembled heterocellular connexin43-positive gap junction plaques when co-cultured, while gap junction plaques in mixed cultures of epithelial cells and fibroblasts were rare. Dye microinjection studies were used to show that there was little gap junction intercellular communication between epithelial cells and fibroblasts. However, intermediate cells were able to communicate with epithelial cells and, to a lesser extent, fibroblasts and could transfer dye to both epithelial cells and fibroblasts when all three cell types were cultured together. Fibroblasts that were stably transfected with a cDNA encoding E-cadherin had a greater tendency to aggregate and exhibited a more epithelial-like phenotype but heterocellular gap junction intercellular communication with epithelial cells, which endogenously express E-cadherin, was not enhanced. These results suggest that mutual expression of E-cadherin is insufficient to stimulate gap junction formation between epithelial cells and fibroblasts. Moreover, our results also demonstrate that communication gaps between epithelial cells and fibroblasts can be bridged by intermediate cells, a process that may be important in mammary gland development, growth, differentiation and cancer.

scholarly journals Recognition of the laminin E8 cell-binding site by an integrin possessing the alpha 6 subunit is essential for epithelial polarization in developing kidney tubules.

1990 ◽  
Vol 111 (3) ◽  
pp. 1265-1273 ◽  
Author(s):  
L Sorokin ◽  
A Sonnenberg ◽  
M Aumailley ◽  
R Timpl ◽  
P Ekblom
Keyword(s):  
Epithelial Cells ◽  
Binding Site ◽  
Apical Cell ◽  
Cell Types ◽  
Metanephric Mesenchyme ◽  
Kidney Tubule ◽  
Cell Binding ◽  
Developing Kidney ◽  
A Chain ◽  
Integrin Alpha 6

It has been previously shown that A-chain and domain(E8)-specific antibodies to laminin that inhibit cell adhesion also interfere with the establishment of epithelial cell polarity during kidney tubule development (Klein, G., M. Langegger, R. Timpl, and P. Ekblom. 1988. Cell. 55:331-341). A monoclonal antibody specific for the integrin alpha 6 subunit, which selectively blocks cell binding to E8, was used to study the receptors involved. Immunofluorescence staining of embryonic kidneys and of organ cultures of metanephric mesenchyme demonstrated coappearance of the integrin alpha 6 subunit and the laminin A-chain in regions where nonpolarized mesenchymal cells convert into polarized epithelial cells. Both epitopes showed marked colocalization in basal areas of tubules, while an exclusive immunostaining for alpha 6 was observed in lateral and apical cell surfaces of the tubular epithelial cells. Organ culture studies demonstrated a consistent inhibition of kidney epithelium development by antibodies against the alpha 6 subunit. The data suggest that the recognition of E8 cell-binding site of laminin by a specific integrin is crucial for the formation of kidney tubule epithelium from undifferentiated mesenchymal stem cells. In some other cell types (endothelium, some ureter cells) an exclusive expression of alpha 6 with no apparent colocalization of laminin A-chain in the corresponding basement membrane was seen. Thus, in these cells, integrins possessing the alpha 6 subunit may bind to laminin isoforms that differ from those synthesized by developing tubules.

scholarly journals Chibby promotes ciliary vesicle formation and basal body docking during airway cell differentiation

2014 ◽  
Vol 207 (1) ◽  
pp. 123-137 ◽  
Author(s):  
Michael C. Burke ◽  
Feng-Qian Li ◽  
Benjamin Cyge ◽  
Takeshi Arashiro ◽  
Heather M. Brechbuhl ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Basal Body ◽  
Membrane Trafficking ◽  
Apical Cell ◽  
Primary Cultures ◽  
Physical Interaction ◽  
Vesicle Formation ◽  
Nucleotide Exchange ◽  
Apical Cell Membrane ◽  
Tracheal Epithelial Cells

Airway multiciliated epithelial cells play crucial roles in the mucosal defense system, but their differentiation process remains poorly understood. Mice lacking the basal body component Chibby (Cby) exhibit impaired mucociliary transport caused by defective ciliogenesis, resulting in chronic airway infection. In this paper, using primary cultures of mouse tracheal epithelial cells, we show that Cby facilitates basal body docking to the apical cell membrane through proper formation of ciliary vesicles at the distal appendage during the early stages of ciliogenesis. Cby is recruited to the distal appendages of centrioles via physical interaction with the distal appendage protein CEP164. Cby then associates with the membrane trafficking machinery component Rabin8, a guanine nucleotide exchange factor for the small guanosine triphosphatase Rab8, to promote recruitment of Rab8 and efficient assembly of ciliary vesicles. Thus, our study identifies Cby as a key regulator of ciliary vesicle formation and basal body docking during the differentiation of airway ciliated cells.

Morphology and Ultrastructure of the Dufours and Venom Gland in the Ant Myrmecia-Gulosa (Fabr) (Hymenoptera, Formicidae)

1990 ◽  
Vol 38 (3) ◽  
pp. 305 ◽  
Author(s):  
J Billen
Keyword(s):  
Endoplasmic Reticulum ◽  
Fine Structure ◽  
Epithelial Cells ◽  
Cell Membrane ◽  
Control Mechanism ◽  
Smooth Endoplasmic Reticulum ◽  
Apical Cell ◽  
Venom Gland ◽  
Apical Cell Membrane ◽  
Cuticular Layer

The morphology and fine structure of the two major sting glands in the primitive Australian bull ant, Myrmecra gulosa, are described. The cells of the glandular epithelium of the tubiform Dufour's gland are characterised by a well developed vesicular smooth endoplasmic reticulum, numerous lamellar inclusions, and microvillar differentiations of the apical cell membrane. The cells of the secretory filaments of the venom gland contain a very extensive granular endoplasmic reticulum and numerous Golgi vesicles. The highly proteinaceous secretion reaches the filament lumen through the intracellular end apparatus. Passage through the convoluted gland probably accompanies the modification or production of additional secretory components, as is suggested by the ultrastructural organisation of the convoluted gland cells. The large venom gland reservoir is lined with squamous epithelial cells and a thick cuticular layer, that protects the ant from self-toxication by the powerful venom. Each sting gland opens separately through the sting, and possesses its own muscular control mechanism that allows independent discharge of secretion.

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