scholarly journals Morphology, behavior, and interaction of cultured epithelial cells after the antibody-induced disruption of keratin filament organization.

1983 ◽  
Vol 96 (2) ◽  
pp. 494-509 ◽  
Author(s):  
M W Klymkowsky ◽  
R H Miller ◽  
E B Lane
Keyword(s):  
Epithelial Cells ◽  
Cultured Cells ◽  
Mitotic Rate ◽  
Microtubule Organization ◽  
Adhesion Junctions ◽  
Cultured Epithelial Cells ◽  
Filament Bundles ◽  
Keratin Filament ◽  
Epithelial Sheets ◽  
Filament Network

The organization of intermediate filaments in cultured epithelial cells was rapidly and radically affected by intracellularly injected monoclonal antikeratin filament antibodies. Different antibodies had different effects, ranging from an apparent splaying apart of keratin filament bundles to the complete disruption of the keratin filament network. Antibodies were detectable within cells for more than four days after injection. The antibody-induced disruption of keratin filament organization had no light-microscopically discernible effect on microfilament or microtubule organization, cellular morphology, mitosis, the integrity of epithelial sheets, mitotic rate, or cellular reintegration after mitosis. Cell-to-cell adhesion junctions survived keratin filament disruption. However, antibody injected into a keratinocyte-derived cell line, rich in desmosomes, brought on a superfasciculation of keratin filament bundles, which appeared to pull desmosomal junctions together, suggesting that desmosomes can move in the plane of the plasma membrane and may only be 'fixed' by their anchoring to the cytoplasmic filament network. Our observations suggest that keratin filaments are not involved in the establishment or maintenance of cell shape in cultured cells.

scholarly journals Mouse differentiation-specific keratins 1 and 10 require a preexisting keratin scaffold to form a filament network.

1993 ◽  
Vol 120 (5) ◽  
pp. 1251-1261 ◽  
Author(s):  
T Kartasova ◽  
D R Roop ◽  
K A Holbrook ◽  
S H Yuspa
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filament ◽  
Filament Formation ◽  
Stable Transfectants ◽  
Cytoskeletal Network ◽  
Filament Bundles ◽  
Keratin Filament ◽  
Differentiation Stage ◽  
Nih 3T3 ◽  
Filament Network

Keratins 1 (K1) and 10 (K10) are the predominant cytoskeletal intermediate filaments of epidermal cells during transition from the proliferative to the terminal differentiation stage. In situ, formation of the K1/K10 intermediate filament network occurs in the cytoplasm of cells with a preexisting cytoskeleton composed of keratins 5 and 14. To define cytoskeletal interactions permissive for formation of the K1/K10 filamentous network, active copies of mouse K1 and K10 genes were introduced into fibroblasts (NIH 3T3) which do not normally express these proteins. Transient and stable transfectants, as well as heterokaryons produced by fusions with epithelial cells, were evaluated for expression of K1 and K10 proteins and filament formation using specific antibodies. In contrast to keratin pairs K5/K14 and K8/K18, the K1/K10 pair failed to form an extensive keratin filament network on its own, although small isolated dense K1/K10 filament bundles were observed throughout the cytoplasm by EM. K1 and K10 filaments integrated only into the preexisting K5/K14 network upon fusion of the NIH 3T3 (K1/K10) cells with epithelial cells expressing endogenous K5/K14 or with NIH 3T3 cells which were transfected with active copies of the K5 and K14 genes. When combinations of active recombinant gene constructs for keratins 1, 5, 10, and 14 were tested in transient NIH 3T3 transfections, the most intact cytokeratin network observed by immunofluorescence was formed by the K5/K14 pair. The K1/K14 pair was capable of forming a cytoskeletal network, but the network was poorly developed, and usually perinuclear. Transfection of K10 in combination with K5 or K1 resulted in cytoplasmic agglomerates, but not a cytoskeleton. These results suggest that the formation of the suprabasal cytoskeleton in epidermis is dependent on the preexisting basal cell intermediate filament network. Furthermore, restrictions on filament formation appear to be more stringent for K10 than for K1.

Cultured epithelial cells derived from human foetal pancreas as a model for the study of cystic fibrosis: further analyses on the origins and nature of the cell types

1988 ◽  
Vol 90 (1) ◽  
pp. 73-77
Author(s):  
A. Harris ◽  
L. Coleman
Keyword(s):  
Cystic Fibrosis ◽  
Tissue Culture ◽  
Epithelial Cells ◽  
Culture System ◽  
Cultured Cells ◽  
Cell Types ◽  
Cultured Epithelial Cells ◽  
Different Cell Types ◽  
Foetal Pancreas

The establishment of a tissue-culture system for epithelial cells derived from human foetal pancreas has recently been reported. Further analyses have now been made on these cells in vitro, together with parallel investigation of the distribution of different cell types within the intact foetal pancreas. Results support the view that the cultured cells are ductal in origin and nature. Pancreatic epithelial cell cultures have also been established from foetuses with cystic fibrosis.

scholarly journals Signals from the spindle midzone are required for the stimulation of cytokinesis in cultured epithelial cells.

1996 ◽  
Vol 7 (2) ◽  
pp. 225-232 ◽  
Author(s):  
L G Cao ◽  
Y L Wang
Keyword(s):  
Epithelial Cells ◽  
Mitotic Spindle ◽  
Actin Filaments ◽  
Cultured Cells ◽  
Critical Role ◽  
Equatorial Region ◽  
Culture Cells ◽  
Cultured Epithelial Cells ◽  
Early Anaphase ◽  
Spindle Midzone

The interaction between the mitotic spindle and the cellular cortex is thought to play a critical role in stimulating cell cleavage. However, little is understood about the nature of such interactions, particularly in tissue culture cells. We have investigated the role of the spindle midzone in signaling cytokinesis by creating a barrier in cultured epithelial cells with a blunted needle, to block signals that may emanate from this region. When the barrier was created during metaphase or early anaphase, cleavage took place only on the sides of the cortex facing the mitotic spindle. Microtubules on the cleaving side showed organization typical of that in normal dividing cells. On the noncleaving side, most microtubules passed from one side of the equator into the other without any apparent organization, and actin filaments failed to organize in the equatorial region. When the barrier was created after the first minute of anaphase, cells showed successful cytokinesis, with normal organization of microtubules and actin filaments on both sides of the barrier. Our study suggests that transient signals from the midzone of early anaphase spindles are required for equatorial contraction in cultured cells and that such signaling may involve the organization of microtubules near the equator.

Altered phenotype of cultured urothelial and other stratified epithelial cells: implications for wound healing

2006 ◽  
Vol 291 (1) ◽  
pp. F9-F21 ◽  
Author(s):  
Tung-Tien Sun
Keyword(s):  
Epithelial Cells ◽  
Wound Repair ◽  
Cultured Cells ◽  
Normal Epithelium ◽  
Corneal Epithelial Cells ◽  
Tissue Specific ◽  
Corneal Epithelial ◽  
Cultured Epithelial Cells ◽  
Normal Differentiation

The differentiation of cultured stratified epithelial cells can deviate significantly from that of normal epithelium, leading to suggestions that cultured cells undergo abnormal differentiation, or a truncated differentiation. Thus cultured epidermal and corneal epithelial cells stop synthesizing their tissue-specific keratin pair K1/K10 and K3/K12, respectively. The replacement of these keratins in the suprabasal compartment by K6/K16 keratins that are made by all stratified squamous epithelia during hyperplasia rules out a truncated differentiation. Importantly, the keratin pattern of in vivo corneal epithelium undergoing wound repair mimics that of cultured rabbit corneal epithelial cells. Although cultured urothelial cells continue to synthesize uroplakins, which normally form two-dimensional crystalline urothelial plaques covering almost the entire apical urothelial surface, these proteins do not assemble into crystals in cultured cells. Cultured epithelial cells can, however, rapidly regain normal differentiation on the removal of mitogenic stimuli, the use of a suitable extracellular matrix, or the transplantation of the cells to an in vivo, nonmitogenic environment. These data suggest that cultured epithelial cells adopt altered differentiation patterns mimicking in vivo regenerating or hyperplastic epithelia. Blocking the synthesis of tissue-specific differentiation products, such as the K1 and K10 keratins designed to form extensive disulfide cross-links in cornified cells, or the assembly of uroplakin plaques allows epithelial cells to better migrate and proliferate, activities that are of overriding importance during wound repair. Cultured urothelial and other stratified epithelial cells provide excellent models for studying the regulation of the synthesis and assembly of differentiation products, a key cellular process during epithelial wound repair.

scholarly journals p38 MAPK-dependent shaping of the keratin cytoskeleton in cultured cells

2007 ◽  
Vol 177 (5) ◽  
pp. 795-807 ◽  
Author(s):  
Stefan Wöll ◽  
Reinhard Windoffer ◽  
Rudolf E. Leube
Keyword(s):  
P38 Mapk ◽  
Intermediate Filament ◽  
Cultured Cells ◽  
Physiological Stress ◽  
Tissue Homeostasis ◽  
Epithelial Tissue ◽  
Down Regulation ◽  
Granule Formation ◽  
Keratin Filament ◽  
Filament Network

Plasticity of the resilient keratin intermediate filament cytoskeleton is an important prerequisite for epithelial tissue homeostasis. Here, the contribution of stress-activated p38 MAPK to keratin network organization was examined in cultured cells. It was observed that phosphorylated p38 colocalized with keratin granules that were rapidly formed in response to orthovanadate. The same p38p recruitment was noted during mitosis, in various stress situations and in cells producing mutant keratins. In all these situations keratin 8 became phosphorylated on S73, a well-known p38 target site. To demonstrate that p38-dependent keratin phosphorylation determines keratin organization, p38 activity was pharmacologically and genetically modulated: up-regulation induced keratin granule formation, whereas down-regulation prevented keratin filament network disassembly. Furthermore, transient p38 inhibition also inhibited keratin filament precursor formation and mutant keratin granule dissolution. Collectively, the rapid and reversible effects of p38 activity on keratin phosphorylation and organization in diverse physiological, stress, and pathological situations identify p38-dependent signalling as a major intermediate filament–regulating pathway.

scholarly journals The keratin-binding protein Albatross regulates polarization of epithelial cells

2008 ◽  
Vol 183 (1) ◽  
pp. 19-28 ◽  
Author(s):  
Masahiko Sugimoto ◽  
Akihito Inoko ◽  
Takashi Shiromizu ◽  
Masanori Nakayama ◽  
Peng Zou ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Binding Protein ◽  
Cell Polarization ◽  
Junctional Complex ◽  
Apical Surface ◽  
Apical Side ◽  
Apical Junctional Complex ◽  
Keratin Filaments ◽  
Keratin Filament ◽  
Filament Network

The keratin intermediate filament network is abundant in epithelial cells, but its function in the establishment and maintenance of cell polarity is unclear. Here, we show that Albatross complexes with Par3 to regulate formation of the apical junctional complex (AJC) and maintain lateral membrane identity. In nonpolarized epithelial cells, Albatross localizes with keratin filaments, whereas in polarized epithelial cells, Albatross is primarily localized in the vicinity of the AJC. Knockdown of Albatross in polarized cells causes a disappearance of key components of the AJC at cell–cell borders and keratin filament reorganization. Lateral proteins E-cadherin and desmoglein 2 were mislocalized even on the apical side. Although Albatross promotes localization of Par3 to the AJC, Par3 and ezrin are still retained at the apical surface in Albatross knockdown cells, which retain intact microvilli. Analysis of keratin-deficient epithelial cells revealed that keratins are required to stabilize the Albatross protein, thus promoting the formation of AJC. We propose that keratins and the keratin-binding protein Albatross are important for epithelial cell polarization.

Improved micro-impedance spectroscopy to determine cell barrier properties

2020 ◽  
Vol 4 (3) ◽  
pp. 150-155 ◽  
Author(s):  
Md. Mehadi Hasan Sohag ◽  
Olivier Nicoud ◽  
Racha Amine ◽  
Abir Khalil-Mgharbel ◽  
Jean-Pierre Alcaraz ◽  
...  
Keyword(s):  
Impedance Spectroscopy ◽  
Epithelial Cells ◽  
Lipid Bilayers ◽  
Cultured Cells ◽  
Cell Model ◽  
Measurement Techniques ◽  
Cell Monolayer ◽  
Microfluidic Channel ◽  
Small Surface ◽  
Resistance Pathway

AbstractThe goal of this study was to determine whether the Tethapod system, which was designed to determine the impedance properties of lipid bilayers, could be used for cell culture in order to utilise micro-impedance spectroscopy to examine further biological applications. To that purpose we have used normal epithelial cells from kidney (RPTEC) and a kidney cancer cell model (786-O). We demonstrate that the Tethapod system is compatible with the culture of 10,000 cells seeded to grow on a small area gold measurement electrode for several days without affecting the cell viability. Furthermore, the range of frequencies for EIS measurements were tuned to examine easily the characteristics of the cell monolayer. We demonstrate significant differences in the paracellular resistance pathway between normal and cancer kidney epithelial cells. Thus, we conclude that this device has advantages for the study of cultured cells that include (i) the configuration of measurement and reference electrodes across a microfluidic channel, and (ii) the small surface area of 6 parallel measurement electrodes (2.1 mm2) integrated in a microfluidic system. These characteristics might improve micro-impedance spectroscopy measurement techniques to provide a simple tool for further studies in the field of the patho-physiology of biological barriers.

Vimentin organizing centre in cultured epithelial cells

Pathology ◽  
1984 ◽  
Vol 16 (4) ◽  
pp. 393-395 ◽  
Author(s):  
John S. Pedersen ◽  
J.R. Underwood ◽  
B.H. Toh
Keyword(s):  
Epithelial Cells ◽  
Cultured Epithelial Cells

Stimulation of both human bronchial epithelium and neutrophils is needed for maximal interactive adhesion

1996 ◽  
Vol 270 (1) ◽  
pp. L80-L87 ◽  
Author(s):  
P. G. Bloemen ◽  
M. C. Van den Tweel ◽  
P. A. Henricks ◽  
F. Engels ◽  
M. J. Van de Velde ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cultured Cells ◽  
Bronchial Epithelium ◽  
Bronchial Epithelial Cells ◽  
Intercellular Adhesion Molecule ◽  
Tnf Alpha ◽  
Epithelial Cell Line ◽  
Ifn Gamma ◽  
Bronchial Epithelial ◽  
Stimulation Of

It has become clear that the bronchial epithelium is not just a passive barrier but plays an active role in inflammation. It can produce several inflammatory mediators and does express cell adhesion molecules of which intercellular adhesion molecule (ICAM)-1 can be upregulated by cytokines like interferon (IFN)-gamma. In the present study, we analyzed in detail the interaction of neutrophils with human bronchial epithelial cells, both primary cultured cells and the bronchial epithelial cell line BEAS-2B. Confluent monolayers of epithelial cells were incubated with freshly isolated 51Cr-labeled neutrophils for 30 min at 37 degrees C; then the nonadherent cells were removed by washing gently. Stimulation of the epithelial cells with IFN-gamma or the combination of IFN-gamma and tumor necrosis factor-alpha (TNF-alpha) (which doubles the ICAM-1 expression) increased neutrophil adhesion. Activation of the neutrophils themselves with N-formylmethionyl-leucyl-phenylalanine (fMLP), platelet-activating factor, or TNF-alpha also caused a profound enhancement of the adhesion. A significant additional increase was found when the epithelial cells had been exposed to IFN-gamma and the neutrophils were stimulated with fMLP simultaneously. This effect was even more pronounced with epithelium preincubated with IFN-gamma and TNF-alpha. With the use of monoclonal antibodies against CD18 and ICAM-1, it was demonstrated that the increased adhesion was mainly mediated by the ICAM-1/beta 2-integrin interaction. This study highlights that both the activation state of the bronchial epithelial cells and the activation state of the neutrophils are critical for their interactive adhesion.

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